Matra viruddha ss-mys

i

“CONCEPT OF SAMSKARA AND MATRA VIRUDDHATWA OF MADHU-

AN ANALYTICAL AND EXPERIMENTAL STUDY”

By

Dr. A. ANNAPOORANI, B.A.M.S.,

Dissertation submitted to the

Rajiv Gandhi University of Health Sciences, Karnataka, Bangalore

in partial fulfillment of the requirements for the degree of

DOCTOR OF MEDICINE (AYURVEDA)

in

AYURVEDA SIDDHANTA

Under the Guidance of

Dr. N. ANJANEYA MURTHY, M.D. (Ayu)

Former Professor & H.O.D,

Department of Post-Graduate Studies in Ayurveda Siddhanta

G.A.M.C, Mysore.

CO-Guides

Dr. K. VENKAT SHIVUDU, M.D. (Ayu)

H.O.D,

Department Of Basic Principles,

J. S. S. Ayurveda Medical College, Mysore.

&

Dr. ANAND KATTI, M.D. (Ayu)

Lecturer

Department of Post-Graduate Studies in Ayurveda Siddhanta

GOVERNMENT AYURVEDA MEDICAL COLLEGE

MYSORE.

2009

Ayurmitra

TAyComprehended

ii

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES, KARNATAKA

DEPARTMENT OF POST GRADUATE STUDIES IN

AYURVEDA SIDDHANTA


MYSORE.

DECLARATION

I hereby declare that this Dissertation “CONCEPT OF SAMSKARA AND MATRA

VIRUDDHATWA OF MADHU-AN ANALYTICAL AND EXPERIMENTAL

STUDY” is a bonafide and genuine research work carried out by me under the guidance

of Dr. N. Anjaneya Murthy, Former Professor & H.O.D, Department of Post Graduate

Studies in Ayurveda Siddhanta, Government Ayurveda Medical College, Mysore.

Date : Signature of the Candidate

Place : Mysore Dr. A. Annapoorani

iii



AYURVEDA SIDDHANTA


MYSORE.

CERTIFICATE BY THE GUIDE

This is to certify that the dissertation entitled “CONCEPT OF SAMSKARA AND MATRA

VIRUDDHATWA OF MADHU-AN ANALYTICAL AND EXPERIMENTAL STUDY” is a

bonafide research work done by Dr. A. Annapoorani in partial fulfillment of the

requirement for the degree of Doctor of Medicine (Ayurveda).

Date : Signature of the Guide

Place : Mysore Dr. N. Anjaneya Murthy, M.D. (Ayu)

Former Professor & H.O.D,

Department of P. G. Studies

in Ayurveda Siddhanta,

Government Ayurveda Medical

College, Mysore.

iv



AYURVEDA SIDDHANTA


MYSORE.

CERTIFICATE BY THE CO-GUIDE

This is to certify that the dissertation entitled “CONCEPT OF SAMSKARA AND

MATRA VIRUDDHATWA OF MADHU-AN ANALYTICAL AND

EXPERIMENTAL STUDY” is a bonafide research work done by Dr. A. Annapoorani

in partial fulfillment of the requirement for the degree of Doctor of Medicine

(Ayurveda).

Date : Signature of the Co-Guide

Place : Mysore Dr. K. Venkat Shivudu, M.D (Ayu)

H.O.D,

Department of Basic Principles,

J. S. S. Ayurveda Medical

College, Mysore.

v



AYURVEDA SIDDHANTA


MYSORE.

CERTIFICATE BY THE CO-GUIDE

This is to certify that the dissertation entitled “CONCEPT OF SAMSKARA AND

MATRA VIRUDDHATWA OF MADHU-AN ANALYTICAL AND

EXPERIMENTAL STUDY” is a bonafide research work done by Dr. A. Annapoorani

in partial fulfillment of the requirement for the degree of Doctor of Medicine

(Ayurveda).

Date : Signature of the Co-Guide

Place : Mysore Dr. Anand Katti, M.D (Ayu)

Lecturer,

Department of P. G. studies

In Ayurveda Siddhanta

Government Ayurveda Medical

College, Mysore.

vi



AYURVEDA SIDDHANTA


MYSORE.

ENDORSEMENT BY THE HOD, PRINCIPAL/

HEAD OF THE INSTITUTION

This is to certify that the dissertation entitled “CONCEPT OF SAMSKARA

AND MATRA VIRUDDHATWA OF MADHU-AN ANALYTICAL AND

EXPERIMENTAL STUDY” is a bonafide research work done by

Dr. A. Annapoorani under the guidance of Dr. N. Anjaneya Murthy M.D. (Ayu), Professor

& H.O.D, Department of Post Graduate Studies in Ayurveda Siddhanta, Government

Ayurveda Medical College, Mysore.

Seal & Signature of the HOD Seal & Signature of the Principal

Date : Date :

Place : Mysore Place : Mysore

vii



AYURVEDA SIDDHANTA


MYSORE.

COPY RIGHT

Declaration by the Candidate

I hereby declare that the Rajiv Gandhi University of Health Sciences, Karnataka shall

have the rights to preserve, use and disseminate this dissertation/thesis in print or

electronic format for academic/research purpose.

Date : Signature of the Candidate

Place : Mysore Dr. A. Annapoorani

© Rajiv Gandhi University of Health Sciences, Karnataka

viii

ACKNOWLEDGEMENT

At this moment of submitting this dissertation, I bow my head to the Almighty for his

blessings and all the spiritual teachers who guided me in accomplishing this study.

I express my heartfelt sense of gratitude to Dr. Shakuntala G. N. Professor and

HOD, Department of PG Studies in Ayurveda Siddhanta, GAMC, Mysore, for her constant

guidance, continuous supervision and help at every stage of this study.

I feel it an honour to express a special thanks to my guide, former HOD,

Dr. N. Anjaneya. Murthy, M.D (Ayu), Joint Director of AYUSH, Govt of Karnataka, for his

inspiration, unstinted support, continuous scrutiny, guidance and immense help at every

stage of this study.

I express my sincere thanks with accolades to Dr.KR.Anilakumar, Scientist ‘D’,

for his incessant encouragement, scholarly guidance and pin-point suggestions to carry

out the analytical and experimental study at Defence Food Research Laboratory, Mysore.

I am grateful to my co-guide Dr. K.Venkat Shivudu, HOD, Dept. of Basic

Principles, JSSAMC, Mysore, for his constant support, time to time guidance and

inspiration in carrying out the study.

I express my gratitude to my co-guide Dr.Anand Katti, Lecturer, Department of

PG studies in Ayurveda Siddhanta, GAMC, Mysore, for his immense support, time to

time guidance and encouragement during the course of this work.

I am also thankful to Dr. Ashok D. Satpute, Principal, GAMC, Mysore, for his

guidance, and support that he had provided during the period of my study.

I owe my sincere gratitude and thanks to Dr.A.S.Bawa, Director, DFRL, Mysore

and Dr. Farahat Khanum, H.O.D, Biochemistry and Nutrition Discipline, DFRL,

Mysore, who provided the valuable opportunity to carry out the part of dissertation work

in the esteemed institution.

ix

I am thankful to Dr. Srinivasa Prasad, Associate Professor, Dept. of Pathology

Mysore Medical college, for providing the histopathological report.

I am very much thankful to Vidwan Gangadhar Bhat, Professor, Tarka

Vibhaga,Maharaja Sanskrit College, Mysore who has given Sanskrit grammatical corrections

for this study.

I thank Dr. Lancy D’Souza for helping in statistical analysis and interpretation.

I wish to place my sincere gratitude to Dr. K. Naseema Akhtar, Dr. Shantala

Priyadarshini, Dr. Shrivatsa and Dr Vasudev A Chate for their inspiration and constant

guidance at every stage of my study.

I would like to place on record my deep sense of gratitude to my beloved teachers,

Dr. T. D. Ksheersagar, Dr. Chandramouli, Dr. D. L. Balakrishna, Dr. V. Rajendra,

Dr. Shantaram, Dr. Gopinath, Dr. R. C. Mythreyi and Dr. Adarsh for their inspiration

during my P.G. studies.

I would like to thank Dr. Nagamani, H.O.D, dept of Dravya Guna, Dr.P.Sudhakar

Reddy, H.O.D, dept of Swastha vrutta, Dr.Anil Kumar, Asst. Professor, dept of Kaya

chikitsa, and Dr.Satish Pai, lecturer in JSSAMC, Mysore, for their help and guidance in the

study.

I would like to acknowledge with thanks the help rendered by Dr.R.K.Sharma,

principal, JSSAMC, Mysore for permitting me to utilize the library facilities and to discuss

with the eminent scholars f the college.

I express my sincere thanks to Dr.Jayashree.M.N. M.D (Ay.Sid), and Dr. Prakash in

Dept. of Agada tantra, G.A.M.C, Trivandrum for their support and help in collection of

literary data.

x

I bow my head to my teacher Dr. S. Varalakshmi. M.D. (Ayu), for her incessant

guidance and immense support, without whom I could not have joined Post graduation

course.

I convey my special thanks to my UG lecturers Dr. Vijaya Seshadri, Dr.Srirama

Krishnan, Dr.Giridhara Krishna, Dr.Sampath, Dr. Rangaprasad Bhat, Dr. Hema,

Ms.Rama and Dr.Sriman Narayana for their encouragement and guidance.

I express my gratefulness to Dr. P.L.T.Girija and Mr.Mukundan for imparting in

me an unquenchable thirst for knowledge in Ayurveda.

I express my thanks to Dr. D. Arun kumar, Mrs. Shrimati Arun Kumar and

Sushruta for their kindness and support during the times of my frustration and for

offering valuable suggestions and inspiration. I would also like to thank

Dr.A.S.Chandrashekar for his support and guidance.

I would like to thank all my colleagues Dr. Aparna K, Dr. Triveni, Dr. Sidram

Guled, Dr. Shivanand, Dr. Chitralatha, Dr. Mahesh Hiremath, and Dr. Akarshini who

made my stay in college very joyful and educative.

I convey my thanks to my seniors Dr.H.D.Vijayalakshmi, Dr.Soubhagya,

Dr.Savita Shenoy, Dr.Kedar Sharma and Dr.Yogesh for their help during the time of

synopsis and also at the commencement of this research work.

I also convey my wishes and thanks to juniors Dr. Geetha.P, Dr.Ramesh Kumar,

Dr.Ranjith kumar Shetty, Dr. Kalyani Bhusane and Dr.Kavitha.S of Ayurveda Siddhanta

and all junior PGs of Kayachikitsa dept. for their overall help.

I express my gratitude and thanks to Ms.Saritha.V,SRF and Ms.Bhawya,

Research fellow, for extending their help and moral support to conduct the work in

Biochemistry Lab, DFRL, Mysore.

I am indebted and thankful to Mr.K. Manjunath and Smt.R.Sridevi, ‘Sri

Gurudutt pharmaceutical distributors’ and Sneha for their love, kindness and co-

operation during my stay in Mysore.

xi

I express my immense thanks to Mr. Shekhar B. S. and Smt. Mangala for their

extreme support throughout my stay at Mysore.

It is a great pleasure to express my deep sense of gratitude to my brother,

Dr.Rajesh.M. Bhat; my cousin, Dr. Aparna. J, M.D. (Paed) for their valuable suggestions

and immense support throughout my PG studies.

I express my sense of gratitude for my friends Mrs. AkilaShankar and

Dr.Alarmel Mangai for their love, kindness and support.

I am ever thankful to my parents, my uncles, aunts, my brothers, sisters and all

my family members who patiently bear my pains and supported me for the successful

completion of this work.

I extend my regards to our dept attender Annaiachar and librarians Mr. Venkata

Subbaiah, Smt. Varalakshmi, Mr. Somasundar, Mr. Basavaraju and Mr. Raju GAMC,

Mysore.

I also extend my regards to Mr. Naveen, librarian of J. S. S. Ayurveda Medical

College, Mysore for their generous help during the course of my work.

I am extremely grateful to the computer technicians Mr.Mahesh, Mr.Shrikanth

and Mr.Raju, Manish enterprises, Mysore for their technical support.

I am ever grateful to all those who have helped me directly and indirectly in

completion of this project work successfully.

Dr.A.Annapoorani

xii

ABBREVIATIONS

AH-Ashtanga Hrudaya

ALP- Alkaline Phosphatase

AS-Ashtanga Sangraha

BN- Bhavaprakasha Nighantu

BS-Bhela Samhita

CD- Conjugted Dienes

CS- Charaka Samhita

DGH- Dravyaguna hastamalaka

DN- Dhanwantara Nighantu

DPPH- Diphenyl picryl hydroxyl

E.C- Enzyme Classification

FFA-Free Fatty Acids

GGT- Gamma glutamyltranspeptidase

GSH-Px - Glutathione peroxidase

GSSG-Red - Glutathione reductase

GST- Glutathione S-transferase

G-6-PDH- Glucose-6-phosphate dehydrogenase

HMF- Hydroxy Methylfurfuraldehyde

HPR- Histopathology reading

KN-Kaiyyadeva Nighantu

MN- Madanapala Nighantu

PFA- Prevention of Food Adulteration Act

RN- Raja Nighantu

SKD- Shabda Kalpadruma

SOD- Superoxide dismutase

SS- Sushruta Samhita

TBARS - Thiobarbituric acid reactive substance

Vachas- Vachaspatyam

VSS- Vaidyaka Shabdasindhu

YR- Yogaratnakara

Viruddhatwa of Madhu – An Analytical and Experimental study

Dr. A. Annapoorani 1

ABSTRACT

Background and Objectives-

Food is a powerful source of energy for all human activities. It undergoes

deterioration due to the contradictory qualities, unsuitable combination and opposed

processing procedures thereby results in incompatibility. Honey and ghee are the two

food substances used widely in our diet. The commercial honey is heat-processed; honey

and ghee are mixed and served for food at times that absolutely contradicts the Ayurvedic

concept. Hence a detailed study has been aimed in order to analyze the physico-chemical

properties and to evaluate the toxic and deleterious effects, if any; on consumption of a)

heated honey b) honey mixed with equal amount of ghee and c) heated honey mixed with

heated ghee in animal models.

Methods-

1) Analytical study-

The following physico-chemical properties of honey and ghee samples are

studied- Ash value, Acid Insoluble ash, pH, Specific gravity, Organoleptic

characteristics, HMF, Browning index, Total phenolic compounds, Flavonoids, Total

Antioxidant activity, Peroxide value, FFA, TBA value and Saturated to unsaturated fatty

acid ratio.

2) Experimental study-

The acute and sub-chronic toxicity studies were carried on Wistar strain albino

rats. The biochemical study on serum, organ weights and histo-pathology readings were

conducted to evaluate the gross and microscopic changes. The results were analyzed

statistically.



Results- On heating HMF content, browning and antioxidant activity of honey were

found to be increased significantly. There was a significant raise in ALP, GST,

Conjugated dienes and GGT of rats fed with honey mixed ghee and heated- honey mixed

ghee. Serum total proteins showed significant increase in rats fed with heated honey,

Honey mixed ghee and heated- honey mixed ghee.

Interpretation & Conclusion

It is evident from the analytical and sub-chronic toxicity study that Ushna Madhu

(heated honey -140 C for more than 15sec) and Sama Ghruta Madhu (honey mixed ghee)

produced early signs of hepato-toxicity in Wistar strain albino rats.

Keywords-

Incompatibility, HMF, Antioxidant activity, Phenols, Flavonoids, Peroxide,

Browning, FFA, TBA, Conjugated dienes.



CONTENTS

Particulars Page no.

Acknowledgement viii

Abbreviations xii

Abstract xiii

List of Tables xvii

List of Flow charts and Figures xviii

INTRODUCTION 1-2

REVIEW OF LITERATURE 3-69

I. REVIEW ON VIRUDDHA

Historical Review 3-6

Derivation 7

Definition 7

Lakshanas 7

Synonym 8

Terms related to Viruddha ahara 9-11

Types of Viruddha and its detail review 11-26

Review on Matra Viruddha 17-18

Review on Samskara Viruddha 19-21

Effects of Viruddha ahara 26-28

Factors that nullify the effects 28

Treatment principles 29-30

Modern Review 31-34



II. REVIEW ON MADHU 35-56

Derivation 35

Definition 35

Synonyms 35-36

Regional names 37

Types of Madhu 37-45

Instructions for the intake of Madhu 45-46

Qualities of different kinds of Madhu 47

Modern Review on honey 48-56

III. REVIEW ON GHRUTA 57-69

Derivation 57

Definition 57

Synonyms 58

Regional names 59

Classification of Ghruta 59-60

Qualities of Ghruta in general 61-62

Review on Go-ghruta 63-65

Modern Review on ghee 66-69

MATERIALS AND METHODOLOGY 70-92

OBSERVATIONS 93-106

DISCUSSION 107-127

CONCLUSION 128-129

RECOMMENDATIONS FOR FURTHER STUDY 130

SUMMARY 131-132

BIBLIOGRAPHY 133-142



LIST OF TABLES

S.N Tables Page No

1. Synonyms of Viruddha 8

2. Ahita Ahara and Viruddhatwa related to it 9

3. Types of Viruddha according to different Acharyas 13

4. Viruddha aharas for different seasons 14

5. Viruddha aharas for different Agni of individuals 15

6. Matra viruddhata according to different Acharyas 18

7. Viruddha Ahara janya rogas 27

8. Synonyms of Madhu 36

9. Types of Madhu based on types of honey bees 37

10. Colour of different kinds of honey 40

11. Colour of different kinds of honey bees 40

12. Properties of different kinds of honey 47

13. Synonyms of Ghruta 58

14. General properties of Ghruta 61

15. Properties of different kinds of Ghruta 64

16. Biochemical parameters adopted to study the samples 82

17. Physical characteristics of honey and ghee samples 96

18. Physico- chemical characteristics of honey samples 98

19. Physico- Chemical characteristics of ghee samples 99

20. Effects of feeding of honey and ghee on rat SGOT, SGPT, creatinine, total proteins, albumin and globulin

100

21. Effects of feeding of honey and ghee on rat serum alkaline phosphatase, total cholesterol, triglycerides, HDL, LDL

101



22. Effects of feeding of honey and ghee on rat serum urea- B, uric acid, total bilirubin, direct bilirubin and glucose

102

23. Effects of feeding of honey and ghee on hepatic lipid peroxides 103

24. Effects of feeding of honey and ghee on detoxifying enzyme of liver 104

25. Effects of feeding honey and ghee on liver enzymes 105

26. Showing Viruddha sevanajanya Sroto dushti 114

LISTS OF FLOW CHARTS Pg No.

1. Viruddha Aharajanya Roga Chikitsa 30

2. Method of Ghee preparation 67

LISTS OF FIGURES Pg No.

1. Food intake of rats 93

2. Weight gain of rats 93

3. Effects of feeding of honey and ghee on relative weight of rat liver 93

4. Effects of feeding of honey and ghee on relative weight of rat kidney 94

5. Effects of feeding of honey and ghee on relative weight of rat heart 94

6. Effects of feeding of honey and ghee on relative weight of rat spleen 95



INTRODUCTION

When a human being is in harmony with laws of nature, no problem is unsolvable,

irrespective of how strong and disturbing the so-called symptoms. – Faw Hardy

Food is the basic requirement for man and is the source of nutrients. It provides

the energy required for all activities of a human, such as growth, repair of the damaged

tissues, reproduction and sustenance.When food is consumed according to the dietetic

laws, it bestows health. Similarly when the laws of dietetic regimens are broken it leads

to numerous diseases. Food related diseases include nutritional deficiency, food

contamination, food intolerance and also food incompatibility.

According to Ayurvedic definition, the incompatible foods i.e. Viruddha ahara are

those which causes derangement in equilibrium of the doshas and remain antagonistic to

the tissues.

Honey and ghee are the two substances used in regular foods since ancient time.

Certain precautionary measures that are emphasized for the intake of honey are not

strictly followed in present scenario. It is said that Ushnam cha Samaghrutam Madhu

marayati || i.e heated and equal quantity of ghee mixed honey causes hazardous effects

that eventually end in death. So the incompatibility involved in their usage needs a proper

insight.

Hence to re-establish the truth and to expand our knowledge by funding with

newer comments and fresh examples, this study has been undertaken.



RELEVANCE OF THE STUDY

Life on the earth is undergoing change at each & every fraction of time. The

scientific era is demanding evidences for the classical concepts. Viruddha ahara/

Incompatible foods remain as a silent cause for many diseases; hence a rational

assessment is made to understand the concept in newer dimension.

OBJECTIVES OF THE STUDY

1. To review and compile the literature on Madhu.

2. To study in detail about the Samskara and Matra Viruddhatwa of Madhu.

3. To analyze the physico-chemical properties of unprocessed, processed and

heated honey and also equal quantity of ghee mixed honey.

4. To assess the probable toxicity on Wister Strain albino rats after administering

the unprocessed, heated and equal quantity of ghee mixed honey.



REVIEW OF LITERATURE

Introduction-

Food is an indispensable part of our life. Our wrong notions regarding it have

made Viruddha Aharas, a part and parcel of our life. The effect of Viruddha Ahara is

slow and spread over a long time that we fail to notice its effects in the body. The basic

treatment principle of Ayurveda is identification and removal of cause and curtailing the

progression of disease. As it is rightly quoted that “Ahara sambhavam vastu Rogastu

Ahara sambhavaha ||” The man made out of food also suffers from the disease due to the

derangement in the dietic patterns. Hence to identify the cause of a disease, knowledge of

Viruddha Ahara is most essential that explains about the Viruddhas originating from

Samyoga, Samskara, Kala, Desha, Matra etc., Identification of a particular Viruddha in

dietic habits of a patient helps the physician to give better treatment.

Historical Review-

A clear idea about the concept of Viruddha in different time periods and

according to different Acharyas explains about the relevance and importance of it in the

society.The prevalence of concept of Viruddha in different kala (time period) has been

given in detail as follows-

There are no direct references of Viruddha ahara in the Veda, Upanishads and

Purana kala. But the concepts of Annarakshana vidhi, Ahara upayogi vidhi which

indirectly refer to the avoidance of Krama Viruddha, Vidhi Viruddha, Sampat Viruddha

etc., are seen in those days. The concept was well established only during Samhita kala,

the details of which are given below-



Samhita kala-

Charaka Samhita- Acharya Charaka has explained the concept of Viruddha in

Sutrasthana-Atreyabhadrakapyeeya Adhyaya, after the explaination of

Rasapanchaka.Under the heading Viruddha Ahara, Acharya mentioned the

definition, different factors responsible for Viruddha, types of Viruddha with a

brief explanation and illustrations, the effects i.e diseases that are produced by

Viruddhanna sevana, treatment principles to be adopted and also the factors that

nullify the effect of Viruddha sevana.1 While explaining the concept of

Madhvaama, Upachara Viruddha has been explained.2

Sushruta Samhita- Acharya Sushruta explained the concept of Viruddha in

Sutrasthana- Hitaahiteeya adhyaya. In which he explained the three types of diet

regimens Viz., Atyanta hita, Atyanta ahita and Hitahita. Acharya is of the opinion

that no substance in the world can be absolutely hita or ahita and it is the Vaidya

who decides according to the svasthyata of the individual. After this, there is a

direct explanation of few types of Viruddha, its harmful effects, prophylaxis and

counteraction.3

Bhela Samhita- In Sutrasthana- Atreyakhandakapyeeya adhyaaya, Acharya Bhela

explained the rasa varga, after which the explanation of different Viruddha ahara

is seen. Here the specific diseases that arise out of certain types of Viruddha have

been mentioned. In another context Acharya emphasizes to prohibit Viruddha

anna sevana in samashanaparighaneeya adhyaya which is the previous chapter.4

Kashyapa Samhita- In Kalpasthana- Bhojanakalpa Adhyaya, while answering the

disciples for beneficial manner of dietic regimen, effects of excess and less food



intake etc., Acharya has explained the concept of Ahita sevana and its effects.

While quoting he gave due importance for the Agni and mentioned that the person

eating for a long time, repeatedly, irregularly, excessively and contrary foods

(Viruddhahara) etc., will develop Agnimandya that results in many diseases like

Jwara etc.5

Sangrahakala-

Ashtanga Sangraha- Acharya Vagbhatta has dedicated a chapter on Viruddha, i.e.,

Viruddhanna vijnaniya adhyaya in Sutrasthana. Here which he explained

definition, different kinds of Viruddha ahara, bad effects of it, treatment and the

factors that nullify the effects. He also mentioned that the Viruddha ahara can act

like a Pathya i.e medicine in certain conditions. For example, Visha in Udara.6

Ashtanga Hrudaya- In Sutrasthana- Annaraksha adhyayathe concept of Viruddha

ahara has been dealt in detail.7

Adhunika kala-

Some of the text books like Bhavaprakasha, Yogaratnakara and Bhaishajya

ratnavali has explained Viruddhashana in the context of nidana for various

diseases like jwara, Krimi etc., Anandakanda have mentioned the concept of

Viruddha in brief.8

According to modern parlance, the concept of food incompatibility, food

intolerance and hypersensitivity reactions to certain foods etc., can be matched

with the concept of Viruddha.



Previous Research works-

Shobhana.M.C- A study on the role of Viruddha ahara in the occurrence of

diseases-Dept. of Swasthavrutta- Trivandrum-2002

Inderkumar Parvani- A conceptual and applied study of Viruddha ahara with

particular reference to Eka Kushta- Dept. of Basic Principles- IPGT&R,

Jamnagar-2003

Aparna Deshpande- A study of Viruddha Ahara w.s.r. to Samyog Viruddha and

Virya Viruddha Ahara- Dept. of Ayurveda Siddhanta- Pune- 2004.

Rasmi.P.R.- Experimental evaluation of toxic effect of Ghruta and Madhu in

equal quantity on oral administration w.s.r. to Matra Viruddha concept in

Ayurveda- Dept. of Agada Tantra- Trivandrum-2007



Derivation-

“Vi + Rudh+ Kta- Virodha Vishishta”|| (SKD)

The term Viruddha is formed from the the root ‘Rudhir’ ‘Avarane’ added with a

prefix ‘Vi’ and a suffix ‘Kta’ which means “Virodha Vishesha” i.e. contrast or opposition

in particular.

Definition-

Virodha yukte | (Vachas)

That which resists or hinders is known as Viruddha.

Viruddham- Deshakaalaprakruti samyoga vipareete | (V.S.S)

Act in contrary to the place, time, nature and combination is Viruddha.9

Lakshanas-

That which acts antagonistic to the deha dhatus (tissues of the body) is known as

Viruddha.10

The substances that cause vitiation of Doshas in the body but do not expel them

are known as Viruddha and so they remain antagonistic to the dhatus.11

The drugs and diets that dislodge the doshas but do not expel them from the body

are regarded as unwholesome.12

Drugs and diets which are unwholesome for the normal dhatus and doshas of the

body, are infact opposed to their proper growth wherein some act due to their

mutually contradictory qualities; some by combination; some by the method of



preparation; some by the virtue of place, time and dose and some others by their

inherent nature.11

No substances in this world can entirely be wholesome or unwholesome. Because,

a substance which is compatible for Vata dosha will be antagonistic to Pitta

dosha. The articles in their natural form or in combinations may be either of these

three- 1. Atyanta Hita (Always beneficial) Eg- Jala, Ghruta, Ksheera etc., being

congenial by nature is always beneficial to human beings. 2. Atyanta Ahita

(Always non- beneficial) Eg- Agni, Kshara and Visha that does Dahana, Pachana

and Marana respectively, are always non- beneficial 3. Hitaahita (Mixed values)

some other substances though wholesome may become equivalent to poisons,

when used in various combinations.13

Table.1. Synonyms of Viruddha14

S.N Synonyms Meanings

1 ÌuÉÂ®: Incompatible, Antagonistic

2 ÌuÉmÉUÏiÉ: Adverse or acting contrary

3 ÌuÉsÉ¤ÉhÉ: Act of distinguishing

4 ÌuÉxÉ…¡ûiÉ: Peculiar to the group

5 ArÉÉåarÉ: Unsuitable

6 AxÉ…¡ûiÉ: Unpreferred

7 AxÉÇpÉÉurÉ: Incomprehensible

8 AxÉÇpÉuÉ: Impropreity

9 AxÉÇÎxjÉiÉ: Wrongcombination/ Inconsistence

10 AlÉÑmÉrÉÑ£ü: Unsuitable for use

11 mÉUxmÉU ÌuÉÂ®: Mutually contradictory

12 MÉëirÉlÉÏMü Antagonistic



Various English equivalents for the term ‘Viruddha’-

Opposed, Hindered, Incompatible, Inconsistent with, Contrary, Contradictory,

Adverse, Unfavourable, Prohibited.

Terms related to Viruddhahara-

Some of the classical terminologies can be considered under the concept of

Viruddha ahara. They are-

Ahita Ahara (Unwholesome food) - Acharyas has listed down some of the food

articles which are unwholesome by nature and it has to be avoided by the people.

Eg-Yavaka, Masha etc., which can be considered as Swabhava Viruddha.

Table. 2. Ahita Ahara and Viruddhatwa related to it

S.N Type of Food articles Ahita Ahara Viruddhatva

1 Shooka dhanya Yavaka Swabhava

2 Shami dhanya Masha Guna

3 Jalavarga River water in rainy season Kala

4 Lavana Ushara lavana Desha

5 Shaka Sarshapa Swabhava

6 Mruga mamsa Go mamsa Swabhava

7 Pakshi mamsa Kapota mamsa Swabhava

8 Matsya Chilichima Swabhava

9 Meats of animals living in hole

(Bhileshaya)

Frog meat Swabhava

10 Ghruta Avikam sarpi Swabhava

11 Ksheeram Avikam Swabhava

12 Vegetable fats (Sthavara sneha) Kusumbha sneha Swabhava

13 Fats of Marshy land (Anupa vasa) Mahisha vasa Swabhava



14 Fats of aquatic animals (Matsya

vasa)

Kumbheeravasa (crocodile

fat)

Swabhava

15 Fats of aquatic animals (Jalachara

vihanga vasa)

Kakamadgu (fat of water

foul)

Swabhava

16 Fats of gallinecious birds (Vishkira

shakuni vasa)

Chataka (sparrow) Swabhava

17 Fats of herbivorous animals

(Shakada medas)

Elephant fat Swabhava

18 Fruits (Phala) Nikucha Swabhava

19 Rhizomes and roots (Khanda) Aluka Swabhava

20 Sugarcane preparations (Ikshu

Vikara)

Phanita Samskara

Mityahara (Improper food)- Intake of diet abiding the rules i.e ‘Ashtaaharavidhi

Vishesha ayatanas’ is regarded as Mitya Ahara sevana. The eight factors that

should be considered for beneficial effects of food are- Prakruti, Karana,

Samyoga, Rashi, Kala, Upayoga Samsta and Upabhokta. Habitual intake of food

in a proper way will be more useful but food taken in improper manner is always

harmful. So it should be avoided.

Samashana-Intake of wholesome and unwholesome food at a time in a meal is

Samashana. Eg- Raktashali + Yavakanna

Adhyashana- Intake of food before the digestion of previous meal is Adhyashana.

If proper time is not given for digestion of previous meal and stomach still

occupied with previous food, the food taken in such conditions mixes with the

partially digested food produces Amavisha. There will be disturbance in the

secretion of gastric juices and peristalsis.



Vishamashana- Taking food in improper quantity (too much or too little), at

irregular times (either too early or too late) is Vishamashana. Above three types-

Samashana, Adhyashana and Vishamashana produces Hikka, Shwasa and Kasa

and also lead to detrimental diseass ending in death.

Atyashana- Intake of excess quantity of food is known as Atyashana. It produces

Ama and Agni mandya.

Apathyas- These are Aharas specific to the roga and the rogi. This can be

interpreted as Avastha Viruddha.

Types of Viruddha-

Acharyas have classified Viruddha Aharas into different types based on many

factors.

Acharya Charaka has classified Viruddha Aharas into 18 types Viz.,

1. Desha Viruddha

2. Kala Viruddha

3. Agni Viruddha

4. Matra Viruddha

5. Satmya Viruddha

6. Dosha Viruddha

7. Samskara Viruddha

8. Veerya Viruddha

9. Koshta Viruddha

10. Avastha Viruddha

11. Krama Viruddha

12. Parihara Viruddha

13. Upachara Viruddha

14. Paka Viruddha

15. Samyoga Viruddha

16. Hrudaya Viruddha

17. Sampat Viruddha

18. Vidhi Viruddha



Apart from these eighteen types, Acharya Charaka has also mentioned about

Parasparaguna Viruddha (eg- Matsya+ Milk) and Swabhava Viruddha (eg- Visha).

Acharya Chakrapani has included Shareera Viruddha under Desha Viruddha.

Acharya Sushruta classifies Viruddha in to 6 types-

1. Samyoga Viruddha

2. Karma Viruddha

3. Maana Viruddha

4. Rasa Viruddha

5. Veerya Viruddha

6. VipakaViruddha

According to Ashtanga Sangraha, Viruddha can be classified as follows-

1. Asadrushat Viruddha

2. Sadrushat Viruddha

3. Ubhaya Viruddha

4. Samskara Viruddha

5. Matra Viruddha

6. Desha Viruddha

7. Kala Viruddha

8. Samyoga Viruddha

9. SwabhavaViruddha



Table.3 .Types of Viruddha according to different Acharyas

S.N Types of Viruddha CS SS AH AS BS

1 Agni Viruddha + - - - -

2 Avastha Viruddha + - - - -

3 Asadrushat Viruddha - - - + -

4 Desha Viruddha + - - + -

5 Dosha Viruddha + - - - -

6 Hrudaya Viruddha + - - - -

7 Kala Viruddha + - - + -

8 Koshta Viruddha + - - - -

9 Krama Viruddha + - - - -

10 Matra Viruddha + + + + -

11 Parihara Viruddha + - + - -

12 Paraspara Viruddha + - - - -

13 Paka Viruddha + - - - -

14 Rasa Viruddha - + - - -

15 Samyoga Viruddha + + + + +

16 Sampat Viruddha + - - - -

17 Sadrushat Viruddha - - - + -

18 Samskara Viruddha + + * + + +

19 Satmya Viruddha + - - - -

20 Swabhava Viruddha + - - + -

21 Upachara Viruddha + - - - -

22 Ubhaya Viruddha - - - + -

23 Vidhi Viruddha + - - - -

24 Vipaka Viruddha - + - - -

25 Veerya Viruddha + + + - -

*Acharya Sushruta has mentioned it as Karma Viruddha in place of Samkara Viruddha



Detail review on types of Viruddha-

1. Desha Viruddha (Incompatibility in context to habitat) –

The diet which has similar properties as those of the geographical region of the

residence is antagonistic in context with desha (habitat).

Eg- a) Intake of rooksha (dry), teekshna (sharp) and other substances that aggravate Vata

in arid region and unctuous, cold and other substances that aggravate Kapha in marshy

region b) Ashes and dusts also cause unwholesomeness to diets and drugs. c) Lavana

from Ushara desha

The diet that is incompatible to a person i.e deha desha considered as Shareera

Viruddha also can be included here. Eg- Honey is unwholesome when used by a

person whose body is afflicted by heat.

2. Kala Viruddha (Seasonal / Time Incompatibility)-

Food substances having similar quality as that of the season/ Rtu is considered as

Kala Viruddha.

Table.4. Viruddha aharas for different seasons

S.N Seasons Viruddha Ahara

1 Shishira Katu, Tikta, Kashaya,Laghu, Sheeta,Vatavardhaka Ahara

2 Vasanta Amla, Madhura,Guru,Snigdha Ahara

3 Greeshma Lavana,Amla, Katu, Ushna Ahara

4 Varsha Udamantham, Nadhi jalam,

5 Sharad Vasa, Taila, Avashyayam , Audaka, Anoopa mamsa, Ksharam, Dadhi

6 Hemanta Laghu, Vatala Ahara





3. Agni Viruddha (Incompatible to the power of digestion) -

Based on the Prakruti, Agni is of four kinds Viz., Mandagni, Teekshnagni,

Vishamagni and Samagni for Kapha, Pitta, Vata and Sama Prakruti respectively. One

should always take food after considering the four types of Agni, if not it will lead to

many diseases due to Agni Viruddhata.

Table.5. Viruddha ahara according to different Agni of individuals

S.N Type of Agni Viruddha Ahara Rogotpatti

1 Mandagni Guru, Snigdha, Madhura Ahara Grahani, Udara

2 Teekshna Laghu, Teekshna, Katu Ahara Bhasmaka roga

3 Vishama Rooksha, Laghu, Ushna, Snigdha,

guru, Sheeta Ahara at variance

Vibandha, Adhmana,

Udavarta

4 Sama Atimatra, Alpamatra, Vishamakala,

Vishama desha without following

Ahara vidhi

Aama doshas-(Alasaka,

Visoochika)

4. Matra Viruddha (Incompatible in quantity)-

For positive maintenance of health, one should take food in proper quantity. The

quantity of food again depends upon the power of digestion and metabolism (Agnibala).

The amount of food which without disturbing the equilibrium of doshas and dhatus, gets

digested and metabolized in proper time is regarded as Matravat Ahara. It certainly helps

the individual in bringing about strength, complexion, happiness and longevity. In

Aharamatra, total quantity as well as quantity of different items is considered. The former

is known as Sarvagraha (consideration of the food as a whole) and the latter as

“Parigraha” (consideration item-wise). This is very important from the view of the



concept of “balanced diet” because each item is considered with regard to the

requirement of the body.

It is not possible to decide in general the specific quantity of food to be consumed

by an individual as the quantity of food intake again depends on Agni (Matra

Agnimapekshaya).Considering these points, some guidelines to decide appropriate

quantity of food that is sufficient for a person’s good health has been explained. Stomach

is imagined to be divided into three equal parts and the food items should be taken

accordingly. One part filled with solid food, the second part with liquids and third part

should be left for Tridosha sanchara (movement of doshas).15

Symptoms of Samyak Matra (Appropriate quantity)-

1. No pressure on the stomach

2. No interference in the proper functioning of heart

3. No excessive heaviness in the lateral side of chest

4. No excessive heaviness in the abdomen

5. Proper nourishment of the senses

6. Relief from hunger and thirst

7. Feeling of comfort in all functions like standing, sleeping, walking and talking

8. Food taken in the morning should be digested by the evening and that of evening

should be digested by the next morning



Review on Matra Viruddha-

Food taken in improper quantity is considered as Matra or Mana Viruddha. It may

be of four kinds. They are-

1. Sarvagraha hina matra (deficient quantity of food as a whole)

2. Parigraha hina matra (deficient in the quantity of ingredients)

3. Sarvagraha atimatra (excessive quantity of food as a whole) and

4. Parigraha atimatra (excessive quantity of ingredients)

General symptoms of Amatravat Ahara are as follows-

Symptoms of Hinamatra sevana (Inadequate quantity of food)-

Impairment of the strength and complexion; Unable to have feeling of satiety;

Causing Udavarta; Impairment of longevity, virility and ojas; Affliction of body, mind,

intellect, senses; Impairment of the excellence of Dhatus; Causing inauspicious

conditions; Cause of eighty types of Vatavyadhis; Feeling of dissatisfaction;

Symptoms of Adhika Matra (Excess quantity of food)-

Adhika Matra (type of Matra Viruddhahara) aggravates all the Doshas, producing

different types of diseases Viz.,

Vata- Colicky pain, distension of abdomen, body ache, dryness of mouth,

giddiness, agnivikriti, various types of pains etc.

Pitta- Jwara, Atisara, Antardaha, thirst, giddiness and delirium.

Kapha- Vomiting, anorexia, cold, fever, laziness and heaviness in the body.



Table.6. Matra Viruddhata according to different Acharyas

S.N Examples CS16 SS17 AS18 AH19

1 Honey + Ghee in equal quantity + + + +

2 Honey + rain water in equal quantity + - - -

3 Honey + seed of the east Indian lotus + - - -

4 Honey + water in equal quantity - + + -

5 Honey + Oily foods in equal quantity - + - +

6 Honey + Oil + Ghee + muscle fat - - - +

7 Honey + Oily foods + rain water - + + -

8 Honey + Ghee + muscle fat - - - +

9 Honey + muscle fat - - - +

10 Honey + Oil - - - +

11 Water + any oily foods in equal quantity - + - -

5. Satmya Viruddha- (Incompatibility in context with suitability)

Satmya means substances of pleasure to one self. A substance conductive to an

individual is called “Satmya” and the use of such substances result in the well being of

that individual. Satmya Viruddha is intake of food against homologation. Intake of sweet

and cold substances by a person accustomed to pungent and hot substances.

Eg – Diet with hot, penetrating, oily and non-vegetarian articles along with alcohol is

contraindicated for a person suited to sweet, soft and vegetarian diet. This type of

unsuited food will produce even allergic disorders.

6. Dosha Viruddha (Incompatibility in context with humours)-

Utilization of drugs, diets and regimens having similar qualities with doshas but at

variance with the habit of the individual causes Dosha Viruddha.



Eg – A person of Vataja constitution, or in whom there is increase of Vata dosha, dry, hot

and spicy food is regarded as Dosha Viruddha.

7. Samskara Viruddha (Incompatibility in context with processing)

Samskara means processing the food items to change its qualities. Samskara is a

synonym of Karana. Karana (preparation) is one of the eight factors of dietetics codes

known as “Ashta aharavidhi viseshaayatana.” Karana (Preparation) is the process

performed to modify the natural properties of substances i.e., establish new properties by

destroying the natural properties of those dravya/substances.

“Samskaro Gunantaradaanamuchyate | Te Gunaastoyaagnisannikarsha Shoucha

Manthana Desha Kaala Vaasana Bhaavanaadibhi Kalaprakarsha

Bhajanadibhishcaadheeyante ||” (Cha. Vi.1/21)

The process is that which radically transforms the properties of substances. This

transformation is effected by Dilution, Application of heat, Clarification, Emulsification,

Storing, Maturing, Flavouring, Impregnation, Preservation and Container.20

For eg-

When rice is prepared by cleansing and applying heat, it attains lightness.

Curd by nature aggravates oedema, when churned it turns into buttermilk and

thereby it alleviates oedema. This illustrates the transformation of attributes by

churning.

On storing Amalaki (Emblica officinalis) along with other drugs inside the heap of

ash transforms the quality of Amalaki.

Kala is the time for maturation or the time prescribed for preservation of a

particular substance.Ingredients of Duralabhasava are required to be kept in a jar



smeared with ghee for fifteen days before use. This illustrates the transformation

by preservation.

A container also brings transformation of attributes, eg- Triphala Rasayana is to be

prepared in a new iron container smeared with the paste of Amalaki; At the time

of eating, one should keep Yusha, Vyanjana and Mamsarasa in silver vessels;

milk in golden vessel; buttermilk in bronze vessel; ghee in iron vessels; cooked

milk in copper vessels and, drinking water and boiled rice in mud etc.

Samskara Viruddha- The incompatibility of preparation is that where food/food

substance is converted in to poison during the course of preparation. Various

methods of preparation are used for processing of diet / food substances to get

more qualitative properties, but when these method, are not according to rules and

regulations of dietetics, one cannot get more qualitative properties but instead get

the harmful effect of food substances. So this type of Samskara (preparation) is

called Samskara Viruddha due to harmful effects it produces on the body tissues.

Eg :-

Curd or honey on heating becomes harmful to the body.

Madya and Bhallataka in a hot state.

Meat of harita bird held in a Daruharidra (Berberis aristata) stick or the same meat

cooked inside hot ash and sand along with honey.

Meat of peacock roasted on castor spit.

Meat of pigeon fried in mustard oil.



Meat of Godha, Lava, Tittiri, Mayura and Kapinjala cooked in castor oil using

castor plant or Daruharidra (Berberis aristata) wood as fuel.

Meat of Balaaka fried with fat of pig along with cocconut.

Meat of Bhaasa cooked on an iron rod.

Meat of Kapota processed in milk produces Majja Vyapat.

Haridraka (a kind of tuber used as a vegetable) fried in mustard oil cause

profound increase of Pitta.

Upodika (Basella alba) cooked with paste of tila.

Kampillaka (Mallotus philiphinensis) prepared with butter milk.

Pippali (Piper longum) processed in oil used for frying fish.

Kakamachi (Solanum nigrum) cooked in a pot in which fish or ginger has

already been cooked.

Leaves of Pushkara (Inula racemosa) and Rohinika fried in mustard oil.

Intake of ghee stored in bronze vessel more than ten days

8. Paka Viruddha (Culinary incompatibility)-

Preparation of food with a bad or rotten fuel and under-cooking, over-cooking

during the process of preparation are all considered as Paka Viruddha. The effect of

Mrudupaka, Madhyamapaka, Kharapaka, Aamapaka and Dagdha Paka has been

mentioned in the context of Sneha paka.

9. Veerya Viruddha (Incompatible in potency)-

Substances having cold potency in combination with those of hot potency is

regarded as Veerya Viruddha. Example- A food item prepared by using milk or fish



together becomes antagonistic as milk is of Sheeta veerya while fish is Ushna veerya.

Apart from this, the following combinations of Rasas produce Veerya Viruddha.

Madhura + Amla/ Lavana/ Katu; Amla + Tikta/ Kashaya; Lavana + Tikta/ Kashaya and

Katu rasa+ Tikta/ Kashaya

10. Koshta Viruddha-

It is the incompatibility in context with bowel. For example-

Administration of a mild purgative in a small dose for a person of Krura Koshta.

Administration of a strong purgative in higher doses for Mrudu Koshta person.

11. Avastha Viruddha (Incompatibility in context with state of a person)-

Incompatibility with reference to stage or condition of a person is known as

Avastha Viruddha.

Intake of Vata aggravating food by a person after exhaustion, sexual act, and

physical exercise.

Intake of Kapha aggravating food by a person after sleep or drowsiness.

In old age a light dry and cold diet is antagonistic as the age dominated by Vata

dosha.

Kashaya rasa Viruddha in Nava jwara

Uccha bhashana in Bastidana kala.

12.Krama Viruddha- (Contrary to the method or manner)

Incompatibility in context with the sequence. When a person takes his

meal without relieving himself of faeces and urine or eats without the feeling of

hunger or does not eat in spite of severe hunger then it turns out to be Krama

Viruddha.



13.Parihara Viruddha- (Incompatibility of the rules of Prohibition)

This is incompatibility of the rules of prohibition, where a person takes hot (food)

substances after a meal of the flesh of a boar and similar animals. ‘Parihara’ means ‘the

food substances which are to be avoided according to the condition. This condition may

be healthy or unhealthy. In diseased condition, intake of food substances, which are

mentioned as Apathya (unwholesome) in that particular disease is called Parihara

Viruddha. Eg- Intake of hot things after taking pork.

14.Upachara Viruddha- (Incompatibility of the rules of prescription)

Upachara word is derived from 'Upachaarya" that means- 1. Practice of medicine

2. Treatment 3. Chikitsa (Apte’s dictionary).Hence Upachara Viruddha is the act in

contrary to treatment /incompatibility of the rules of ingestion. Eg given as follows-

Intake of cold things after taking ghee.

Payasa (milk pudding) followed by mantha (solution of flour and sugar) is

incompatable and causes profound increase of Kapha.

Honey and ghee of unequal proportion along with rain water taken as after

drink.

15.Samyoga Viruddha (Incompatibility due to combination)-

Substances that are non- beneficial when used in combination are as follows-

Intake of Sheeta+Ushna/ Nava+Purana/ Aama+Pakwa food articles

Intake of Sour fruits or any sour things + Milk

Meat of the domestic, swampy, aquatic animals+ newly germinated paddy/ fat/

honey/milk/ jaggery/ Masha

Leaves of Rohinika, Pushkara and Jatuka taken with milk and honey



Meat of balaaka with wine or gruel

Kakamachi+ Piper longum/Piper nigrum/Madhu/Guda

Honey and seeds of Pushkara

Pilu + Kareera

Bisa (Lotus stalk) + Germinated grains

Sharkara (wine prepared with sugar) + Maireya (wine prepared with dates) +

Honey

Chilicima fish + Milk RaktaVyapatti, Mrutyu

Pork + Meat of dog + Milk Shankha roga, Galagraha and Sadyomrutyu

Wild roots and fruits+ Pork + Meat of dog+ Milk Shankha roga, Galagraha and

Sadyomrutyu

Lakucha fruit + Madhu/Milk Aandhya, Badirya and Mrutyu

Lakucha fruit+ Dadhi/Guda/Madhu/Ghee/Soup of Masha

Kadali phala+ Dadhi/ Takra/ Talaphala

16. Hrudaya Viruddha- (Incompatibility of palatibility)

This is the incompatibility of palatability where an article or substances of food is

unpleasant in taste. Here by term “Hrudaya” one should take “Manas” Any substances

which are not pleasant in taste. One should not take a food item for which the person has

apathy, when he is not in a good mood and so does not want to eat.

17. Sampat Viruddha- (Incompatibility with reference to the richness of quality)-

This is incompatibility with reference to richness of quality. For example- Intake

of substance that is not matured or over matured or putrified.Intake of food articles which

are not having edible qualities can be considered as Sampat Viruddha.



Over stale and over ripened fruits, meat of diseased animal, substances that are artificially

produced (by chemical manures) and of cross bred variety agricultural, poultry or cattle

products are prohibited. (Encyclopedia)

18. Vidhi Viruddha- (Contrary to the rules of dietetics)

One should take food substances / diet according to Upayoga Samstha mentioned

in the eight factors of diet and dietetics (Ashta aharavidhi Visesayatana). One should eat

food, which is hot, unctuous, non-antagonistic in potency and in due measure, after full

digestion of the previous meal, in a congenial place, provided with all the accessories,

neither too hurriedly, nor too leisurely, without talking or laughing with full

concentration and having proper regard to oneself. If a person is not following the rules,

then it is considered as Vidhi Viruddha. That is incompatibility of the rules of eating.

Eg- Food taken in a solitary place or in public.

Apart from these eighteen types explained by Acharya Charaka, Some more types

of Viruddha are added by Acharya Sushruta. They are as follows-

Rasa Viruddha (Incompatibility due to taste)-

Intake of food having contradictory tastes act contrary to the tissues and

considered as Rasa Viruddha.

Eg:-

Madhura rasa taken along with Amla, Lavana, Kashaya, Katu and Tikta rasa.

Amla rasa taken along with Lavana, Katu, Tikta, and Kashaya rasa.

Lavana dravyas with Katu, Tikta and Kashaya rasa.

Katu rasa dravyas with Tikta and Kashaya rasa.

Tikta rasa dravyas along with Kashaya rasa.



Vipaka Viruddha (Incompatibility due to rasa produced after digestion)-

Intake of food having contradictory Vipakas are incompatible to the tissues of the

body. Following rasas produce contradictory Vipaka in the body-

Madhura rasa with Katu, Tikta and Kashaya rasa.

Amla rasa with Katu, Tikta and Kashaya rasa.

Lavana rasa with Katu, Tikta and Kashaya rasa.

Acharya Vagbhatta explained three more types which are different from the rest-

a) Sadrushat Viruddha- i.e Even when the rasa, guna, veerya, vipaka are

similar some drug acts antagonistically. For eg- Ksheera and Panasa both

possess Madhura rasa, Madhura vipaka and sheeta veerya but when combined

produces incompatibility.

b) Asadrushat Viruddha- i.e Ksheera and Kulattha have contrary veerya and

vipakas i.e Sheeta and Madhura, Ushna and Amla respectively. Hence they

produce incompatibility.

c) Ubhaya Viruddha- Ksheera and Matsya possess Madhura rasa and Madhura

vipaka but Ksheera is sheeta and Matsya is ushna in veerya. It produces

incompatibility.

Effects of Viruddha Ahara-

Viruddhahara is similar to Visha in terms of the effects it produces in the body. A

person who regularly follows a diet which is marked by Viruddhashana, Adhyashana or

Ajeernashana ultimately reaches a stage of Aama visha which is as fatal as any other

Visham.21 Visham acts in different way inside the body- Teekshna Visha causes

immediate death or toxic effects whereas Gara or Dooshivisha have a long term effects.



The effect of Viruddhahara has to be studied in the lines of Visha i.e., it can cause

immediate death as that of teekshna visha or produce a devastating diseases as that of

Gara and Dooshi visha. The Viruddha Aharajanya rogas has been classified based on the

dhatus, srotas and rogamargas affected for clear understanding.

Table.7.Showing Viruddha Ahara Janya rogas

S.N Viruddha Aharajanya Rogas CS22 SS23 AS24 BS25

1 Jwara + - - -

2 Pandu + - - -

3 Klaibya + - - -

4 Agninasha - - + -

5 Visarpa + - - -

6 Visphotaka + - + -

7 Rakta dushti - - - +

8 Kilasa + - - -

9 Kushta + - + -

10 Raktapitta - - + -

11 Gulma - - + -

12 Galagraha + - - +

13 Prameha - - + -

14 Moorcha + - - -

15 Majja Vyapat - - - +

16 Vata vyadhi - - + -

17 Shandyam - - -

18 Santana Dosha - - -

19 Balanasha - + -

20 Moodagarbha - - + -

21 Stanya roga + - - -

22 Yakshma - - + -



23 Aadhmana + - - -

24 Aama Visha + - - -

25 Grahani + - + -

26 Trushna + - - -

27 Shotha + - + -

28 Ashmari - - + -

29 Smrutinasha - - + -

30 Indriyanasha - + + -

31 Chitta nasha - - + -

32 Unmada + - - -

33 Mada + - + -

34 Arshas - - + -

35 Vidradhi - - + -

36 Shankha roga - - - +

37 Bhagandara + - + -

38 Aandhya + - - +

39 Badhirya - - - +

40 Peenasa + - - -

41 Kasa + - - -

42 Arshas - - + -

43 Udakodara + - - -

44 Krimija rogas - - - +

Factors that nullify the effects of Viruddha Ahara

Under certain conditions Viruddha ahara will remain harmless to the body and the

effects are nullified. If the incompatibility is homologous to the person and if he has

strong Jataragni, it will not disturb the harmony of the tissues. The ill effects of Viruddha

ahara will be nullified as long as the person is young, performs vyayama and take snigdha

ahara regularly. The jataragni will be able to deal with the harmful effects to certain



extent if Viruddha ahara is of lesser quantity. If the quantity is more, it produces morbid

effects.

Viruddha Ahara which have become accustomed and which are in smaller

quantity do not cause trouble to the body. Instead they even cure the diseases by causing

variations in the Doshas.

Eg- 1) Curd +Milk given in certain morbid conditions.

2) Sarpa visha in condition of Udara

3) Sthavara visha as an antidote for Jhangama Visha

Treatment principles in Viruddha Ahara

Choice of treatment in Viruddha Aharajanya roga is Shodhana followed by Shamana and

Abhisamskruti.

Shodhana-

Depending on the nature of the disease, selection of shodhana can be done. For

example Vamana in case of Kushta and Unmada, Virechana in case of Visarpa and

Udara.

Shamana-

After Shodhana, Shamana is done by administering aushadas to the specific rogas

produced by Viruddha Ahara.



Abhisamskruti-

Abhisamskruti is the final step to strengthen the body in order to withstand the

effects of Viruddhahara which is achieved by Hitaharas including Rasayanas.

Flow Chart.1. Viruddha Ahara janya Roga Chikitsa

Viruddhajanya roga

Chikitsa sutra

Shodhana

Abhisamskruti

Hitaharas & Rasayanas

Shamana

Aushadas specific to rogas



Modern concepts of Viruddha Ahara

When one look into the equivalent concepts to Viruddha Ahara, spectrum of

disease ranging from food intolerance to food poisoning can be considered. Hence a brief

description of relevant conditions has been dealt.

Incompatibility-

The term ‘Incompatible’ generally means ‘antagonistic in action.’ The quality of

not being mixed without a chemical change, or without countering the action of the other

ingredients in a compound is known as incompatibility.26

Food intolerance-

An abnormal non-immunological response to the ingested food is regarded as

food intolerance. The basis for intolerance may be pharmacological, enzymatic,

metabolic or toxic.27

Food allergy/ Food Hypersensitivity-

It is an immunologic reaction to food to which a patient has become sensitized. It

requires a first exposure (sensitization), which stimulates the production of IgE

antibodies; subsequent exposure produce symptoms. Sensitivity to almost any food may

develop but it develops most frequently to milk, egg, wheat, shellfish and chocolate.

Food allergies are Type-I reactions, symptoms can appear within minutes. Mild

symptoms such as urticaria, eczema, abdominal cramps and GIT upset are most common,

but food allergies also can cause life threatening systemic anaphylaxis.28



Milk-Protein allergy in infancy-

It is also called as Cow’s Milk Protein Intolerance(CMPI) that manifest within the

initial few months of life, usually a few weeks after the initial exposure to cow’s

milk.Clinical symptoms include diarrohea, vomiting, abdominal colic, irritability, failure

to thrive and GIT haemorrhage. It generally subsides by second year of life.29

Lactose intolerance-

It is developed in children due to the deficiency of the enzyme lactase, a non-

specific disaccharide in the intestinal mucosa. It is of three kinds Viz., (1) Congenital

lactase deficiency occurring in new born. It can be compared to Dosha Viruddha. (2)

Lactase deficiency associated with premature infants. It can be compared to Avastha

Viruddha. (3) Acquired Lactase deficiency occurring in adults and children.29

Sucrase and Isomaltase deficiency-

Congenital sucrose- isomaltase deficiency has been reported to occur in some

subjects and they do not tolerate sucrose or isomaltose.

Lathyrism-

It is a crippling disease characterised by paralysis of the leg muscles occurring in

adults consuming large quantities of the seeds of Lathyrus sativus or other Lathyrus

species over long periods. The toxin present in lathyrus seed has been identified as -

oxalyl amino alanine (BOAA) that produces neuro-lathyrism in humans and osteo-

lathyrism in animals.30



Food Poisoning-

It is acute gastro-enteritis caused by the ingestion of food or drink contaminated

with either living bacteria or their toxins or inorganic substances and poisons derived

from plants and animals. The condition is characterized by (a) history of ingestion of

common food or in communal feeding (b) attack of many persons at a time (c) similarity

of signs and symptoms in majority.

Food additives-

Food additives are non-nutritious substances which are added to food, generally in

small quantity, to improve its appearance, flavour, texture or storage properties.

Uncontrolled or indiscriminate use of food additives may pose health hazards. For

example, certain preservatives such as nitrites and nitrates lead to the production of

nitrosamines that have been implicated in cancer aetiology. According to PFA, any food

that contains additives which are not permitted; if it exceeds the permissible limit then the

food is considered as adulterated.

Foodborne diseases-

It is defined as a disease, usually either infectious or toxic in nature, caused by the

agents that enter the body through the ingestion of food. Foodborne disease may develop

due to (a) Foodborne intoxicants and (b) Foodborne infections.

Epidemic dropsy-

The outbreak of Epidemic dropsy is reported in India, due to the contamination

of mustard oil with argemone oil. Sanguinarine is a toxic alkaloid isolated from argemone

oil found to be the causative agent that interferes with the oxidation of pyruvic acid.



Endemic ascites-

In Madhya Pradesh the outbreak of ascites has been reported. Studies conducted

by National institute of Nutrition, Hydrebad showed that the local population subsit on

millet which gets contaminated with the weed seeds of Crotalaria. On Chemical analysis,

crotalaria seeds were found to contain pyrrolizidine alkaloids which are hepato toxic.31



REVIEW ON MADHU

Madhu is the natural sweetner used since ancient time. It is considered auspicious

and also recommended as a daily food especially in the rainy season. Madhu also has

many medicinal properties and is used widely as adjuvants for many medicines. A detail

review of Madhu is given below-

Derivation

The term Madhu is derived from the root word ‘Man’ ‘Avabodhane’ and suffix

‘U’ i.e. “Mani +dha Madha + ‘U’nadi Madhu || (Siddhanta kaumudi)

Definition

Manyante visheshena jananti janaa yasmin| (SKD)

That which is honoured by the people specifically is Madhu.

Manyateabhilashyate madhu pushparasaakhyam|| (Amarakosha)

That which is known for its speciality is known as Madhu

Synonyms of Madhu

The word ‘Madhu’ has many synonyms mentioned in various texts, the

knowledge of which will be more beneficial in understanding different aspects of Madhu.

The synonyms are listed below-



Table.8. Synonyms of Madhu32,33,34,35,36,37,38

SN Synonyms SKD DN RN BN KN MN DGH

1 qÉkÉÑ – Honey + + + + + + +

2 ¤ÉÉæSì-Species of honey + + + + - - +

3 qÉÉ¤ÉÏMÇü - Belonging to bee + + + + - + +

4 qÉÉÍ¤ÉMÇü- Honey + + + - - - -

5 qÉÉÍ¤ÉMüÉ- Honey - - - + - - -

6 mÉÑwmÉÉxÉuÉÇ - Decoction of flowers + + + - + + -

7 mÉÑwmÉUxÉÇ – Nectar + + + - + + -

8 mÉÑwmÉUxÉÉÀûrÉ- Derived from nectar + - - - - - -

9 mÉÑwmÉUxÉÉå°uÉÇ - Derived from nectar + - - + - - +

10 MÑüxÉÑqÉÉxÉuÉÇ - Flower liquor + + + - - - -

11 xÉÉUbÉÇ - Derived from bees + + - + + - +

12 mÉÌuÉ§ÉÇ – Honey + - + - - - -

13 ÌmÉ§rÉÇ – Honey + - + - - - -

14 qÉÉkuÉÏMÇü - Intoxicating drink + - - + - - +

15 pÉ×ÇaÉuÉÉliÉÇ - Vomitus of bees + - - + - - +

16 qÉÉÍ¤ÉMüuÉÉliÉÇ - Vomitus of bees + - - - - - +

17 uÉUOûÏuÉÉliÉÇ - Vomitus of wasp + - - - - - +

18 qÉÉÍ¤ÉMüÌuÉOèû - Excretion of bees - - - - + - -

19 qÉMüUÇS:- Juice of flowers + - - - - - -

20 qÉUlS/qÉUlSMü:- Nectar of flowers + - - - - - -



Regional names/ Vernacular names-39

Sanskrit- Madhu

Hindi- Shahad

Kannada- Jenutuppa

Telugu- Thena

Tamil- Then

Malayalam- Then

Gujarati- Madha

Marathi- Madha

Bengali- Madhu

Urdu- Shahad

Arabic- Asal

Parsi- Shahad, Agaveen

English- Honey

Latin- Mel

Types of Madhu-

Madhu can be classified into different types based on the source (honey bee),

storage time, method of collection and quality. A brief classification has been mentioned

here and the explanation will be given later.

I. Based on the types of honey bees – Eight types viz. Makshikam,

Bhramaram,Kshoudram, Pouttikam, Chaatram, Aarghyam, Auddhalakam and

Dalam

Table. 9. Types of Madhu based on the types of honey bees 40,41,42,43,44,45,46,47,48

S.No Types CS SS AS YR DN RN BN KN MN

1 Makshikam + + + + + + + + +

2 Bhramaram + + + + + + + + +

3 Kshoudram + + + + + + + + +

4 Pouttikam + + + + + + + + +

5 Chatram - + - + + + + + -

6 Aarghyam - + - + + + + + -

7 Uddhalakam - + - + + + + + -

8 Dalam - + - + + + + + -



II. Based on the duration of storage-

Two types viz. a) Nava Madhu- Fresh honey b) Purana Madhu- Old/ Stored honey

III. Based on the method of collection-45

Two types viz.- i) a) Sadosha- Contaminated honey b) Nirdosha Madhu-

Uncontaminated honey. ii) a) Aama- Unripened honey b) Pakwa Madhu- Ripened honey

IV. Based on the quality –45

a) Dushta Madhu- Spoiled honey b) Shreshta Madhu- Pure honey

V. Based on the state in which it should be consumed-45

a) Sheeta Madhu b)Ushna Madhu

Madhu Varga according to different Acharyas

Madhu one of the Jangama dravyas has been placed under different categories by

different Acharyas which has been listed below-

Charaka Samhita (CS) - Ikshu varga; Vamanopaga ghana; Shonitasthapana ghana.

Sushruta Samhita (SS) - Madhu varga

Ashtanga Sangraha (AS) - Ikshu varga, Niruhopayogi; Shirovirechanopayogi;

Sandhaneeya and Rudhira sthapana Ghana

Ashtanga Hridaya (AH) - Ikshu varga

Dhanwantari Nighantu (DN) - Suvarnadi varga

Raja Nighantu (RN) - Paneeyadi varga

Bhavaprakasha Nighantu (BN) - Madhu varga



Kaiyyadeva Nighantu (KN) - Oushadi varga

Priya Nighantu (PN) - Drava varga

Madhanapala Nighantu (MN) - Ikshu varga

Samanya Madhu gunas- General properties of Madhu

Madhu (Honey) possesses the following properties in general. Acharya Charaka

opines the qualities as follows- Rasa- Madhura; Anurasa- Kashaya; Guna- Guru,

Rooksha, Sookshma, Vishada; Veerya- Sheeta; Doshakarma- Kaphapitta shamaka and

Vatala. Acharya Sushruta differs in his opinion that Madhu has Laghu guna; Rooksha

Veerya and Tridosha shamaka. The other properties of Madhu are- Deepana, Grahi,

Lekhana, Balya, Sandhana, Srotovishodhana, Swarya, Varnya, Vrshya, Hrdya,

Prasadajanaka, Soukumarya, Medhya and Ruchikaraka. It is good in alleivating the

ailments of Netra, Chardi, Visha, Hikka, Shwasa Kasa, Shosha, Atisara, Raktapitta,

Krimi, Trshna, Moha, Medoroga, Kushta, Arshas, Kshata and Kshaya.

Special quality of Madhu-

Madhu has a unique property i.e. ‘Yogavahitwa’. It is the quality of Madhu that

enhances the potency of drug in unison and helps to reach towards the target organ.

Qualities of different kinds of Madhu-

The qualities of different kinds of Madhu- explaining different colours of the

honey, different colours of honey bees etc., are dealt in detail below-



Table.10 . Colour of different kinds of honey

S.N Types DN RN BN KN MN DGH YR

1 Makshikam Taila + + + + + +

2 Bhramaram Shweta + SpK + SpK SpK SpK

3 Kshoudram Kapila + + + + + Kapisha

4 Pouttikam Ghritam + + + + + Taila

5 Chatram Apeeta + PK PK - - PK

6 Aarghyam Pingala

+ - Like Madhuka

niryasa

- - Shweta

pishangaka

7 Uddhalaka Suvarna + - - - - PK

8 Dalam Paatala Apeeta - - - - Nanavarna

Note-SpK- Sphatika; PK- PeetaKapila.

Table.11. Colour of different kinds of honeybees

S.No Honey bees RN BN KN DGH

1 Makashika Kapila Pinga Pingala Pinga

2 Bhramara Anjana - - -

3 Kshudraa Pingala Kapila Pinga Kapila

4 Puttikaa Pinga Krshna Krshna Krshna

5 Saragha (Chatra ) Peetapingala Kapilapeeta Peetakapila Kapilapeeta

6 Aarghya/ Shwetaka - Shwetaka Peeta Shwetakam

7 Uddhalaka Kapila Kapila Pingala Kapila

8 Dala Indraneeladala - Pingala -



I. Based on the types of honey bees-

Madhu collected from different kinds of honey bees possesses different

qualities. Among all the types of Madu, Makshika Madhu is considered as the

best. The qualities of different Madhu are explained below.

1. Makshika Madhu- The honey collected from reddishbrown coloured, big-sized,

wild variety of honeybees that feed on different kinds of flowers is known as

Makshika Madhu. It has Madhura rasa; Laghu, Rooksha guna; Sheeta veerya and

acts as Vata Shamaka. Such honey is good in Netrarogas, Kamala, Arshas, Vrana,

Kshaya and Shwasa rogas.

2. Bhramara Madhu- The honey collected from collyrium coloured minute sized

honey bees is known as Bhramara. It possesses Madhura rasa; Madhura vipaka;

Guru, Rooksha, Picchila, Abhishyandi gunas and Sheeta veerya. It alleivates

Raktapitta, Mukhajadya, Mootrajadya and Atyantajadyahara.

3. Kshoudra Madhu- It is the kind of honey collected from reddish coloured, smaller

sized honey bees. It has Laghu, Rooksha, Sookshma, Abhishyandi, Picchila

gunas; Sheeta veerya; acts as Pitta Vata Shamaka and is lekhaneeya, prameha

hara and netramayahara.

4. Pouttika Madhu- It is the kind of honey collected from the poisonous honeybees

which are reddish brown, dark coloured, smaller in size resembling the

mosquitoes (mashaka). The bees generally reside in the hollow burrows of trees

and feed on poisonous flowers and rice (anna). Such a honey is called Pouttika. It

possess the following properties such as Laghu, Rooksha guna; Ushna veerya;



alleviates Kapha but aggravates Pitta and Vata thereby causes Raktapitta and

Daha. It is indicated in Mootrakrchra,Granthi, Kshata and Shosha.

5. Chatra Madhu- It is the kind of honey collected from the honeybees which are

reddish in colour found in the Himalayan forests and their hives are of the shape

of umbrella. Such a honey has Madhura rasa; Madhura vipaka; Guru, Picchila

guna; Sheeta veerya; allievates Rakta, Pitta vikaras, Krimi, Shwitra and Prameha

but aggravates Vata dosha.

6. Aarghya Madhu-The honey collected from the yellow coloured honey bees

having sharp stings is called as Aarghya Madhu. It possess Kashaya, Tikta,

Madhura rasa; Katu vipaka; alleviates Kapha and Pitta. It acts as Balapushtikaraka

and Chakshushya.

7. Auddalaka / Auddhala Madhu- The honey which is collected from the reddish

brown coloured bees that resides inside the soil near the ant-hills is known as

Auddhala Madhu. It possess Kashaya,Amla, Tikta rasa; Katu vipaka; Ushna

veerya; aggravates Pitta. It is indicated in Kushta, Visha, Aruchi and also acts as

Svarya.

8. Dala Madhu- It is the kind of honey collected from the honey bees which are red

or brown coloured, smaller in size and that reside inside the burrows of trees.

Generally the honey is collected from the leaves of trees and is known as Dala

Madhu. It possesses Katu, Amla, Madhura rasa; Kashaya anurasa; Vatala;

Lekhaneeya. It is generally indicated in Chardi, Aruchi and Prameha.



II. Based on the duration of storage

Madhu differs in its qualities based on the duration of storage as storage has effect

on the properties of Madhu.

a) Nava Madhu/ Fresh honey-

Nava Madhu or freshly collected honey has the following properties- Rasa-

Madhura bhooyishta; Guna- Guru, Snigdha, Abhishyandi and Sara; Karma-

NatiShleshmahara, Pushtikaraka, Sthoulyakara.

b) Purana Madhu/ Stored honey-

Purana Madhu or stored honey has Rooksha, Anabhishyandi, Grahi gunas; acts as

Atilekhana, Medohara and Sthoulyagna. It also acts as Tridosha shamaka. Madhu

attains puranatva after one year of collection.

III. Based on the method of collection

Madhu should be collected in the prescribed manner as explained by Acharyas,

failing which it leads to contamination.

i) Sadosha and Nirdosha Madhu-

The honey that is extracted by beating the bee- hives with a stick is considered to

be as effective as Rasayana and called as Nirdosha Madhu. It alleviates Hikka,

Gudhankura, Trividha Shopha, Kapha and Vrana. The honey collected otherwise is

considered as Sadosha Madhu.



ii) Aama and Pakwa Madhu-

Aama Madhu/Unripened honey aggravates Vata and Shosha but alleviates

Aama, Gulma, Ruja, Pitta, Daha, Asradosha and Antravruddhi. Whereas Pakwa

Madhu/Ripened honey is Tridoshashamaka and alleviates Ruja, Jadya,

Jihwaamaya. It acts as Ruchya, Balakaraka and Virya vruddhikaraka.

IV. Based on the quality

During collection of honey due care should be given to avoid spoilage of Madhu.

a) Dushta Madhu/ Spoiled honey-

The honey which has not been strained; contains insects, dead bees, stings

and other parts of bees; that which has undergone fermentation due to prolonged

storage; that which is produced in home are all considered as Dushta Madhu and is

not good for health.

b) Shreshta Madhu/ Pure honey-

The honey which has the opposite qualities that of the Dushta Madhu is

considered as Shreshta Madhu.

V. Based on the state in which it should be consumed

a) Sheeta Madhu-

The honey generally is collected from the bees which feed on different

kinds of flowers including poisonous flowers and at times also from poisonous

honey bees, therefore it is advised by our Acharyas to consume honey in cold

state which bestows health.



b) Ushna Madhu-

When honey is heated or consumed with hot drugs, hot things, hot

seasons, and hot region and administered in diseases which produce heat in

the body results in deleterious effects and may eventually lead to death.

Instructions for the intake of Madhu- 49

Certain rules and regulations are to be adopted while consuming Madhu as it is a

product of nectar fromed from different flowers including poisonous flowers, collected

also from the bees which may be poisonous or from poisonous region. Hence due

precautions should be taken before administering Madhu.

Madhu- Sevanayogyata-

In natural state

By Kapha prakruti persons

People of Marshy land

Children and Young persons

In little quantity

Madhu – Sevana Ayogyata-

Heated honey

Honey with equal ghee

For Vata Prakruti

People of arid land

In Old age

In large quantity



When consumed in Ayogya conditions it produces the following effects-

When heated honey and honey with equal ghee is taken it results in Samskara and

Matra Viruddhatwa respectively. When Madhu is taken by Vata Prakruti persons or in

old age it aggravates Vata. If taken in large quantity, it causes Madhvama and also

produces emergency conditions of Vata rogas because of its rooksha guna and Kashaya

Anurasa.



Table.12. Properties of different kinds of honey

S.N Madhu types

Rasa Anu Rasa

Guna Veerya Vipaka Dosha Karma

Samanya Karma

1 Makshikam Madhura - Laghu, Rooksha Sheeta - Vata Shamaka

Chakshushya, Kamala, Arsha, Vrana hara, Kshata, Hikka, Shwasa hara

2 Bhramaram Madhura - Guru, Rooksha, Picchila, Abhishyandi

Sheeta Madhura - Rakta vikara hara, Atyanta jadya, Mootra jadya hara

3 Kshoudram - - Laghu, Sukshma, Abhishyandi, Picchila

Sheeta - Pitta Vata shamaka

Lekhaniya, Prameha & Netra roga hara

4 Pouttikam - - Laghu, Rooksha Ushna - Kapha hara, Pitta Vata kara

Raktapitta & Dahakaraka, Mootrakrucchra, Gulma & Kshaya hara

5 Chatram Madhura - Guru, Picchila Sheeta Madhura - Switra, Prameha, Trushna, Bhramam, Krimi Raktapitta & Vishapaha

6 Aarghyam Kashaya, Tikta, Madhura

- - - Katu Kaphapitta shamaka

Chakshushya

7 Uddhalaka Kashaya, Amla, Tikta

- - Ushna Katu Pittala Kushta, Visha, Svarya, Ruchya

8 Dala Katu, Amla Madhura

Kashaya Laghu, Rooksha - Pittakara Kaphahara

9 Nava Madhurabhooyishta - Guru, Snigdha, Abhishyandi Sara

- - Kaphahara Pushtikrut and Brhmana

10 Purana - - Rooksha, Anabhishyandi, Grahi

- Tridosha Shamaka

Atilekhana, Medohara and Sthoulyagna

11 Sadosha - - - - - Kapha hara Rasayana, Hikka, Arshas Shopha & Vrana hara

12 Aama - - - - - Vatakrut Pittahara

Aama,Gulma, Ruja, Pitta,Daha, Asradosha & Antravruddhi hara

13 Pakwa Tridosha Shamaka

Ruja, Jadya, Jihwaamaya hara Ruchya, Balya, Vrushya



MODERN REVIEW ON HONEY

Definition

Honey is the food derived entirely from the work of bees operating upon the

nectar of flowers and other sweet exudation of plants. The nectar and saccharine

exudation of plants are gathered, modified and stored in the comb by honey bees. It is

then obtained by allowing the honey to drain from the comb or by submitting the

honeycomb to pressure or is prepared by melting the honey, allowing the scum to rise to

the surface and straining. It must be free from the foreign substances such as parts of

insects, leaves etc., but may contain pollen grain.50

Composition of honey-

Honey contains various principle nutrients. A 100g of honey contains 20g

moisture, 79.7g carbohydrates, 0.1g of protein, 0.01g calcium, 0.02g phosphorous, 0.9

mg iron, 0.01g vitamin B1,0.2mg nicotinic acid, 0.04mg of riboflavin, 0.01mg vitamin B6

and 0.06mg pantothenic acid per 100g. There is no fat and vitamin A. One gm honey

produces 320 KCal of energy.51 The honey shall not contain more than 25% moisture,

0.5% ash and 5% sucrose. It should contain 10% honey dew, 65% reducing sugar and

fructose- glucose ratio shall not be less than 0.95%. 52

Moisture-

Moisture content is one of the most important characteristics of honey, as it

influences the keeping quality, granulation and texture of honey. The natural moisture of

honey in the comb is that remaining from the nectar after ripening. After extraction of the

honey from the comb its moisture content may change depending on the conditions of



storage. It may range between 13 and 25 percent. If honey has more than 25 percent

moisture and contains a sufficient number of yeast spores, it will ferment. Such honey

should be pasteurized, that is, heated sufficiently to kill such organisms.

Sugars-

Honey is a carbohydrate material, with 95 to 99.9 % of the solids being sugars.

Dextrose (glucose) and levulose (fructose) are the main sugars account for about 85 % of

the solids in honey. Apart from those 25 others sugars have been found. All of these

sugars are more complex than the monosaccharides, dextrose and levulose. Twelve

dissaccharides, ten trisaccharides and two more higher saccharides- isomaltotetraose and

isomaltopentaose have been identified. Most of these sugars are present in quite small

quantities. The complex sugars do not occur in nectar, but are formed during the ripening

of honey.

Proteins-

In honey, 40 to 65 percent of the total nitrogen is in protein. The presence of

proteins causes honey to have a lower surface tension than it would have otherwise,

which produces a marked tendency to foam and form scum and encourages formation of

fine air bubbles. The quantity of amino acids in honey is small and of no nutritional

significance. Amino acids are known to react slowly or more rapidly by heating, with

sugars to produce yellow or brown materials. Part of the darkening of honey with age or

heating may be due to this.

Acids-

The acids of honey are responsible for the stability against micro-organisms. It



account for less than 0.5 percent of the solids. Gluconic acid is the major one found in

honey. It arises from dextrose through the action of an enzyme called glucose oxidase.

Minerals-

When honey is dried and burned, a small residue of ash invariably remains, which

is the mineral content. In general, ash content is highly variable ranging from 0.02% to

over 1%. Potassium accounts for about one-third of the total amount. The other minerals

in honey are sodium, calcium and magnesium. Less abundant elements found are iron,

manganese, copper, chlorine, phosphorus, sulphur and silicon.

Enzymes-

One of the characteristics that set honey apart from all other sweetening agents is

the presence of enzymes. These conceivably arise from the honey bee and also from

traces of plant enzymes. Those most prominent enzymes are added by the honey bees

during the conversion of nectar to honey. Some of the most important honey enzymes are

α - glucosidase, β- glucosidase, glucose oxidase, catalase, acidphosphatase, and α-

amylase and β- amylase.

α-glucosidase- It is probably the most significant honey enzyme and is responsible for

inversion of nectar disaccharide sucrose into its constituent monosaccharide.

Glucose oxidase- It is responsible for the antibacterial property of honey. This

antibacterial agent is produced by the action of glucose oxidase on its glucose substrate

i.e dextrose that produces hydrogen peroxide and gluconic acid. The acidity of this is

largely responsible for the low pH of honey resulting in low water activity which helps to

stabilize honey against fermentation.



Catalase- It is responsible for converting hydrogen peroxide into water and oxygen.

Diastase- It is added to nectar by bees during ripening and is a mixture of α- and β-

amylase. The activity of honey diastase is known to be reduced upon heating and is

measured to estimate the extent of heating to which a honey has been exposed.

Properties of honey-53

A. Physical Properties

Honey is a thick, syrupy liquid, the colour of which varies from pale yellow,

amber to darkish red and nearly black. The variations in colour are almost entirely due to

the plant source although the climate may modify the colour through the darkening action

of heat.

Honey is translucent when fresh but becomes opaque and granular due to the

crystallization of dextrose. It has a characteristic odour and sweet, faintly acrid taste.

Aroma and flavor of honey is ascribed to the various sugars, aminoacids, tannins and

minor, non-volatile substances. The additional contribution to flavor is due to alkaloid

compounds specific to plant sources. The honey is levorotatory at 200º C slightly acid to

litmus paper. The specific gravity of honey is 1.35-1.36 at 20ºC.

B. Physico-chemical Properties

The polyphenol compounds in honey causes astringency and also responsible for the

development of final colour, taste and aroma. The oxidation of phenolic compounds in

the presence of polyphenolase enzymes result in brown discolouration.

The heat induced browning reactions take place in two ways- Caramelization and

Maillard reaction.



Caramelization reactions-The sugars in dry state or in syrup when heated undergo

reaction depending upon the temperature and the presence of catalyst. There will be

reactions leading to the equilibrium of aanomeric and ring forms, inversion of sucrose,

condensation reactions leading to the formation of oligosaccharides and polysaccharides,

isomerisation, dehydration and fragmentation. All these lead to the formation unsaturated

polymers with brown colour.

Maillard reactions- It is a complex group of many reactions wherein carbonyl group of

acyclic sugars combine with basic amino group of proteins, peptides, aminoacids etc.,

The sugar amines polymerize to form red brown to dark brown compounds.

Granulation of honey is due to the dextrose which can spontaneously crystallize. The

granulated honey becomes more susceptible to spoilage by fermentation and

objectionable to consumer. Honey that has granulated can be returned to liquid state by

direct or indirect heating. Stirring accelerates the dissolution of crystals.

The honey is hygroscopic i.e. absorbs water from the surrounding. The honey has

lower water activity as it is a saturated mixture of monosacharides – fructose and glucose.

It create a poor environment for the growth of micro- organisms. pH of honey ranges

between 3.2- 4.5. The range of pH is maintained by gluconolactone and gluconic acid in

it.

C. Medicinal properties-

i) Anti- bacterial activity-

Honey act as bacteriostatic rather than bactericidal. It is not a suitable medium

for bacteria for two reasons-i.e. it is fairly acid and has too high sugar content. The



killing of bacteria by high sugar content is due to the osmotic effect. The presence of

the enzyme inhibine in honey is also responsible for its antibacterial activity. Usually

gram (+) ve strains are first affected followed by gram (-) ve strains which takes up to

48 hrs.

ii) Anti- fungal activity-

Honey acts against yeast and some species of Aspergillus.

iii) Antioxidant activity-

Honey contains many phenolic and non- phenolic antioxidant molecules

which protects the cell from damage when consumed regularly.

Production of honey-

Honey bees ingest nectar and store it in their honey sac. The honey sac weighs 40-

70mg when full and can account for more than 90% of the weight of the bee. Enzymes

are secreted and mixed with the sac contents. Then the contents are passed to the house

bees that carry out the ripening process.

Ripening of honey-

This consists of alternately expelling and ingesting the honey sac fluid for 15-20

min. When the nectar has lost about half of its moisture, small droplets of this semi-

ripened honey are deposited in the comb cells. In 1-3 days numerous enzyme- mediated

changes occur and honey attains density. The cells are then capped with wax and the

honey stored as food. Compositional changes continue during ripening, the most

prominent result being the production of an increase in complex array of sugars due to



the continuous enzyme activity. Hydrogen peroxide is also formed by the enzymatic

reactions in order to stabilize honey against micro- organisms.

Processing of honey-

The honeys that are available in the market are processed honey. The primary

objectives of commercial honey processing include maintaining the desired texture, to

reduce granulation and also keep it free from fermentation.

Bee keepers remove the hive units and then remove the cappings by mechanical

slicing. The honey is then extracted by centrifugal separation. Raw honey contains

pollen, wax, sugar-tolerant yeast and often some crystals of glucose monohydrate.

The methods that are well established and accomplished since many years for

processing honey before large scale marketing are as follows-

Pasteurization- The honey is generally heated at 70-75º C to destroy yeasts to dissolve

sugar crystals.

Pressure filtration- This process is used to produce liquid honey after controlled heating.

Liquification- Central Bee Research and Training Institute in Pune have recently

developed a honey liquefier which serves as a pre- heating unit for further honey

processing.

The other commercial processes are straining, micro- filtration and vacuum evaporation.

Generally the application of heat to honey requires careful control so that the colour,

flavor and aroma of honey are not impaired.



Quality deterioration of honey- 54

The honey quality can be affected in number of ways. The common factors that

contribute to quality deterioration includes- Fermentation, Thermal treatment and

formation of Hydroxy Methyl Furfuraldehyde (HMF).

Fermentation-

Fermentation of honey is caused by the action of yeasts upon the sugars dextrose

and levulose, resulting in the formation of ethyl alcohol and carbon dioxide. The alcohol

in the presence of oxygen is broken down into acetic acid and water. As a result, honey

that has been fermented may taste sour. The yeasts responsible for fermentation occur

naturally in honey. It is osmophilic thus can germinate and grow at much higher sugars.

Thus, the water content is one of the factors concerned in spoilage by fermentation. The

other factors that results in fermentation are yeast count, temperature of storage and

granulation.

Thermal treatment-

Honey processing frequently requires heating both to reduce viscosity, and to

prevent crystallization or fermentation.55

Honey heating is generally carried in two ways: (1) In air- ventilated chambers, at 45-

50˚C for 4/7 days or (2) By immersion of honey drums in hot water. Although, the second

method is more efficient, the first is most common.56

Effects of thermal treatment

Thermal treatment applied to honey produces a non- enzymatic browning due to

maillard reaction. There will be formation of brown pigments and development of



antioxidant activity during the reaction. But heating honey may also result in the adverse

effects as follows- change in flavours and viscosity, melting of sugar crystals, change in

sugar compositions, destruction of vitamins and bionutrients, decrease in diastase activity

and increase in hydroxymethyl furfural (HMF) content.

As heating produce severe quality loss, the boundary thermal treatment assumed

as the most severe conditions able to produce a permissible quality loss was at 140 C

during 15 sec in the transient stage and 30 sec in isothermal stage.

Hydroxy Methyl Furfuraldehyde (HMF)-

HMF is used as an indicator of heat and storage changes in honey. Heating of

honey results in HMF, which is formed during acid-catalyzed dehydration of hexoses.

Heat increases the rate of reaction. The increase in speed of the reaction is exponential to

the increasing heat. HMF occurs naturally in honeys and usually increases with the heat

treatment of honey. The codex level of HMF must not exceed 80 mg/kg honey in tropical

countries. The factors influencing HMF production in honey are (a) Temperature and

time of heating57 (b) Storage conditions; use of metallic containers58 (c) The chemical

properties of honey from which floral source honey has been extracted- like pH, total

acidity and mineral contents.59



REVIEW ON GHRUTA

Ghruta, obtained from the class Mammalia of the animal kingdom (Jangama

dravya) is an essential food. According to Ayurveda, it is one among the ‘Nitya

Sevaniya Aharadravya’. Ghruta is a Shreshta sneha dravya as it is used from birth and

is Satmya to all the people and for all age groups. It acts as Tridosha shamaka since it

alleviates Vata by its Snigdhata and Madhuryata, Pitta by Saumya guna and Sheeta

Veerya and Kapha by Samskara. Ghee is a clarified butter fat, considered as vehicle

for various medicines, as it increases the potency of medicine. It also acts as a

Medhya dravya and increases intellectual power.

Derivation-

The term ghruta is derived from the root word ‘Ghru’ and added with the prefix

‘kta’. The root word indicates ‘Ghru’‘Ksharana deeptyo’ and ‘Ghru’ ‘Prasravane’

which means to scarify, to shine and to sprinkle respectively.60

Definition-

Navaneetaja Padartha:| (Amarakosha)

Ghruta is the product of navaneeta (butter).

Pakwa navaneetam | Pakwa navaneete sarpimshi cha | (SKD)

Ghruta (ghee) is the one produced by heating the butter.

Sarpirvileenam Ajyam tu Ghaneebhootam Ghrutam Bhavet| 61

When it is in liquid state it is termed as ‘Sarpi’ and when in denser

(semi solid) state called as ‘Ghruta’.



Table.13. Synonyms of Ghruta- 62,63,64,65,66,67,68,69

S.No Synonyms SKD DN RN BN KN MN DGH VSS

1 bÉ×iÉÇ- Ghee + + + + + + + +

2 AÉerÉqÉç- Clarified butter + + + + + + + +

3 WûÌuÉ:- Oblation + + + + + + + +

4 xÉÌmÉï:- Liquid ghee + + + + + + + +

5 mÉÌuÉ§ÉqÉç- Auspicious + + + - + - - +

6 lÉuÉlÉÏiÉMüqÉç-Made of butter + - - - - - - +

7 lÉuÉlÉÏiÉeÉqÉç- Born out of butter - + + - + - - -

8 eÉÏuÉlÉÏrÉÇ-Vitalising - + - - + - - -

9 AqÉ×iÉqÉç- Used for yagna + + + - - - - +

10 AqÉëÑiÉÉÀûrÉ-Acts like Nectar - - - - + + - -

11 AÍpÉbÉÉU:- Sprinkled over fire + + + - + - - +

12 AÉkÉÉUÇ-Helps to maintain fire - - - - + + - -

13 WûÉåqrÉqÉç- Used for an oblation + - + - - - - +

14 AÉrÉÑ:- Bestows life + - + - - - - +

15 iÉæeÉxÉqÉç -Offering in Agni + - + - - - - +

16 AÉeÉqÉç- Clarified butter + - - - - - - +

17 mÉÑUÉåQûÉzÉ- Clarified butter - - - - - - - +

18 iÉÉårÉSÇ- Gives rain by offering

ghee in oblation

- - - - - - - +

19 uÉÌ»ûpÉÉåarÉÇ- Increase digestive

fire

- - - - - - - +

20 mÉÉæjÉÇ- Auspicious - - - - - - - +



Regional Names or Vernacular Names-70

Sanskrit – Ghruta

Hindi – Ghi

Kannada – Tuppa

Tamil – Neyi

Telugu – Neyi

Malayalam – Neyyu

Marathi – Tup

Gujarati – Ghee

Thulu – Nai

Urdu – Ghee

Bengali – Ghee, Ghruta

Arabic – Shamana

Latin – Butarum deparatum

English – Clarified butter, Ghee

Classification of Ghruta

Classification is grouping of drugs into different categories on the basis of

certain criteria. Classification is mainly done on the basis of its source, consistency

and duration of storage.

I. Based on the source- 71 A) 1. Navaneetotha ghruta- Samanya ghruta prepared

out of navaneeta 2. Ksheerottha ghruta- Prepared out of cream derived

from ksheera

B) 1. Go ghruta 2. Maahisha ghruta 3. Aja ghruta 4. Oushtra ghruta 5.

Avi/Aavika ghruta 6. Ashwa ghruta 7. Kharabha ghruta 8. Hastini ghruta

9. Naari ghruta 10. Ekashapha ghruta 11. Vana Chaga ghruta

(Among these Goghruta – Best; Avighruta - Worst)

II. Based on the consistency-

1. Sarpi- Liquid

2. Ghruta- Semisolid



III. Based on duration of storage-

“Older the ghruta more the beneficial effects.” Based on the duration of

storage ghee has been classified as follows-72

1. Nootana ghruta - Fresh ghee

2. Hyastana Dugdhotta ghruta- Ghee prepared out of one day old milk

3. Purana ghruta- 1-10 yr old

4. Prapurana ghruta- More than 10 yr old

5. Koumbha ghruta- 11-100 yr old

6. Maha ghruta- More than 100 yr old

Ghruta gana vargeekarana -

Ghruta is categorized under different vargas by different Acharyas. It has been listed

below-

Charaka Samhita (CS) – Go-rasa varga

Sushruta Samhita (SS) - Ghruta varga

Ashtanga Hridaya (AH)- Ksheera varga

Ashtanga Sangraha (AS) - Ksheera varga

Bhavaprakasha Nighantu (BN) – Ghruta varga

Kaiyyadeva Nighantu (KN) – Ghruta varga

Raja Nighantu (RN) – Ksheeraadi varga

Dhanwantari Nighantu (DN) -Suvarnaadi varga

Madhanapala Nighantu (MN) – Paneeyadi varga

Priya Nighantu (PN)- Drava vargas



General qualities of Ghruta-

Ghruta generally means the ghee prepared out of Navaneeta or butter. It has

the following properties-

Rasa- Madhura; Guna- Snigdha, Mrudu, Shlakshna, Guru, Yogavahi,

Alpabhishyandi, Soumyam; Veerya- Sheeta; Vipaka- Madhura ; Dosha karma-

Tridosha Shamaka.

The other karmas of Ghruta are listed in the table below

Table.14. General properties of Ghruta-

Pharmacological actions CS73 SS71 AH74 BPN72 KDN73 DN75

Nirvapana + - - - - -

Mrudukara + - - - - -

Svaraprasada + + + + + -

Varnaprasada + - - - + -

Smruti vruddhi + + + + + +

Buddhi vruddi + + + - + +

Agni vruddhi + + + + + +

Meda vruddhi - + + + + +

Kanti vruddhi - + + + + +

Soukumarya - + + - - +

Balakara - + + + + +

Ayushya - + + + - +

Vayasthapana - + + + + -

Chakshushya - + + + + +

Rasayana - - - + - -

Ojo vruddhi + + + + + -

Tejo vruddhi - + - + - -



Lavanya vruddhi - + - + - -

Rochana - - - + - -

Jeevana - - - - + -

Brihmana - - - - + -

Dhatu karma

Rasa vruddhi + - - - - -

Shukra vruddhi + + + + + +

Medo vruddhi + - - - - -

Jeevana - - - - + -

Rakta shamaka - - - + - -

Rogaghnata

Unmada + + + + - -

Shosha + - + + - -

Jwara + + + + - -

Mada + - - - - -

Apasmara + + - + - -

Murcha + - - - - -

Gara + - - - - -

Yoniroga + - - - - -

Karnaroga + - - - - -

Shiroroga + - - - - -

Udavarta - + - + - -

Kustha - - - + - -

Shoola - + - + - -

Anaha - + - + - -

Kshataksheena - - + - - +

Parisarpa - - + + - +

Vrana - - - + + -

Timira - - - + - -



Go Ghruta

Among all the varieties, Ayurveda recommends Goghruta as the best for both

food and medicinal purposes. Hence if not specified, the word ghruta always refers to

Goghruta.

It is the best among the ghrutas. It possesses the following properties- Rasa-

Madhura; Vipaka – Madhura; Veerya –Sheeta. Ghruta, because of its varied qualities

acts in number of ways (Sahasra veeryam vidhibhi karma sahasrakrut ) to impart

good health. Dosha Karma- pacifies the Tridoshas. Samanya karma- Shreshta

Chakshaushya, Balya, Vrushya and Vayasthapaka; Increases Shukra and Agni;

Promotes dhee, smriti, medha and kanti; Good for bala, vruddha and sukumara;

relieves shrama (fatigue) and enhances shareera sthirata.Rogaghnata- It pacifies

Kshata produced by Shastra and agni, Visarpa, Unmada, Shosha, Jwara and Visha.71

The properties of all other types of ghruta are given in table below.



Table.15. Vividha Ghruta Gunas -Qualities of different kinds of Ghruta

Vividha Ghrutas Rasa Guna Veerya Vipaka Dosha karma Karma

Aja Madhura Laghu Laghu, Katu Kaphahara Deepana, Chakshushya, Vrushya, Balya, Indicated in Kasa, Shwasa, Arshas and Kshaya

Mahisha Madhura Guru Sheeta Guru, Madhura

Vata-sheshmahara*

Raktapittaghna, Dhrutikara, Kantikara, Balya, Varna-prasada, Cures Durnama, Grahani, acts as Deepana, Hrudya, Chakshushya, Vrushya

Oushtra Madhura Sheeta Katu Kapha-vatahara Indicated in Shopha, Krumi, Kushta, Visha, Gulma, Udara, Moorcha, Moha, Unmada, Garavisha, Jwara, Apasmara, Shosha, acts as Deepana

Aavika Katu Ati guru Laghu, Katu Kapha-vatahara, Na Cha Pitta prakopaka

Indicated in Yoni dosha, Shopha, Kampa, Shosha, Ashmari, Sharkara, acts as Chakshushya, Agnisandhookshya And Asthi vardhaka

Ashwa Katu, Madhura, Kashaya

Guru** Ushna Laghu, Katu Kaphahara, Vata karaka

Indicated in Moorcha, Visha, Netra roga, Daha, acts as Deepana, Tarpana, Mootra baddhakara

Hastini Kashaya, Tikta

Laghu Kapha-pittahara Indicated in Kushta, Visha, Krumi, Rakta dosha, acts as Agni vardhana, Mootra sangrahi, Vishtambhakaraka

Karabha/Gardabhi Kashaya,,Katu Ushna Laghu Kapha-vatahara Indicated in Mootra dosha, Visha, Arshas, Krumi, Shosha, Kushta, Gulma, Udara, acts as Balya

Stri/ Naari

Madhura Laghu Tridosha shamaka

Indicated in Yoni dosha, Rakta dosha, acts as Chakshushya, Amruta,



Agnivardhaka, Vishapaha, Ruchikaraka

Ksheerottha Sheeta Tridosha shamaka

Indicated in Raktapitta, Bhrama, Moorcha, Mada, Netraroga, Daha, acts as Sangrahi, Tarpana

Ekashapha Sakashaya Laghu Ushna Katu Kaphakara Mootrabaddhata, Agno deepana Vanachaga Ateeva uru Buddhipatavakara, Balakara Nootana Indicated in Balakshaya, Pandoroga,

Kamala, Netraroga, acts as Tarpana, Shramahara

Haiyangaveena Indicated in Jwara, acts as Chakshushya, Deepana, Brumhana, Ruchikara, Vrushya, Balya

Purana Tikta, Katu Rooksha Sara, Teekshna

Ushna Katu Tridoshahara Indicated in Shwasa, Peenasa, Kasa, Yonishoola, Shirashoola, Akshishoola, Karna shoola, Shopha, Moorcha, Kushta,Visha,Unmada,Apasmara, Timira, Graha dosha, acts as Vranashodhaka, Lekhana,Chedana, Deepana, Sroto vishodhana

Prapurana Laksharasa sannibha vilayana, Ugragandhi

Koumbha Rakshoghna Maha Kaphahara, Vata

vardhaka Indicated in Timira, acts as Balya, Pavitra, Medhya, Bhootahara

Note- * According to Sushruta, it is Vata-pittahara.

** According to Kaiyadeva Nighantu, it is Laghu



Modern Review on Ghee

Definition

Ghee is Indian clarified butter especially from the milk of a cow or

buffalo.76

As per PFA, A.11.02.21. Ghee means the pure clarified fat derived solely

from milk or curd or from butter to which no colouring matter or

preservative is added.77

In another definition, Ghee is the Indian name for clarified butter fat, and

is usually prepared from cow’s milk, buffalo’s milk or from mixed milks.

Composition of ghee

Ghee is largely made up of glycerols, free fatty acids, phospholipids, cholesterol,

fat soluble vitamins, glyceryl ethers and alcohol. The total proportion of polyunsaturated

fatty acids is about 3-4 % in ghee. The fatty acid composition greatly influences the fat

contents and rheological properties such as melting and crystallization behavior,

solubility, surface activity, and ability to form emulsions.

Average Analytical constants of cow’s ghee

According to Food and Nutrition chart, 100gm of ghee contains 0% moisture,

100% fat, 600 I.U of Vit- A, 2 mg of Vit-E. According to Encyclopedia of Food science,

Butyrorefractometer reading- 43.2, Saponification value- 227.3, Reichert- Miessl value-

26.7, Polenske value- 1.76, Kirschner value- 22.1, Iodine value- 33.7 and Melting point-

28-41 C



Method of preparation of ghee by curd or cream method has been shown in the following

flow diagram.78

Flow Chart.2. Method of Ghee preparation

Milk

Centrifugal separation Fermentation

Cream Curd

Ripened / Unripened Churning

(Indigenous)

Churning (Western)

Cream butter Desi butter

Direct clarification

Prestratification

Ghee

Quality of ghee-

Flavour-

Aroma and taste constitute the flavour of ghee. The preferred ghee flavours range

from ‘slightly curdy’ to ‘pronounced curdy’. The most important flavour components of

ghee are as follows- FFAs: 6-12mg/g of fresh ghee, methyl ketones: 6-12p.p.m, a

complex mixture of 44 lactones, both δ- lactones and γ- lactones, 29-43p.p.m; aldehydes,



ketones and alcohols. Recent research work shows that - Flavour is greatly influenced by

the fermentation of the cream or butter and the heating processes. Carbonyls, lactones

and free fatty acids are reported to be the key ghee flavouring compounds.

Texture-

Granulation of ghee is an important criterion for its selection and it develops a

lower level of rancidity than the ghee kept in liquid state. Milk fat has the unique

property of forming grains because it is made up of a wide variety of complex

triacylglycerol mixtures with varying melting points. The presence of FFAs markedly

increases the grain size.

Colour-

In cow’s ghee carotene imparts a yellow colour where as buffaloes’ ghee is white

because of the absence of carotene. Ghee produced by the direct cream method has a

brownish color compared to that prepared by creamy butter process. Stratification results

in a light colour. A more intense heating will result in a darker colour, especially if the

raw material has been fermented.

Quality deterioration

The quality attributes of ghee are affected by many factors, such as quality of

base material, extent of lactic acid fermentation, time and temperature of heating, rate of

cooling, package type, filling conditions, presence of oxygen and contaminants such as

iron and copper, exposure to sunlight, temperature, and duration of storage.



Benefits of ghee :-

It contains natural antioxidants, Vitamin A and E

It enhance digestion and absorption

It contains no Transfatty acids

It is the natural alternative for the hydrogenated cooking oils.

It acts as anticarcinogenic as it contains high amounts of linoleic acid.

Ghee in the diet lowers the prostaglandin levels in serum but also decreases the secretion

of leukotrienes by macrophages.



MATERIALS AND METHODS

Materials

The study has been conducted in three steps-

Literary study

Analytical study

Experimental study

Literary Study

Materials-

The materials used for the study were obtained from

Vedic scriptures

Sanskrit lexicons

Ayurved ka Vaignanika Itihaas

Ayurved ka Bruhat Itihaas

Bruhattrayis and Laghutrayis with commentaries

Various dravyaguna books like nighantus

Text books of Biochemistry

Food and nutrition text books

Guidelines for experimental study

Published articles in reputed journals

Various media like internet etc.

Related research works



Source-

Library of Government Ayurveda Medical College and Hospital, Mysore.

Library of JSS Ayurveda Medical College and Hospital, Mysore.

Technical library of Defence Food Research Laboratory, Mysore.

Methodology-

Literature survey is done using the required Tantrayuktis (scientific technique),

and various Nyayas (methods). For e.g.; Adhikarana, Nirvachana are applied in

Ayurvedic reviews where as for modern reviews, Atidesha tantrayukti and

Simhavalokana nyaya are used. Apadesha, Yoga, Uhya are used for the discussion to

elicit the reason behind the results and Prayojana is used to arrive at the conclusion.

Analytical study

The Analytical and Experimental study has been carried out in Biochemistry and

Nutrition division, Defence Food Research Laboratory, Mysore under the valuable

guidance of a renowned scientist, Dr.Anila Kumar. K.R.

Materials for Analytical Study-

For Analytical study, the materials used are- 1.Drugs 2.Chemicals and

3. Instrumentation

1. Drugs-

Unprocessed natural honey (raw) procured from Madikeri forest officer,

Karnataka

Processed honey (Dabur) procured from market.



Both the honey samples were heated to 60ºC and 140ºC

Cow’s ghee was procured from the local village, Tamilnadu

Processed ghee (Nandini) was procured from Mysore dairy

Cow’s ghee mixed with equal quantity of unprocessed honey

Heated cow’s ghee mixed with equal quantity of 140ºC heated

unprocessed honey

2. Chemicals-

Source- All the biochemicals used in these investigations were of highest purity

and procured from Sigma company, USA; Merck, Germany; Sisco Research

Laboratory, Mumbai; Across Organics, Mumbai; Spectrochem, Mumbai and S.D.

Fine Chemicals, Mumbai. All the organic Solvents were of “Analytical Reagent”

grade.

3. Instrumentation-

o Micro weighings- Schimatzu AEL- 160 model electronic balance

o Other weighings- Kern or Anamed electronic balance

o Pyknometer

o Soxhlet apparatus for fat extraction

o Eutech 5102 model pH meter

o Spectro photometer

o Gas chromatography (Chemito model no.8610)



Methodolgy-79

After careful quality check measures of the samples i. e evaluation of total and

reducing sugars and moisture evaluation, the analytical study has been conducted.

1) Total Ash

Ash content in a food sample indicates the presence of inorganic residue after the

destruction of organic matter. The ash value was calculated as described in AOAC

International. The tare weight of silica dish (initial weight) was noted. A known weight of

the sample was taken in silica dish and ignited. Then the silica dish was transferred to

muffle furnace and was ashed at 600 ± 20 °c for 4- 6 hrs. The dish was cooled after

removing from muffle furnace and the weight (final weight) was noted. The difference in

initial and final weight gave the total ash content and is expressed in percentage.

2) Acid insoluble Ash

The above silica dish was taken and 25 ml of dilute HCl was added to it. Then it

was gently boiled over a low flame for 5 minutes and filtered using an ash less filter

paper. The dish was washed thoroughly with hot water to remove the traces of ash

particles. Then the filter paper was returned to the dish, ignited and kept in muffle

furnace for 4- 6 hrs. After cooling the final weight was noted. Acid insoluble ash of the

sample was derived by finding the difference in final weight and dish tare weight. It is

then expressed in percentage.

3) Specific gravity

Specific gravity is the term used to express the relative masses of equal volumes

of materials and water being measured at a stated temperature. For this measurement, a



specific gravity bottle known as pyknometer has been used. Specific gravity of the sample

was determined by AOAC method.

Weight of Sample Specific gravity =

Weight of water

4) Organoleptic characteristics

Sensory evaluation or organoleptic characteristics was done by group of quality

control panelists on colour, aroma, odour, flavour, taste and texture using a nine point

hedonic scale.

5) p H value

pH is defined as the logarithm of the reciprocal of hydrogen ion concentration in

g per litre. It is of importance as a measure of active acidity which influences the flavour

or palatability of a product. It was determined using pH meter

6) Hydroxymethylfurfuraldehyde (HMF)

HMF in honey was studied using spectro photometry as mentioned in AOAC

international (1983). Accurately weighed 5 gm of honey was taken in a small beaker and

transferred to volumetric flask with 25 to 50 ml of water. Then 0.5 ml of Careez Solution

I was added and mixed. Then 0.5 ml of Careez Solution II was added, mixed and the

volume was diluted with water. Then the contents were filtered through filter paper

discarding the first 10 ml of filtrate. 5 ml of filtrate was pippeted into each of two 18 ±

150 mm test tubes. 5 ml of water was added to one test tube (sample) and 5 ml NaHSO3

solution was added to other test tube (reference). Then it is mixed well in votex mixer

and the absorbance of the sample was determined against the reference at 284 & 336 nm.



Calculation:-

HMF in mg / 100 g honey = (A284 - A336) ± 14. 97 ± 5 /g of sample

7) Browning index

About 5 gm of sample was dissolved in 67% of alcohol and the sample was

incubated for half an hour to one hour. Then it was filtered through Whatman filter no.1

and the absorbance A was noted in the filtrate @ 420 nm.

8) Estimation of Polyphenols

Aliquots of sample extract were taken in clean dry test tubes i.e. 0.1ml, 0.2ml and

0.4ml. To this required amount of distilled water was added to make up the volume up to

3.0ml. Then 0.5ml of Folin`s Ciocaletus reagent [1: 1 dilution in distilled water] was

added and incubated for 3 minutes. Then 2 ml of 20% sodium bicarbonate was added

and placed in boiling water bath for 1 minute and cooled. The colour was measured in the

spectrophotometer at 650nm.Gallic acid was used as standard for making the calibration

curve.

9) Estimation of flavonoids

The determination of flavonoids was performed according to a colorimetric assay.

Aliquots of sample extract were taken in clean dry test tubes i.e. 0.1ml, 0.4ml and 0.8ml.

To this required amount of distilled water was added to make the volume up to 1ml. Then

75µl of 5% of sodium nitrite and 150µl of 10% aluminum chloride was added to it and

incubated for 5 minutes. Then 0.5ml of 1M sodium hydroxide and 0.7ml of distilled

water was added. The absorbance was taken immediately at 510nm. Catachin was used as

standard for making the calibration curve.



10) Antioxidant activity by Diphenyl picryl hydroxyl (DPPH) method

The antioxidant activity is determined by spectrophotometric method. 0.1g/ml of

sample was taken in a test tube and 2.9ml of methanol was added. It was centrifuged at

5000 rpm for 15 minutes. To the filtrate 0.5ml of Diphenyl picryl hydroxyl radical

(DPPH reagent) was added and allowed to stand for 45 minutes. In the mean time a

control sample was also maintained by taking 3.0ml of methanol and 0.5ml of DPPH was

added and incubated along with sample tube. After 45 minutes, optical density was

measured in spectrophotometer at 515nm.

The antioxidant activity of the sample was calculated using the formula-

% scavenging activity = Absorbance control – Absorbance sample x 100

Absorbance control

11) Estimation of Peroxide value

Peroxide value was determined according to the Association of Official

Analytical Chemists (1993). In an Iodine flask 5 gm of sample was taken with 100 ml of

chloroform and kept for one hour in a flask shaker. Next day the contents were filtered.

Then 20 ml of extract was taken in flask, and then 30 ml of glacial acetic acid and 1 ml of

saturated potassium iodide was added and kept in dark for 15 - 20 min. Then 50 ml of

distilled water and 2- 3 drops of 5% starch indicator was added. The solution turned blue.

Similarly the experiment was repeated in blank sample and titrated against 0.1 N sodium

thiosulphate. The end point was milky white.

Per oxide value was calculated in milliequ of O2 / Kg sample.

Peroxide Value = TitreValue ± Normal quantity of Sodium thiosulphate ± 1000

Weight of fat in 20 ml extract



12) Estimation of free fatty acid (FFA)

In an iodine flask, 5 gm of sample was taken with 100 ml of chloroform. The

contents were kept one hour in a flask shaker. Then 20 ml of extract was taken in flask a

and kept in hot air oven for half an hour till all the chloroform gets evaporated and the

flask was cooled. Then 1: 1 methanol + benzene were added to the flask. Then 2- 3

drops of phenolphthalein indicator was added and titrated against 0.1 µ std. alcoholic

potassium hydroxide (KOH) solution. The end point was pale pink.80

Free fatty acid value was measured in % of oleic acid by the following formula-

FFA = (Sample titre value - Blank titre value) ± Normality of KOH ± 56.1

Weight of fat in 20 ml extract

13) Estimation of Thiobarbituricacid (TBA) value

In an iodine flask, 5 gm of sample was taken and 7.5 ml of 3NHCl and 75 ml of

distilled water were added. Then the distillate was collected. A 20 ml distillate was taken

in a boiling tube; 2 ml of 0.67 % TBA solution was added and heated in boiling water

bath for 35 min. It was then cooled and optical density was noted at 540 nm. TBA value

was measured in mg malonaldehyde per kg fat. It was calculated as follows-81

TBA value = Optical density X 3.2

Weight of sample X 0.15

14) Estimation of Saturated to Unsaturated fatty acid ratio

Appropriate aliquots of the fat thus extracted from ghee samples in duplicates

were processed for the preparation of methyl esters of the constituent fatty acids using

borontrifluoride.



The methylated samples were analyzed for their fatty acid profile by gas

chromatography using FID detector coupled with data station. The operational conditions

of Gas Chromotography were standardized, using both the individual fatty acid methyl

esters. The chromatographic conditions employed were as follows: the initial column

temperature maintained at 140˚C was programmed to rise to a maximum of 250˚C at the

rate of 6˚C per minute. The temperature of the injector and detector were kept at 240˚C

and 250˚C respectively. The carrier nitrogen gas was monitored to maintain a flow of rate

of 25 ml/ minute.

The methylated samples of the fats extracted from various processed products

were diluted with petroleum ether to obtain 20 mg concentration of the 40-60˚C fatty

acids per ml. An aliquot (1.0µl) of the diluted material was injected and the peaks

obtained as a result of separation of various constituent fatty acids were identified by

comparing the retention time values with those obtained using standards of not only the

different individual fatty acid methyl ester but also a standard mix of the various fatty

acid methyl esters.82

Experimental study

The experimental study has been carried out on male albino rats to evaluate the

toxicity that arise on consuming Ushna Madhu (heated honey) and Sama Ghruta Madhu

(Equal quantity of ghee and honey). Moreover it is believed that heating honey produces

peroxides that produce harmful effects in the body; hence a new attempt is made to find

the effect of equal mixture of heated honey and heated ghee.



Procurement of animals-

Healthy male rats (Rattue norvegicus) of Wistar strain reared in the Defence Food

Research Laboratory animal facility, Mysore has been taken for study.

Selection of animals-

The rats that weigh about 100-150 g were used in the entire studies. Clearance of

experimental design by the Institutional Ethical committee for rats was taken.

Instrumentation-

Wired cages for each rats

Weighing machine

Aluminum containers

Incubator

Micro pipette

Plastic vials

Centrifuger


II

Schimatzu model uv1601- Spectrophotometer

Chemicals-

All the organic Solvents were of “Analytical Reagent” grade. The reagents used in the

biochemical assays were of highest purity and procured from AGAPPE Diagnostics, Kerala,

Crest Bio-Systems, Goa and Span Diagnostics, Surat.

Methodology-

Treatment of experimental rats

The rats before attaining the weight were kept on pellet diet (Nutrilab Rodent) procured

from Tetragon chemie Pvt.Ltd, Bangalore for a period of 7 days before the commencement of

experiments. The pellet diet was powdered and stored in air tight container at room temperature

during the course of study. The powdered pellet diets were weighed out into feeding cups and

water was added to the diet and mixed into paste; spillage of diet was largely prevented by this

method. Food and water were given ad libitum.

Grouping-

Rats were divided into six groups of six rats each.

Group I: fed with normal pellet diet served as a control.

Group-II: fed with honey along with pellet diet.

Group-III: fed with heated honey with pellet diet.

Group-IV: fed with ghee with pellet diet.

Group-V: fed with honey mixed ghee with pellet diet.

Group-VI: fed with heated- honey mixed ghee with diet.

Fixation of dosage-

The dose of honey and ghee was decided as prescribed in Ayurvedic literature. As per


III

Ayurvedic practice, for 70 kg men, the normal dose of honey recommended is 48g/day (1 pala

matra) and recommended dose of ghee is 30 ml/ day (hraseeyasi matra).

Hence the dose for 150g rats was calculated as follows-

Dose of honey 48 X 150 = 102 mg

70

Dose of ghee 30 X 150 = 64 µl

70

Thus the dose of honey was fixed at 102 mg of honey /day and that of ghee was

calculated to be 64µl /day.


IV

Study design for animal study-

The animals were kept at ambient temperature and exposed to light- dark cycle of 12

hours each.To find out the acute toxicity the rats were fed with the drugs along with pellet diet

and observed for 48 hours. As there was no lethality, the study was continued for a period of six

weeks to find sub-chronic toxicity.

The following observations were made in rats-

During the course of study- The observations made on

Weight gain (weekly)

Food intake (weekly)

Serum biochemical parameters

After sacrifice- The observations were made on

Relative organ weight

Liver enzyme study

Histopathology reading

Weight gain and Food intake-

The left over diet was collected, dried and weighed every day to determine the food

intake. Weekly food intake and weight gain were monitored.

Methods for Serum Study-

Procurement of serum-Blood was collected from the orbital plexus of the rats on 0 day and at

the end of six weeks before sacrifice and properly stored in plastic vials. The vials were


V

centrifuged at 3000 rpm for 15- 20 minutes to collect the serum.

Biochemical study-All the serum bio-chemical parameters have been estimated by the

spectrophotometric method using cuvette of 1cm light path at 37ºC temperature and in

wavelength of 505nm.with the help of respective experimental kits.

Table. 16. Biochemical parameters adopted to study the samples

S.N Bio-chemical Parameter Methods adopted

1 SGOT and SGPT Tietz (1970)

2 Serum creatinine Modified Jaffe’s Kinetic method as described by Bowers et

al (1980)

3 Total protein End point assay as described by Jhonson et al (1999)

4 Serum albumin End point assay as described by Jhonson et al (1999)

5 Serum globulin Total proteins - Serum albumin

6 ALP Schlebusch et al (1974)

7 Total cholesterol Allain et al (1974)

8 HDL cholesterol Precipitation method as described by Grillo et al (1981)

9 LDL Cholesterol Freidwald’s Formula- LDL Cholesterol (mg/ dl) = (Total

Cholesterol) – (Triglyceride/5) – (HDL Cholesterol)

10 Urea Modified Berthelot method as described by Searcy.etal

(1967)

11 Uric Acid Uricase method as prescribed by Fossatj et al (1980)

12 Triglycerides GPO- PAP method as mentioned by Jacobs et al (1960)

13 Bilirubin (total & direct) Modified DMSO method as described by Walter et al

(1980)

14 Glucose GOD- PAP method as described by Trinder et al (1969)

Enzyme study in Liver Sample

Preparatory procedure-


VI

After the completion of experimental period i.e., 6 weeks, the rats were sacrificed under

mild anesthesia (sodium pentobarbitone, 50 mg/ kg body weight i. p.) and liver, kidney, heart,

brain and colon tissues were quickly excised. The tissues were quickly washed in ice- cold

isotonic sodium chloride solution and blotted dry before weighing. The tissue samples were

processed for biochemical and enzymatic assays as per the prescribed procedures.

1) Hydroperoxides

About 0.05 g of the lipid isolated from liver was thoroughly mixed with 5.0 ml of

chloroform: methyl alcohol solvent mixture (2:1). This was followed by centrifugation at 800 x g

for 5 minutes separating the upper phase distinctly. Upper phase is removed using syringe.

Lower layer is recovered into a test tube and taken to dryness under a stream of nitrogen. While

still under the stream of nitrogen 1ml of acetic acid: chloroform solvent mixture (3:2) is added

followed by 0.05 ml of potassium iodide (dissolve 6.0 g of KI in 5.0 ml of water on ice, which

has previously been bubbled with nitrogen gas for 5 min).

The test tubes are then stoppered and kept in dark for 5 minutes exactly. This is followed

by the addition of 3 ml of cadmium acetate (0.5%). The solution mixture is then centrifuged at

800 x g for 15 min. The optical density of the upper phase is read at 353 nm against a blank

containing the complete assay mixture minus the lipid. Standardisation of the reaction was done

by using cumene hydroperoxide as the peroxide standard. The molar extinction coefficient of

cumene hydroperoxide is 1.73x104/M.83

2) Conjugated dienes

About 0.05 g of the lipid isolated from liver was mixed thoroughly with 5.0 ml of

chloroform: methanol solvent mixture (2:1) followed by centrifugation at 800-x g for 10 minutes.


VII

This aided in separation of phases. Most of the upper phase was removed either by suction or by

the aid of a syringe. The lower chloroform layer was placed in a test tube and was taken to

dryness in a stream of nitrogen gas. The lipid residue was dissolved in 1.5ml of cyclohexane and

the absorbance at 233 nm was determined against a cyclohexane blank. The amount of

conjugated dienes produced was calculated using a molar extinction coefficient of 2.52x104/M.83

3) Thiobarbituric acid reactive substance (TBARS) measured as Malondialdehyde

About 0.5ml of tissue homogenate in phosphate buffer, pH 7.0 was added to 0.5ml of

10% TCA. 2.0ml 0f TBA mixture (0.35% TBA, 0.2% sodium lauryl sulphate, 50mM FeCl3 and

0.5mM BHT in 0.1M glycine HCl buffer, pH 3.6) was added to the homogenate containing TCA.

The above reaction mixture was heated in boiling water bath for 10 min. After cooling the

mixture was centrifuged at 1000 x g for 10 min. The OD of the supernatant was measured at

532nm. The concentration of MDA was estimated using the molar extinction coefficient,

1.56x105/ M/cm.84

4) Catalase (EC 1.11.1.6)

Catalase was assayed in tissue homogenates by following the spectrophotometric

degradation of H2O2 at 20˚C. About 1.0 g of liver or colon tissue was homogenized in the cold in

4 volumes of 0.05 M potassium phosphate buffer in 0.9% saline solution (pH 7.4), containing

0.1% (v/v) ethanol, in a motor driven tissue homogenizer equipped with a teflon pestle. The

homogenate was centrifuged at 4˚C for 10 minutes at 700 x g to sediment nuclei and cell debris.

The sediment was re-suspended in the buffer, homogenized and centrifuged. The supernatants

were pooled to give 1:10 final dilution. The supernatant was kept in crushed ice for 30 minutes.

To an aliquot of the supernatant, 10% triton X-100 (w/v) was added to a final


VIII

concentration of 1% and mixed with gentle shaking. Cold isotonic buffer was then added to

produce desired dilutions. One ml of the assay mixture contained 10mM potassium phosphate

buffer, 6 mM H2O2 and 0.1ml of the diluted enzyme.

The reaction was started by adding the H2O2 to the assay mixture and monitored the rate

of decrease in absorbance at 240 nm every 15 seconds against a reagent blank devoid of the

substrate. One unit of activity was defined as the amount of enzyme catalytically degrading H2O2

producing a decrease in absorbance of 0.1 per min.

The enzyme concentration was so adjusted to provide an initial absorption between 0.5 to

0.6 units when the steady state was reached. Mean of three determinations was taken for

calculating enzyme activity. Any non-catalatic degradation of H2O2 was controlled by repeating

the experiment in the presence of 1 mM sodiumazide, a potent inhibitor of catalase.85

5) Glutathione peroxidase (EC1.11.19)

Glutathione peroxidase (GSH-Px) activity was assayed in liver homogenates by UV

method.86


IX

Preparation of tissue sample-

About 0.5g of fresh tissue was homogenised in ice cold 0.1 M phosphate buffer pH 7.0

in a motor driven Potter-Elvehjem tissue homogeniser equipped with teflon pestle. The

homogenate was centrifuged at 4o C for 10 mins at 700 x g to sediment nuclei and cell

debris.The sediment was resuspended in the buffer, rehomogenised and recentrifuged. The

supernatants were pooled to a final 1:5 dilution with the homogenising buffer and kept in

crushed ice for 10 min.

GSH- Px Kit

Procedure

Glutathione peroxidase was determined by spectro photometric method of wavelength

340 nm at 37°C temperature. In cuvette 0.02 ml of diluted samples/ distilled water, 1.0 ml of

reagent and 0.04 ml of cumene was added, mixed and the initial absorbance of the sample and

reagent blank was measured after one minute and read again after one and two minutes.

Calculation:-

Glutathione peroxidase concentration was calculated from the following formula:

U/L of Homogenate = 8412 ± ∆A 340 nm / min.

6) Superoxide dismutase (EC 1.15.1.1)

Superoxide dismutase (SOD) was measured by the inhibition of cytochrome C reduction

mediated via superoxide anions generated by xanthine oxidase and monitored at 550 nm. One

unit of of SOD is defined as the amount required inhibiting the reduction of cytochrome C by

50%


X

SOD kit

Kit contained reagents: R1a - mixed substrate of xanthine- 0.05 mol / l and I.N.T. 0.025mmol/ l,

R1b- Buffer- CAPS 40 mmol / l, pH 10.2 & EDTA- 0.94 mmol/l, R2- Xanthine Oxidase- 80

U/L. Working Reagent was prepared by reconstituting one vial of R1a with 20 ml of R1b and

reconstituting R2 with 10 ml of re-distilled water.

Method of determination

SOD was determined by spectrophotometric method of wavelength 505 nm at 37°C

temperature. In cuvette 0.05 ml of diluted samples / standard was added, then 0.05 ml of ransod

diluent, 1.7 ml of working reagent and 0.25 ml of R2 was added and the initial absorbance of the

sample after 30 secs was read and again after 3 minutes.

Calculation:-

A2 - A1 = ∆A / min of Standard or Samples

3

Sample diluents rate = rate of uninhibited reaction = 100 %

% inhibition = 100 - (∆A / min of std) ±100

∆A / min of sample

7) Glucose-6-phosphate dehydrogenase (EC.1.1.1.49)

Glucose 6- phosphate dehydrogenase was assayed in liver sample.The supernatant of the

tissue homogenate prepared in 100 mM Tris buffer pH 8.2, was assayed for glucose-6-phosphate

dehydrogenase (G-6-PDH) by monitoring spectrophotometrically. The amount of NADPH

formed under specified conditions of pH and temperature was measured in a given time.

Contents


XI

R1- Buffer contains Tri Ethanol Amine Buffer 37.7 mmol/l at p H 7 and EDTA- 3.2

mmol/l, R2 NADP- 0.34 mmol/ l, R3- G- 6 phosphate - 0.58 mmol/ l


G-6-PDH was determined by spectro photometric method of wavelength 340 nm at 37°C

temperature. In cuvette 0.25 ml of samples / standard was pippeted, then 1.0 ml of R1 , 0.05ml

of R2 was added and incubated for 10 min at 37°C and 0.25 ml of R3 was added. The initial

absorbance of the sample after 1 min and again after 2 &3 minutes was noted.87

8) Glutathione reductase (EC1.6.4.2)

Glutathione reductase (GSSG-Red) was assayed in liver homogenates by spectro

photometric method. Glutathione reductase has been used in the detection of hepatic and

malignant diseases.

Preparation of tissue sample

The supernatant was prepared as in the case of the assay of GSH-Px.

GSSG Red kit contains the reagents : R1a - Mixed Substrate of Xanthine- 0.05 mol / l and I.N.T

0.025mmol/ l, R1b- Buffer- CAPS 40 mmol/ l, pH 10.2 & EDTA- 0.94 mmol/l

R2- Xanthine Oxidase- 80 U/L


GSSG- Red was determined by spectro photometric method of wavelength 340 nm at

37°C temperature. In cuvette 0.04 ml of samples, 1.0 ml of R1, 0.2ml was added the initial

absorbance of the sample was measured after 1 min and read again after 2, 3, 4 & 5 minutes.

Glutathione reductase activity was calculated as follows: 88


XII

GSSG-Red value in U/gm liver = 4983 ± ∆A340 nm / min ± 20

160

9) Glutathione S-transferase (EC 2.5.1.18)

Glutathione S-transferase (GST) was assayed in liver homogenate by the

spectrophotometric method.89

Preparation of tissue Sample:-

Freshly excised tissues (1 g) were homogenized in ice-cold phosphate buffer, 0.1M (pH

6.5) keeping the tube cool in a beaker containing crushed ice. The homogenate was centrifuged

at 800 x g for 10 min and the residue was re-homogenised and recentrifuged. The pooled

supernatant was made up to 14.0 ml. This supernatant was diluted 10 times using phosphate

buffer that served as the enzyme source for the reaction.

Procedure-

The reaction was started by adding 0.1 ml of GSH (5 mM) and 0.1 ml of 1 mM 1-chloro-

2, 4- dinitrobenzene (CDNB, as the substrate) to 0.1 ml of the enzyme. The reaction mixture

was made up to 1.0 ml with the phosphate buffer. The rate of increase in absorbance at 340 nm

was monitored every 15 sec against a reagent blank devoid of CDNB.

One unit of the activity was defined as the amount of enzyme in mg protein catalysing the

formation of 1mm of product formed per minute under condition of assay. The enzyme

concentrations were so adjusted as to provide an initial absorption between 0.5 to 0.6 absorption

units when the steady state was reached. Mean of determination was taken for calculating

enzyme activity.

GSH solution was flushed with pure nitrogen gas before addition. The reaction mixture


XIII

was thoroughly shaken before the addition of CDNB so that H2O2 formed was fully decomposed

and did not interfere with CDNB.

10. Gamma glutamyltranspeptidase (EC.2.3.3.2)

0.5 g of tissue was homogenized in 10 volumes of 2mM tris HCl buffer pH 7.4 (4˚C)

containing 50m M D- mannitol. To the homogenate 1M CaCl2 solution was added to achieve a

final concentration of 10mM and the mixture was placed in ice with occasional stirring for 10

min. The homogenate was then centrifuged at 800 x g for 15 min. The reaction was initiated by

adding suitable amount of the supernatant to 0.2 ml of 5mML-γ-glutamyl-p-nitroanilide, 0.2ml

of 0.1N glycylglycine (pH 8.0) and 0.6 ml of tris HCl buffer, maintained at 37˚C. The rate of

release of p-nitroanilide was recorded at 410 nm.90

Histopathology reading (HPR)

At the end of six weeks, the tissues were excised and analyzed for histopathology. Micro section

of the organs Viz., Liver, Kidney, Heart, intestine and Brain have been studied for any toxic changes.

Analysis of Data-

The obtained data were analysed using the following statistical methods-

Descriptive statistics

One way ANOVA

Scheffe’s Post Hoc test

Repeated measure ANOVA


XIV

Statistical Methods Applied

Descriptive statistics

The Descriptive procedure displays univariate summary statistics for several variables in

a single table and calculates standardized values (z scores). Variables can be ordered by the size

of their means (in ascending or descending order), alphabetically, or by the order in which you

select the variables (the default).

One-Way ANOVA

The One-Way ANOVA procedure produces a one-way analysis of variance for a

quantitative dependent variable by a single factor (independent) variable. Analysis of variance is

used to test the hypothesis that several means are equal. This technique is an extension of the

two-sample t test.

Scheffe’s Post hoc test

Once it is determined that differences exist among the means, post hoc range

tests and pair-wise multiple comparisons can determine which means differ. Range tests identify

homogeneous subsets of means that are not different from each other. Pair-wise multiple

comparisons test the difference between each pair of means, and yield a matrix where asterisks

indicate significantly different group means at an alpha level of 0.05. Scheffe’s Post hoc test is

one of the widely used post hoc tests in biological sciences.

Repeated measure ANOVA

GLM Repeated Measures analyzes groups of related dependent variables that represent

different measurements of the same attribute. This dialog box lets define one or more within-


XV

subjects factors for use in GLM Repeated Measures. The order in which one specifies within-

subjects factors is important. Each factor constitutes a level within the previous factor.

All the statistical methods were carried out through the SPSS for Windows (version 16.0)


XVI

Fig. 1. Food intake of rats

Values are Mean ± SD of six rats

Fig. 2. Weight gain of rats

Values are Mean ± SD of six rats

Fig 3. Effects of feeding of honey and ghee on relative weight of rat liver

Values are Mean ± SD of six rats where SD ranges from 0.1-0.3


XVII

Fig 4. Effects of feeding of honey and ghee on relative weight of rat kidney


Fig 5. Effects of feeding of honey and ghee on relative weight of rat heart



XVIII

Fig 6. Effects of feeding of honey and ghee on relative weight of rat spleen

Values above are Mean ± SD of six rats where SD ranges from 0.01-0.02

There is no significant change in the weight gain or food intake of rat groups when

compared to control. There is also no significant difference in the relative weights of rat organs

which are fed with honey, ghee, honey mixed ghee and heated- honey and ghee as compared to

control.


XIX

Table. 17. Physical characteristics of honey and ghee samples

Samples Ash value %

Acid Insoluble

Ash %

pH Specific gravity

Organoleptic characteristic

Unprocessed honey 0.15 ± 0.012 0.01± 0.002 4.16 ± 0.4 1.39 ± 0.14 7.5 ± 0.61

Processed honey 0.18 ± 0.013 0.01± 0.002 3.92 ± 0.4 1.41 ± 0.13 7.0 ± 0.59

60 C heated Unprocessed honey

0.14 ± 0.01 0.01± 0.002 4.50 ± 0.5 1.41 ± 0.12 6.5 ± 0.62

60 C heated Processed honey

0.17 ± 0.03 0.01± 0.002 4.18 ± 0.4 1.43 ± 0.13 6.5 ± 0.62

140 C heated Unprocessed honey

0.14 ± 0.03 0.01± 0.002 4.73 ± 0.5 0.88 ± 0.09 5.0 ± 0.49

140 C heated Processed honey

0.18 ± 0.00 0.01± 0.002 4.64 ± 0.4 0.80 ± 0.08 5.0 ± 0.42

Cow’s Ghee 0.07 ± 0.001 0.00± 0.00 4.31 ± 0.4 0.90 ± 0.07 6.0 ± 0.59

Nandini Ghee 0.07 ± 0.002 0.00± 0.00 4.46 ± 0.4 0.89 ± 0.07 7.0 ± 0.65

Honey and ghee 0.09 ± 0.003 0.00± 0.00 4.40 ± 0.4 1.24 ± 0.10 7.0 ± 0.61

F value (8,9) 447.62 - 21.11 223.63 3.47

P value 0.000 - 0.000 0.000 0.041

Values are Mean±SD of six rats as indicated by Scheefe’s posthoc test where = 0.05


XX

Table. 17 depicts the physical properties of honey samples Viz, Ash value, Acid insoluble

ash, pH, Specific gravity and Organoleptic characteristics of honey and ghee samples.

The results showed that-

There was no significant difference in ash value of unprocessed, processed and

heated honey samples. However, the ash value of ghee was significantly less

when compared to honey samples which imply the low amount of mineral

contents when compared to honey.

The acid insoluble ash was within the normal range for all the samples.

pH of honey and ghee samples showed that there was no significant difference

though a mild elevation was noted in heated honey samples when compared to

unheated honey samples.

Specific gravity of honey and ghee samples showed that there was a significant

decrease in specific gravity of 140˚C heated honey samples (unprocessed and

processed honey) and ghee samples (cow’s ghee and Nandini ghee).

The organoleptic characteristics of honey and ghee samples measured by ‘nine

point Hedonic scale’ suggested that the unprocessed honey was rated ‘very good’,

processed honey as ‘good’, 60 C heatedunprocessed and processed honey

samples ‘good above fair’, 140 C heated honey (both processed and unprocessed)

was ‘fair’ on a’ however the ‘taste’ was not included as a part of the sensory

evaluation. Cow’s ghee was also rated as ‘good above fair’, Nandini ghee and

honey mixed ghee were rated as ‘good’.


XXI

Table.18. Physico-chemical characteristics of honey samples

Samples HMF mg/ kg

Browning index Units

Total Phenols

mg / 100g

Flavonoids mg/ gm

Total Antioxidant

Activity %

Unprocessed honey 25.00±3.0 0.089±0.00 1.43±0.15 0.04±0.003 37.80±5.1

Processed honey 35.00±3.8 0.134±0.02 1.43±0.13 0.04±0.003 39.40±6.2

60 C heated Unprocessed honey 68.80±5.9 0.133±0.02 1.48±0.13 0.04±0.003 50.11±5.1

60 C heated Processed honey 72.40±7.3 0.196±0.03 1.47±0.12 0.04±0.003 52.07±6.3

140 C heatedUnprocessed honey 69.70±5.2 0.210±0.02 1.92±0.15 0.05±0.004 64.18±6.5

140 C heated Processed honey 73.80±6.5 0.213±0.03 1.92±0.17 0.05±0.004 67.56±7.1

F value (5,6)* /(8,9) 87.43* 0.621* 193.84 14.38 53.03

P value 0.000 0.691 0.000 0.000 0.000

Values are Mean ±SD of six rats as indicated by Scheefe’s posthoc test where = 0.05

Table 18 shows the chemical characteristics of honey samples Viz., HMF, Browning

index, Total phenolic compounds, Flavonoids and Total antioxidant ativity. The result shows

that-

There was a significant raise in HMF in 60 C and 140 C heated unprocessed and

processed honey samples. There was a significant raise in browning and antioxidant activity in

heated honey samples when compared to unheated samples. There was also a significant raise in

total phenols and flavonoids in 140 C heated honey samples.


XXII

Table. 19. Physico- Chemical characteristics of honey and ghee samples

Sample

PV

m eq.02 / kg

FFA % oleic

acid

TBA mg MDA/

kg

Browning index

units X10-3

Total Phenolic

compounds mg/ 100g

Flavonoids mg/ g X10-3

Total Antioxidant Activity %

Saturated to unsaturated

fatty acid ratio

Cow’s

Ghee 6.25±0.7 3.07±0.3 0.74±0.08 8.0±0.2 0.41±0.04 4.1±0.20 9.8±0.8 65: 34

Nandini

Ghee 6.25±0.7 3.10±0.3 0.75±0.07 9.0±0.7 0.42±0.03 4.2±0.24 9.4±0.90 65: 34

Honey and

ghee 6.10±0.7 3.08±0.3 0.72±0.08 113.0±1.2 0.47±0.03 4.2±0.14 24.78±1.9 65: 34

F value

(2,3) 0.757 1.113 0.865 83.5 193.83 14.38 53.03 -

P value 0.542 0.435 0.505 0.000 0.000 0.000 0.000 -

Values are Mean ±SD of six rats as indicated by Scheefe’s posthoc test where = 0.05

Table 19 depicts the physico- chemical characteristics of honey and ghee samples. The

results suggests that –

There was no statistical significance in peroxide, FFA, TBA value, browning index, total

phenols, flavonoids and total antioxidant activity of the ghee samples and ghee mixed

honey samples.

There was a significant level of browning and antioxidant activity of honey and ghee

mixture.

There was no change in saturated to unsaturated fatty acid ratio by mixing honey and

ghee in equal quantity.


XXIII

Observations in Experimental study Serum study

Table.20. Effects of feeding of honey and ghee on rat SGOT, SGPT, creatinine, total proteins, albumin and globulin in serum

Rat Group SGOT

units/ml SGPT

units/ml Creatinine

mg % Total Protein

gm/dl Albumin

gm/dl Globulin

gm/dl

Control 18.45 ± 1.17 34.95 ± 4.01 0.59 ± 0.12 6.65 ± 0.29 4.67 ± 0.12 2.44 ± 0.35

Honey 18.83 ± 0.93 31.52 ± 1.73 0.58 ± 0.11 7.08 ± 0.21 4.65 ± 0.15 2.44 ± 0.22

Heated honey 19.05 ± 1.93 32.75 ± 3.53 0.56 ± 0.11 8.50 ± 0.32 4.20 ± 0.18 2.30 ± 0.39

Ghee 18.85 ± 1.79 31.60 ± 3.84 0.55 ± 0.15 7.92 ± 0.44 4.08 ± 0.16 2.84 ± 0.46

Honey& ghee 18.65 ± 1.19 34.62 ± 2.70 0.54 ± 0.15 12.68± 1.60 5.18 ± 0.15 4.50 ± 0.68

Heated honey

& ghee 24.11 ± 2.30 32.81 ± 3.11 0.53 ± 0.18 11.89 ± 3.77 5.65 ± 0.30 4.24 ± 0.41

F value (3,15) 4.29 2.44 22.47 11.15 15.39 14.85

P value 0.000 0.005 0.000 0.000 0.000 0.000

Values are Mean ± SD of 6 rats. Values bearing different superscripts in the same column are

significantly different (p < 0.05) . Table 19 shows the following results-

There was no significant difference in SGOT, SGPT and creatinine in the five groups of

rats fed with honey, heated honey, ghee, honey mixed ghee.

SGOT levels in rats fed with heated- honey and ghee is elevated within the normal limits

as there is a trend of elevation.

There was a significant raise (p < 0.05) in total proteins of rats fed with heated honey,

honey mixed with ghee and heated -honey mixed ghee.

There was a significant raise in serum albumin of rats fed with heated honey, ghee, honey

mixed ghee and heated- honey mixed ghee. However, there was a significant change in

globulin only in rats fed with honey mixed ghee and heated -honey mixed ghee.


XXIV

Table.21.Effects of feeding of honey and ghee on rat serum alkaline phosphatase, total cholesterol, triglycerides, HDL, LDL

Rat Group Alkaline

phosphatase units/lt

Total cholesterol

mg/dl

Triglycerides mg/dl

HDL mg/dl

LDL mg/dl

Control 15.09 ± 1.59 173.20 ± 14.5 112.95 ± 13.60 54.40 ± 7.3 96.21 ± 8.17

Honey 16.08 ± 1.71 182.78 ± 17.8 117.40 ± 11.40 54.65 ± 2.6 104.6 ± 8.42

Heated honey 14.01 ± 1.91 182.48 ± 15.4 119.90 ± 09.25 45.92 ± 3.9 112.55± 8.18

Ghee 15.18 ± 1.31 179.11± 15.5 111.24 ± 12 .80 45.93 ± 2.3 101.9 ± 8.01

Honey and ghee 18.18 ± 0.11 178.22 ±17.4 117.11 ± 11.50 48.91 ± 4.2 105.9 ± 8.32

Heated honey and

Ghee 19.20 ± 0.14 179.21 ± 16.1 218.98 ± 14.30 46.83 ± 4.9 88.58 ± 8.13

F value (3, 15) 3.008 17.54 14.32 5.57 0.388

P value 0.001 0.000 0.000 0.000 0.979


significantly different (p < 0.05)

Table 21 shows the effect of feeding of honey, heated honey, ghee honey mixed ghee

and heated- honey mixed ghee for six weeks on rat serum alkaline phosphatase, total cholesterol,

triglycerides, HDL and LDL cholesterol. It shows that-

There was a significant difference (p <0.05) in alkaline phosphatase of rats fed with

honey mixed ghee and heated- honey mixed ghee.

There was no significant difference in total cholesterol in rats which are fed with honey,

heated honey, ghee and honey mixed ghee.

There was a significant raise in serum triglycerides in rats fed with heated- honey and

ghee.

LDL cholesterol in rats fed with heated- honey mixed ghee showed significant decrease

(p<0.05) as compared to other groups.


XXV

Table.22. Effects of feeding of honey and ghee on rat serum urea- B, uric acid, total bilirubin, direct bilirubin and glucose

Rat Group Urea B mg/dl

Uric acid mg/dl

Total Bilirubin mg/dl

Direct Bilirubin

mg/dl

Glucose mg/dl

Control 32.15 ± 1.09 4.50 ± 0.37 0.73 ± 0.16 0.46± 0.03 87.98 ± 9.2

Honey 35.22 ± 2.33 5.03 ± 0.29 0.78 ± 0.12 0.42± 0.08 87.51 ± 8.9

Heated honey 35.25 ± 6.07 4.88 ± 0.31 0.76 ± 0.08 0.41± 0.03 85.35 ± 8.5

Ghee 35.47 ±5.50 5.03 ± 0.23 0.82 ± 0.07 0.47± 0.09 97.89 ± 9.3

Honey& ghee 33.26 ±2.30 6.47 ± 0.29 0.78 ± 0.07 0.41± 0.01 80.78 ± 7.9

Heated honey & ghee 33.63 ±1.74 6.53 ± 0.50 0.79 ± 0.06 0.45± 0.08 89.23 ± 8.3

F value (3,15) 14.09 3.97 8.602 5.92 2.69

P value 0.000 0.000 0.000 0.000 0.002



These results show that-

Urea-B levels in the rats fed with ghee, honey mixed with ghee and heated- honey

mixed ghee are found to be significantly increased as compared to control groups.

There is a significant increase (p < 0.05) in uric acid levels of rats fed with ghee,

honey mixed ghee and heated honey mixed with heated ghee.

There is no significant difference in total bilirubin, direct bilirubin and glucose levels

in any of the rat groups.


XXVI

Liver enzyme study

Table.23. Effects of feeding of honey and ghee on hepatic lipid peroxides

Rat Groups TBARS* HP** CD **

Control 33.96 ± 0.55 11.71 ± 0.09 51.17± 5.2

Honey 27.56 ± 1.95 11.99 ± 1.40 51.48 ± 5.1

Heated honey 29.01 ± 0.90 11.11 ± 1.10 52.46 ± 5.0

Ghee 34.69 ± 1.09 10.15 ± 1.24 56.27 ± 4.2

Honey& ghee 31.05 ± 1.0 10.29 ± 1.49 60.13 ± 5.9

Heated honey & ghee 35.19 ± 3.3 10.21 ± 1.90 72.56 ± 5.6

F value (5,30) 1.033 16.34 7.49

Pvalue 0.416 0.000 0.000

Note- *- n moles/g ; **- μ moles/g

Values are Mean ± SD of 6 rats. Values bearing different superscripts in the same column

are significantly different (p < 0.05). Table shows the effects on hepatic lipid peroxides of rats-

There was a significant decrease in TBARS content in liver of rats fed with honey,

heated honey and honey mixed with ghee.

The hydroperoxides of liver of all the groups of rats showed no significant decrease.

Conjugated dienes of liver of rats fed with heated- honey mixed ghee showed a

significant increase.


XXVII

Table.24. Effects of feeding of honey and ghee on detoxifying enzyme of liver Rat Groups GSSG-Red σ G-6- PDH ω GSTX102 λ GGT γ

Control 33.43±1.3 06.62±1.20 1.28±0.06 1.35±1.27

Honey 33.43±1.8 11.04±1.23 1.13±0.00 1.38±0.19

Heated honey 31.05±2.3 10.23±0.08 1.72±0.01 1.29±0.10

Ghee 32.95±2.3 11.54±0.44 1.45±0.01 1.30±0.18

Honey& ghee 31.89±2.9 12.01±0.33 1.83±0.02 1.89±0.12

Heated honey 30.98±2.8 12.09 ±0.91 1.88±0.02 1.99±0.11

& ghee

F value (5,30) 1.064 182.45 1.42 1.40

P value 0.4 0.000 0.24 0.20

σ- mmoles NADP produced/ min/ mg proteins; ω- ∆A/ min/ mg proteins; λ- nmoles; γ- nmoles p- nitroanilide released/ min/ mg protein



Table 24 describes the effect of feeding of honey, heated honey, ghee honey mixed ghee

and heated-honey mixed ghee on detoxifying enzymes of liver in rats.

There was no significant difference in glutathione reductase of all groups of rats.

There was a significant difference in glutathione- transferase and GGT of rats fed with

honey mixed ghee and heated honey mixed ghee probably due to toxicity.


XXVIII

Table.25. Effects of feeding honey and ghee on liver enzymes

Ω- mmole conjugate formed / min/mg proteins; δ- Units /min/ mg proteins;

-m moles NADP produced / min /mg proteins;

Values are Mean ± SD of 6 rats. Values bearing different superscripts in the same column

are significantly different (p < 0.05).

Table 25 shows the effect of feeding of honey and ghee on liver enzymes of rats.

There was no significant difference in catalase activity.

The activity of SOD, GSH-Px was significantly less in rats fed with ghee, honey

mixed ghee and heated- honey mixed ghee.

Rat Groups Catalase Ω SOD δ GSH-Px

Control 0.169 ± 0.059 14.11± 0.02 127.44 ± 1.27

Honey 0.179 ± 0.077 14.02± 0.06 127.39 ± 1.28

Heated honey 0.125 ± 0.041 14.05± 0.03 120.76 ± 1.21

Ghee 0.119 ± 0.074 14.07± 0.07 128.65 ± 1.21

Honey& ghee 0.132 ± 0.084 13.52 ± 0.16 123.00 ± 1.22

Heated honey & ghee 0.133 ± 0.041 13.88± 0.05 123.01 ± 1.23

F value (5,30) 20.657 6.236 0.634

P value 0.000 0.000 0.676


XXIX

Histopathology reports of the tissues

At the end of six weeks, the tissues were excised and analysed for histopathology. Micro

section of liver of all groups of rats showed normal architecture with hepatic lobules and

hepatocytes arranged in sheets and cords with central veins were normal. Micro section of

kidney of all groups of rats showed normal cortex and medulla with normal glomeruli and

collecting tubules. Micro section of brain in all groups showed normal texture of cerebral cortex

and cerebellum where as heart tissue showed cardiac muscle arranged concentrically. The

intestinal tissues of all groups of rats on micro section showed normal mucosal lining epithelium

with submucosa and serosal layer and also lymphoid aggregates (Payer’s patches).


XXX

Discussion on title-

“Concept of Samskara and Matra Viruddhatwa of Madhu- An Analytical and

Experimental study”

Annat bhootani Jayante | Jatani Annena Vardhante| (Taitriyopanishad)

Man, made out of food is solely dependent on food for his growth and development. It is

the food that determines the state of health in human beings. The changing scenario in lifestyle

also brought up many changes in food and food habits. Delicious cuisines are more popular than

health foods.

In Ayurvedic classics it is clearly mentioned that on proper administration even an acute

poison can act like an effective drug whereas if improperly administered i.e., by improper Matra,

Samskara, Kala, Samyoga etc. an effective drug can also produce ill effects like poison. This

statement is also applicable in relation with Madhu. Acharyas emphasize certain precautions to

be taken during the intake of Madhu, like ushna and samaghruta Madhu are Samskara Viruddha

and Matra Viruddha respectively. Now, in the present time, these instructions are not strictly

followed. Not only this certain purificatory procedures of honey like liquefaction and filtration

needs application of heat to honey, which is contradictory in Ayurvedic perspective. So, this

inclined me to study the changes in Physico-chemical properties of Madhu in relation with the

method of purification as well as the combination of Madhu and Ghrita in equal quantity.

As it is the era of evidence based medicine, analytical and experimental study is carried

to reassess the established truths. Hence the study has been conducted in two different levels- i.e

Analysis of changes that occur in the drug and Experimental study to study its effects on the

body.


XXXI

Discussion on Viruddha ahara

Discussion on Derivation-

Derivation of the term Viruddha i.e “Vi+ Rudhir- Avarane + kta” Avarane means to

cover that is contextually that which curtails the normal functioning of tissues. There are two

kinds of Virodha Viz.,Padarthagata Virodha and Buddhi gata Virodha.

The Padartha gata Virodha deals about all kinds of Viruddha Ahara which curtails the

normal functioning of tissues whereas Buddhigata Virodha deals about that which masks the

attainment of real knowledge.

Discussion on Definition-

The definition ‘Virodha yukta: Viruddha’ means that which is in combination with

Viruddha, it implies that the dravya though not contradictory by its own but produce adverse

effects when they are under the influence of any of the following factors- Desha, Kala, Prakruti,

Samyoga etc., Eg- Ghruta or Madhu are nutritious foods but when taken in equal quantities

produce adverse effects.

Discussion on Lakshanas-

By compiling all the lakshanas it can be put forth as- Viruddha is a dravya when

consumed causes diffusion of deranged Doshas and remains antagonistic to Dhatus and alter its

normal function (Dhatu pratyanaeekataya sthitam). By this, one can infer that dhatugata chikitsa

helps to pacify the effects of Viruddha ahara.

Discussion on Historical review-


XXXII

The concept of Anna rakshana was most prevalent since Veda kala. The concept of

Viruddha ahara was well established only during Samhita kala. When we look into the relevance

of its context in different texts, a close link between Ahita and Viruddha ahara was observed.

Acharya Charaka after ennumerating Atyanta hita and Atyanta ahita dravyas in

‘Yajjahpurusheeya Adhyaya’ (25th chapter) continued to explain in ‘Atreya Bhadrakapyeeya

Adhyaya’ (26th chapter), the Rasapanchakas for better understanding of Hita and Ahita dravyas.

Further in order to put forth the precise factors that produce adverse effects the concept of

Viruddha Ahara might have been explained in detail. Acharya Sushruta also considered

Viruddha as Atyanta Ahita dravya for both Swastha and Rogi. Acharya Vagbhatta mentioned the

concept of Viruddha in Annarakshana Vidhi. Acharya Bhela and Kashyapa also put forth this

concept in the context of Bhojana Vidhi after the explanation of Ahita Ahara.

Thus one can understand the relation of Annarakshana, Bhojana vidhi with Ahita and

Viruddha Ahara.

Discussion on related terms to Viruddha-

Ahita, Mityahara and Viruddha ahara all these produce an antagonistic effect in the body.

Hence these concepts can be approached with a paradigm of Viruddha ahara.


XXXIII

Ahita ahara-

The meaning of ‘Ahita’ is ‘na hita’ and ‘apathya’. There are three terms in relation to

Ahita i.e Ekanta hita, Ekanta Ahita and Hitaahita. The first two terms is addressed to the

Swastha whereas the latter term is to be understood according to the condition i.e Roga avastha.

Among the three, as Viruddha produces adverse effects to the swastha and also plays an

important role in causing diseases, it is placed under Ekanta Ahita. 91 Hence we can infer that all

Viruddha aharas can be understood as Ahita but not vice versa.

Mithya Ahara-

The term ‘Mithya’ means unsuitable ‘Anuchita’92 and non-conducive ‘Asatmya’.93 It is

considered as the root cause of many diseases. At some context Mithya ahara is used to denote

Viruddha Ahara because in general sense Viruddha is also an improper way of administration of

Ahara.94

Samashana-

It is one of the Mithya yoga (improper combination) of Ahara where there is Samyoga of

Pathya and Apathya dravyas. Hence it can be placed under Samyoga Viruddha.

Adhyashana-

When food is taken before the digestion of previous meals, it hampers the Jataragni,

hence can be considered as Agni Viruddha.


XXXIV

Atyashana-

Too much of anything is good for nothing. The quantity when exceeds produce ill effects

hence can be taken as Matra Viruddha.

Vishamashana-

The term Vishama means “Vigato Viruddho Va sama|” It is the deviation from the normal

path i.e., against the rules hence can be placed under Vidhi Viruddha, Kala Viruddha and Matra

Viruddha.

Apathya-

Apathyas are described as two types. One- Nitya apathyas (i.e food to be avoided as a

regular diet) that can be considered as Swabhava Viruddha and the other- Apathyas to a

particular roga (explained in context of each disease) and rogi (explained in context of

purificatory procedures as ayogya Vyaktis) that can be placed under Avastha Viruddha.

Thus all the abnormal usage of foods can be put under the paradigm of Viruddha.

Discussion on types of Viruddha-

All Acharyas except Charaka did not classify Viruddha but just made mentioning of

different Viruddhas with suitable examples. For clear understanding, the different types

mentioned by each Acharyas has been mentioned in the present study. But it can be included in

the eighteen types of Viruddha Ahara of Acharya Charaka-

Rasa, Vipaka and Veerya Viruddha that is explained on the basis of mutual contradiction

by Acharya Sushruta can be brought under Paraspara guna Viruddha.


XXXV

The classification done by Acharya Vagbhatta- Sadrushat, Asadrushat and Ubhaya

Viruddha which is explained on the basis of combination of two dravyas can be taken as

a sub-classification of Samyoga Viruddha.

All types of Viruddha can be included under Swabhava Viruddha as they produce

adverse effects in general, hence there is no need to consider Swabhava Viruddha as a

separate classification.

Thus all types of Viruddha can be brought under the roof of eighteen types of Viruddha and

can be accepted as a standard scale for assessing Viruddha.

Discussion on Samskara Viruddha

Samskara is a Kriya that imbibes new qualities in a substance. Gunantaradhana i.e.,

change in qualities by Samskara is possible only in case of naimittika gunas, not in the case of

samsiddhika gunas ('Yavat dravya bhavina eva guna') e.g Ushnatva of Agni, Chalatva of Vayu,

Snigdha guna of taila etc. cannot be changed by Samskara.

In case of Madhu, Ushna Samskara leads to the production of HMF (Hydroxy Methyl

Furfurladehyde) which produces harmful effects. A recent study also proved that there was some

evidence of carcinogenic activity in female mice on administration of HMF in a dose of 188 &

375 mg/Kg for a period of 2 yrs. It showed an increase in incidence of hepato-cellular adenoma,

lesions in olfactory and respiratory epithelium.95


XXXVI

Discussion on Matra Viruddhatwa

The prime most factors for the maintenance of health are Matravat Ahara i.e intake of

appropriate quantity of food. As Ayurveda believes in Pratipurusha Siddhanta, the quantity of

food intake differs from each individual based on one’s Agnibala and the quantity of medicine

depends on the severity of the disease and the strength of the individual. Hence the quantity of

food can either be Utkrushta (excess), Madhyama (moderate) and Alpa (little). When the

quantity of food exceeds the prescribed limit it produces Ama pradosha vikaras like Visoochika

and Alasaka and in lesser quantity produces Vata prakopa. Viruddhadhyashana ajeernashana

sheelina punaramadosham amavishamityachakshate | When Viruddha ahara i.e., either in

contradictory quantity or contradictory processing are taken it leads to Ama visha.

In case of honey if it is taken in excess quantity, it hampers Agni and produces Ama

(Madhvama) which is a difficult condition to treat. It can also be considered that intake of

Madhu in equal quantity with Jala/Sneha including Ghruta is difficult for digestion hence

produces hazardous effects in the body. Intake of Sama Ghruta Madhu producing early signs of

hepto toxicity is very much evident from the experimental study.

Discussion on the effects of Viruddhasevana-

The effects of intake of incompatible foods can be well understood in the following manner-


XXXVII

Table.26. Showing Viruddha sevanajanya Sroto dushti

S.N Srotas Dushti Karanas Type of Viruddha involved

1 Pranavaha Rooksha sevana

Performing Vyayama in a hungry state.

Vidhi Viruddha

Avastha Viruddha

2 Udakavaha Ushna, Atishushka Ahara

Trushna nigraha

Vidhi Viruddha

Parihara Viruddha

3 Annavaha Atimatra sevana

Akalasevana

Ahita sevana

Matra Viruddha

Kala Viruddha

Vidhi, Upachara -Viruddha

4 Rasavaha Guru, Sheeta & Atisnigdha Ahara

Atimatra

Samashana, Atichintana

Agni Viruddha

Matra Viruddha

Vidhi Viruddha

5 Raktavaha Vidahi, Ushna Annapana Dosha Viruddha

6 Mamsavaha Abhishyanda, Sthoola, Guru bhojana Agni Viruddha

7 Medovaha Medya atibhakshana

Varuni atibhakshana

Vidhi, Parihara Viruddha

Matra Viruddha

8 Asthivaha Vatalasevana Dosha Viruddha

9 Majjavaha Atyabhishyandi, Viruddhashana Vidhi Viruddha

10 Mootravaha Mootritodaka bhakshana Krama Viruddha

11 Purishavaha Ajeerna, Adhyashana

Atyashana

Vidhi Viruddha

Matra Viruddha

12 Swedavaha Sheetoshnakrama sevana Krama Viruddha


XXXVIII

Viruddha Ahara janya rogas can be categorised under different Srotogata, dhatugata

rogas. They are as follows-

Srotogata dushti- Pranavaha – Rajayakshma; Annavaha- Aadhmana, Amavisha,

Grahani; Udakavaha- Trushna; Mamsavaha –Shotha; Mootravaha –Ashmari;

Manovaha- Smrutinasha, Indriyanasha, Chitta nasha, Unmada, Mada

Dhatugata dushti- Rasa dushti- Jwara, Pandu, Klaibhya, Agninasha; Rakta dushti-

Visarpa, Visphotaka, Kushta, Kilasa, Raktapitta, Gulma; Mamsa dushti- Galagraha;

Medo dushti- Prameha; Majja dushti- Moorcha, Majja vyapat, Vata vyadhi; Shukra

dushti- Shandhyam, Santana dosha, Bala nasha, Moodha garbha

Common Viruddha Aharas followed in day-today life-

Today, the era of urbanization has produced increased number of fast foods and

restaurants. The food habits and life style has also been modified according to the profession/

career of an individual. Hence much importance for taste is given but not for health benefits. The

common incidence of Viruddha Ahara seen in day today life is as follows-

Milk shakes (Fruits including sour fruits with milk) causes Vipaka and Veerya

Viruddha

Intake of Fruit juices/ice-cream/curd/honey/alcohol after a hot, spicy meal results

in Upachara Viruddha

Taking bath after the consumption of food is Vidhi Viruddha.

Taking milk/tea/coffee after consuming fish is Parasparaguna Viruddha.

Taking food in improper time, in road places (public) is Vidhi Viruddha.


XXXIX

Food cooked for a week and refrigerated/Preserved/Tinned foods can be

understood as Samskara Viruddha.

Honey cakes and confectionaries done by baking honey is also Samskara

Viruddha.

Honey along with hot water can be taken as Samyoga Viruddha.

Discussion on modern review of Viruddha-

The definition of incompatibility shows that it brings about any chemical change i.e.

gunantaradana and counteract each of the ingredients in compound. This can be

understood as Samyoga and Samskara Viruddha.

Pharmological intolerance is abnormal response of the body for the ingested food. The

response develops due to sensitivity to certain foods, hence can be correlated to Shareera

Viruddha.

Enzymatic intolerance is due to the inability to digest the food. Enzymes are proteins that

catalyze the chemical reaction and they act on specific substrates. In Ayurveda, enzymes

are correlated to Agni therefore this can be understood as Agni Viruddha.

Food allergy or hypersensitivity develops due to the intake of food antagonistic to Koshta

hence can be to Koshta Viruddha.

Lathyrism that develops due to excessive intake of Lathyrus seeds can be interpreted to

be due to Matra Viruddha.

Milk protein allergy is the condition that develops during infancy only for few months. In

this age, cow’s milk passes across the gut wall and provoke immune responses. As this


XL

condition develop during infancy and got subside by two years of age, it can be

categorized into Kala Viruddha and Avastha Viruddha. This condition is common among

children who are weaned early. Introduction of cow’s milk to children who are

unaccustomed to it develops Satmya Viruddha.

Lactose intolerance is because of congenital absence of enzyme lactase. The congenital

deficiency can be understood as Dosha Viruddha.

Ingestion of contaminated food especially in communal feeding causes food poisoning.

The contaminated food can be understood as Vipannarasa bhojana i.e non-nutritious

food and can be correlated as Sampad Viruddha.

Ingestion of food in public is one another cause for food poisoning which is against the

dietetic rules, hence can be taken as Vidhi Virudha.

Food additives are non- nutritious substances which on indiscriminate use pose health

hazards. Generally they are added to improve the commercial value i.e to impart new

qualities (Gunantaradana). So this can be understood as Samskara Viruddha.

Food borne diseases are broadly classified as-

a) Diseases due to natural poisons in food-Eg- Lathyrism (- oxalyl amino alanine) and

Endemic ascites (pyrrolizidine alkaloids) that simulates Swabhava Viruddha.

b) Diseases due to chemical poisoning that may be accidental or adulterated. Eg- Lead in

canned food, Mixture of argemone oil with mustard oil caused epidemic dropsy,

Pesticide residues. This can be taken as Samyoga, Samskara and Sampat Viruddha.


XLI

c) Diseases due to biological agents that cause infections can be equated to Sampat

Viruddha.

d) Diseases that arise due to food allergy can be taken as Koshta and Shareera Viruddha.

Discussion on Madhu-

Gunas of Madhu-

The difference in opinion in gunas of Madhu can be understood as follows-

Guru Vs Laghu guna -

Acharya Sushruta has mentioned Laghu guna because he considered Purana

Madhu as the one fit for consumption.

Guru refers to guna and Laghu refers to paka. Madhu because of its Guru guna is

heavy to digest and owing to its Laghu paka, it holds the urine and feaces and

aggravate Vata dosha.

Tridosha Shamaka Vs Vatala-

Madhu acts as Tridosha shamaka by its dravya and guna Prabhava.(Dalhana)

Madhu acts as Vatala when it is administered alone or in conditions of Kevala

Vata but in conditions of Kapha or Pitta or Kaphapitta samsrishta Vata, it

alleviates Vata when administered with Vatahara dravya.

Ama and Pakwa Madhu-


XLII

In Ayurvedic literature, the details regarding the properties of Ama and Pakwa Madhu

has been explained but identification of the same kinds of honey is unavailable. It can be

understood with the help of modern literature- Ama Madhu is the one which is collected before

the ripening process of honey i. e when the comb is unsealed and the moisture content is more

than the limits. Pakwa Madhu is the honey collected from the sealed honey combs where the

honey has many complex arrays of sugars and beneficial enzymes.

Dushta Madhu-

The honey which has not been strained; contains insects, dead bees, stings and other

parts of bees is poor in quality. The honey that has undergone fermentation due to prolonged

storage; that which is produced in home are all not good for health as they lack in nutrients

present in nectar and pollen grains.

Discussion on Methodology-

The study is conducted in three steps- 1. Literary study 2. Analytical study 3.

Experimental study.

Literary study-

As among all Pramanas, Aptopadesha is foremost method; a thorough literary survey has

been conducted to understand the concept of Viruddha and a detail review of drugs- Madhu and

Ghruta. The literary study has been conducted methodically using tantra yuktis. After this as per

“Simhavalokhana nyaya” i.e proceeding further after retrospective study, analysis of honey and

ghee has been carried which is followed by experimental study.


XLIII

Analytical study-

Before conducting the study proper, quality control measures have been carried out for

honey and ghee. Then to get proper knowledge a comparative study of Unprocessed honey,

Processed honey, Cow’s ghee, Nandini ghee (control drugs) and heated honey, honey mixed

ghee (test drugs) has been carried out. This study was based on the standard parameters to assess

the physical and chemical properties of each sample.

Experimental study-

To understand the pharmaco-kinetics of Viruddha Ahara, experimental study was

conducted on Albino rats. The rats have been observed to study the changes in two different

levels- a) Gross changes b) Minute changes.

To study the gross changes- weekly weight gain and food intake was monitored

periodically. To study the minute changes- serum study and organ study of the rats has been

conducted. By serum study, hepatic and renal functions has been evaluated critically as they are

principal detoxifying centers in the body. By organ study, the gross changes in weight of the

principal organs, enzymatic changes and microscopic changes in tissues has been evaluated by

comparing with control groups.

Discussion on Study design-

Analytical study design-

As the market available honey is heat processed a comparative study of

Unprocessed and Processed honey has been carried out. Then to observe the changes

when heated at different temperatures- the study of 60C heated and 140C heated honey


XLIV

has been conducted. (To interpret ‘Ushnam Madhu Marayati’ as ‘Ushna Samskarat

Madhu Marayati’). Moreover it is believed that the boundary thermal treatment, assumed

as the most severe conditions able to produce a permissible quality loss was found to be

140˚C, during 15s in the transient stage and 30s in the isothermal stage. (Lucero et al, 2004).

Hence honey was heated to 140C for more than 60s to study the possible quality loss.

Acharyas did not mention the type of Ghee in context of Matra Viruddha. When

unspecificied Cow’s ghee is to be considered. Hence the analysis of Cow’s ghee with

market available ghee has been conducted to compare the standard parameters before

analysing the equal mixture of ghee and honey.

Thus a critical analysis of honey and ghee has been carried out.

Experimental study design-

As the study was aimed to find any toxic effects- Acute toxic study was conducted and

the animals were monitored for 48 hours. (Note- Here the dose is taken as prescribed in

Ayurvedic literature; the usual LD50 design for toxic study was not followed). When no

lethality, no behavioural changes and no gross changes were noticed the study has been

extended to sub-chronic study for the period of six weeks.

Discussion on dose fixation-

On thorough literary survey starting from Samhitas to latest text i.e., Indian Materia

medica, there is no mentioning of dosage for honey and ghee for daily intake as food.


XLV

In case of honey, dosage is not mentioned even when administered as medicine. As

honey is a daily recommended diet and the concept of Viruddha explained in relation to Ahara,

the dosage of honey has been taken as 1Pala matra i.e., 48g for normal human.

For ghee there are different dosages for different purposes i.e Two Pala- Hrasva Matra

given for bruhmana in daily diet; Four Pala- Madhyama Matra given for shamana; Six Pala-

Uttama Matra given for the purpose of shodhana.96 In clinical practice, the smallest dose

recommended (Hraseeyasi matra) is 30ml to test the Agnibala of the patient. As the dosage for

daily intake is unspecified, 30ml is fixed as the dosage of ghee for daily intake.

After the fixation of dose of honey and ghee for a normal human, the dose for the rats

was calculated to be 102mg of honey and 64µl of ghee as the daily dose for rats.

To determine the dose for ‘Sama Ghruta Madhu’ the dosage of each drug has to be

brought in one unit. Hence the dose of honey which was in mg was calculated to be 68 µl. As

the recommended dose of ghee is 64 µl, there was a negligible difference between 68 and 64 µl.

Hence the dose was fixed to be 68 µl + 68 µl of honey and ghee.

According to modern literature, ghee on heating produces peroxides that causes oxidative

rancidity, hence a new attempt is made to find the effect of the equal mixture of heated honey

and heated ghee.

Discussion on method of administration-

To study the effects of Viruddha Ahara, the drugs were administered along with the pellet

diets which are the principal food for the albino rats. The powdered pellets were cooked with

water and allowed to cool then the drugs were mixed thoroughly with it in prescribed dosage.


XLVI

Here to get the effectiveness of Nitya Ahara and to get the proper reading it is administered in

this way. In the context of Vicharana Sneha- Snehasya kalpa: Shreshta: Snehakarmaashu

Sadhanaat|| (A.San.Soo.15/16)

Discussion on observation

Analytical study-

Table 1. shows that-

pH of of 140 C heated honey (both processed and unprocessed) was in elevated level

which indicated a decrease in acid content of honey.

Specific gravity of 140 C heated honey samples was reduced considerably to 0.8 showing

the poor quality of honey on heating. According to P.F.A act, A.07.03, pH of honey

should be 3.2- 4.5 and specific gravity should be 1.3 - 1.5. The decrease in specific

gravity might be because of increase in density of honey and reduced moisture.

Ash value of ghee samples was significantly less when compared to honey samples

which may be due to low mineral content in ghee.

Table. 2 shows that

There was a significant raise in HMF of heated honey indicating the quality loss of

honey on heating. White (1994) proposed that HMF level as the only reliable heating/

storage index in honey. 97 The European Union, in order to simplify and update the

legislation in some food sectors, and to follow the new standards of codex on honey,

published the EU Directive 2001/ 110/ CE (L 10/ 47) where in Annex II honey


XLVII

description and chemical composition are listed. The point 3.2 of ALINORM stresses

the effect of overheating on chemical composition changes and quality loss. The most

important difference was in the HMF level. The normal values of HMF must not

exceed 80 mg / kg of honey coming from tropical regions with ambient temperature.98

There was also an increase in browning of heated samples. Browning is a non-

enzymatic Maillard reaction which occurs when the sugars condense with free amino

acids. It is believed that the Maillard Reaction Products (MRPs) are acting as non-

nutrient antioxidants. 99 The antioxidant properties of MRPs have been reported to be

strongly affected by the physico-chemical properties of the system and by the

processing conditions. Polyphenols, ascorbic acid and other carbonyl compounds

even if formed during oxidative reactions can take part in the Maillard reaction

itself.100

There was a significant increase in antioxidant activity of heated honey samples. The

antioxidant activity and brown pigment formation (BPF) increases with temperature

and time and there is a correlation between the antioxidant activity and increased

brown pigment formation.101

The significant raise in phenol and flavonoids compounds suggests their participation

in browning reaction.

Table 3 suggests that

There was no change in peroxide, FFA, TBA, phenols and flavonoid content in

samples of ghee and ghee mixed honey samples.


XLVIII

The significant raise in browning and antioxidant activity of honey mixed ghee

sample suggests that the phenols present in honey imparts brown color on oxidation

and was due to the correlation between antioxidant activity and brown pigment

formation respectively.

Serum study-102

Serum study in rats aimed at finding any hepato or renal toxicity.


SGOT levels in rats fed with heated honey mixed heated ghee is elevated within normal

limas that marks the trend of elevation.

Significant raise in total proteins in the groups of rats fed with heated honey, honey

mixed ghee and heated honey mixed heated ghee may be due to the early signs of liver

cirrhosis.


The significant raise in ALP of rats fed with honey mixed ghee and heated honey mixed

heated ghee may probably be due to the early signs of biliary tract obstruction.

The significant rise in TGL of rats fed with heated honey mixed heated ghee may

possibly be due to hyperlipidemia or nephritis.


The significant rise in Urea and Uric acid levels of rats fed with honey and ghee,

heated honey mixed heated ghee that was probably due to early signs of impaired

renal functions.

Liver enzyme study-

Table 7 suggests the changes in hepatic lipid peroxides-


XLIX

To study the enzymatic changes in organs, liver tissue after excision is homogenized in a

buffer and lipid peroxides and detoxifying enzymes in it was observed. The results can be

interpreted as follows-

There was a significant decrease in TBARS content in liver of rats fed with honey,

heated honey and ghee mixed honey. This decrease could be due to the rich content of

beneficial phyto-constituents including phenols in honey.

The significant increase in Conjugated dienes of rats fed with heated honey mixed

heated ghee might be due to toxic changes.

Table 8 suggests the changes in detoxifying enzymes in liver-

GGT transfers the gamma glutamyl residues to substrate and is used in the synthesis

of glutathione in the body. It is found in liver, kidney, pancreas, intestinal and

prostate gland. GGT is clinically important as it is one of the enzyme profile to detect

any liver diseases. GGT is an indicative of toxicity because significant increase in

hepatic GGT activity is reported in inflammation and the enzyme is also an effective

biochemical marker for immunotoxicity of xenobiotics.

G-6- PDH is an important enzyme for the production of NADPH which is very

essential in preserving the integrity of RBC. The increase in G-6- PDH in all the rat

groups except the control groups suggests the enhanced activity of honey and ghee

samples.

Table 9 suggests the activity of liver enzymes-

The free radical scavenger enzyme system are principally SOD, GSH-Px, and

Catalase. Catalase is the enzyme located in small organelles which has the enormous

capacity to destroy H2O2.The results can be interpreted as follows-


L

There was a significant decrease in SOD and GSH-Px in rats fed with honey mixed

ghee and heated honey mixed heated ghee which may be probably due to toxicity.

Discussion on Histo-pathology report-

As it was a toxicity study, the vital organs like heart and brain; detoxifying organs

like liver, kidney and for any change in the GIT, small intestines were studied

microscopically. There was no evidence of toxicological changes in the tissues that

suggests that the increase in serum parameters was due to the progressive trend of disease

process towards structural changes in tissues. As the pathogenesis was mild it did not

show any histological changes.


LI

CONCLUSION

“Ahara dravyas”, are not contradictory on their own, but produces contradictory effects

under the influence of Desha, Kala, Matra, Samskara etc.

Viruddha dravyas remain antagonistic to the dhatus, and hence dhatu pratyanika chikitsa

has been recommended.

Viruddha Ahara plays an important role in causing number of diseases hence proper

Bhojana Vidhi and Annarakshana Vidhi has to be adopted.

All Viruddha Aharas can act as Ahita but not vice versa.

Ahita Ahara, Mithya Ahara, Samashana, Adhyashana, Atyashana and Vishamashana are

closely related with Viruddha Ahara. Hence all can be grouped under abnormal dietetic

habits.

Pakwa Madhu is the honey obtained after ripening process.

Samskara Viruddhatwa of Madhu was evident from the formation of harmful HMF on

heating.

Matra Viruddhatwa of Madhu was evident from the elevated levels of ALP, GST and

GGT in liver of rats fed with Samaghruta Madhu.

HMF content of processed honey is more than that of unprocessed honey.

Heated honey has more HMF content when compared to unheated samples.

Browning and Antioxidant activity of heated honey samples was increased.


LII

The specific gravity of 140 C heated honey samples was lesser due to the increase in

density on heating honey.

Serum total proteins of rats fed with heated honey (Ushna Madhu), Honey mixed ghee

(Sama Ghruta Madhu) and heated- honey mixed ghee.(Sama / Ushna- Ghruta Madhu)

showed a marginal rise that may be due to the early sign of biliary obstruction.

The elevated levels of serum triglycerides in rats fed with heated- honey mixed ghee was

probably due to hyperlipidemia.

The significant raise in urea and uric acid levels of rats fed with heated honey mixed ghee

might be due to hyperlipidemia or nephritis.

The rich phytoconstituents in honey led to significant reduction in TBARS content in

liver tissues of rats fed with different honey samples.

The significant increase in ALP, GST, Conjugated dienes and GGT of rats fed with

honey mixed ghee (Sama Ghruta Madhu) and heated- honey mixed ghee was probably

due to liver toxicity.

It is evident from the analytical and sub-chronic toxicity study that Ushna Madhu

(heating honey above 140 C more than 15sec) and Sama Ghruta Madhu (equal honey

mixed ghee) produced early signs of hepato-toxicity in albino rats. Thus the Ushna

Samskara of Madhu and Sama Matra of Madhu and Ghruta found to be toxic in albino

rats.


LIII

RECOMMENDATION OF FURTHER STUDY

The same study can be conducted to elicit the chronic toxicity.

To understand Samskara Viruddha of Madhu a comparative study can be conducted

between Nava and Purana Madhu.

A survey study can be conducted to elicit common Viruddha Aharas in day today

practice.


LIV

Summary

The study titled- “Concept of Samskara and Matra Viruddhatwa of Madhu- An

Analytical and Experimental study” has been carried to re-establish the facts and to understand

the pathogenesis on administration of Viruddha Ahara by taking Ushna Madhu and Sama Ghruta

Madhu as example. The study has been conducted in three steps- 1) Conceptual study 2)

Analytical study 3) Experimental study.

In the conceptual study, there are three chapters. First chapter deals with a detail review

on Viruddha including derivation, definition, lakshanas, synonyms, various English equivalents,

related terms to Viruddha, types, diseases produced, its treatment and modern review. It also

includes elaborative notes on Matra and Samskara Viruddha. Second and third chapter gives a

detail review with critical notes on Madhu and Ghruta respectively.

Analytical and Experimental study had been carried out in Biochemistry and Nutrition

division of Defence Food Research Laboratory, Mysore. A critical analysis of honey and ghee

samples were conducted to assess the physical and chemical changes that appear by heating

honey and mixing honey and ghee in equal proportion.

To assess the pharmacokinetics of the same samples an experimental study had been

conducted on Wistar strain Albinorats. One more sample has been newly tried that is an equal

mixture of heated honey and heated ghee. Rats were divided into six groups of six rats

each.Group I: fed with normal pellet diet served as a control; Group-II: fed with honey along

with pellet diet; Group-III: fed with heated honey with pellet diet; Group-IV: fed with ghee with

pellet diet; Group-V: fed with honey mixed ghee with pellet diet; Group-VI: fed with heated-

honey mixed ghee with diet.


LV

The results revealed that on heating honey there was increase in HMF, a carcinogenic agent.

Serum studies revealed the elevated levels of proteins in heated honey group (Ushna Madhu) ;

raised levels of ALP, proteins, of rats fed with honey mixed ghee (Sama Ghruta Madhu);

elevated of Urea and uric acid levels in heated honey mixed ghee group. Liver enzyme studies in

rats revealed elevation of free radicals i.e., GST, Conjugated dienes and GGT.

All these suggests the early signs of hepato toxicity on feeding heated honey (Ushna Madhu)

and equal honey with ghee (Sama Ghruta Madhu) and also heated- honey mixed ghee. Thus the

fact that Ushnam cha Samaghrutam Madhu marayati | has been proved with evidence and

figures (statistics).


LVI

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LXVI

Annexure

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LXXI

NINE- POINT HEDONIC SCALE

Taste panel Evaluation

Product Name:

Name of the Panelist: Date & Time:

S.No Colour Aroma Taste Texture OAA

1

2

3

4

5

Characteristics and Sample code-

9- Excellent; 8- Very Good; 7- Good; 6- Good above fair; 5- Fair;

4-Fair above Poor; 3- Poor; 2- Very Poor; 1- Extremely Poor


LXXII

NTP TECHNICAL REPORT

ON THE

TOXICOLOGY AND CARCINOGENESIS

STUDIES OF 5-(HYDROXYMETHYL)-2-FURFURAL

(CAS NO. 67-47-0)

IN F344/N RATS AND B6C3F1 MICE

(GAVAGE STUDIES)

Scheduled Peer Review Date: February 27-28, 2008

NOTICE

This DRAFT Technical Report is distributed solely for the purpose of predissemination

peer review under the applicable information quality guidelines. It has not been formally

disseminated by the NTP. It does not represent and should not be construed to represent

NTP determination or policy.

NTP TR 554

NIH Publication No. 08-5895

National Toxicology Program

National Institutes of Health

Public Health Service

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES


73

National Toxicology Program Evaluated and interpreted results and reported findings R.D. Irwin, Ph.D., Study Scientist R.A. Herbert, D.V.M., Ph.D., Study Pathologist J.B. Bishop, Ph.D. D.W. Bristol, Ph.D. J.R. Bucher, Ph.D. R.S. Chhabra, Ph.D. P.M. Foster, Ph.D. M.J. Hooth, Ph.D.

A.P. King-Herbert, D.V.M. G.E. Kissling, Ph.D. D.E. Malarkey, D.V.M., Ph.D. J.H. Roycroft, Ph.D. J.M. Sanders, Ph.D. C.S. Smith, Ph.D. G.S. Travlos, D.V.M. N.J. Walker, Ph.D. K.L. Witt, M.S.

Battelle Columbus Operations Conducted studies and evaluated pathology findings M.R. Hejtmancik, Ph.D., Principal Investigator

J.D. Toft, II, D.V.M., M.S. M.J. Ryan, D.V.M., Ph.D. A.W. Singer, D.V.M.

Experimental Pathology Laboratories, Inc. Provided pathology review M.H. Hamlin, II, D.V.M., Principal Investigator

K.J. Cimon, D.V.M., M.S., Ph.D. R.A. Miller, D.V.M., Ph.D. J.C. Peckham, D.V.M., M.S., Ph.D.

Dynamac Corporation Prepared quality assessment audits S. Brecher, Ph.D., Principal Investigator

S. Iyer, B.S. V. Tharakan, D.V.M.

CONTRIBUTORS R.O.W. Sciences, Inc. Provided SMVCE analysis

G.W. Wolfe, Ph.D., Principal Investigator Y. Wang, M.S.

NTP Pathology Working Group Evaluated slides and prepared pathology report on rats (January 26, 2006) A. Suttie, B.V.Sc., Ph.D., Coordinator ILS, Inc. K.J. Cimon, D.V.M., M.S., Ph.D. Experimental Pathology Laboratories, Inc. C. Colleton, D.V.M. Battelle Columbus Operations J. Cullen, V.M.D., Ph.D. North Carolina State University S.A. Elmore, D.V.M., M.S. National Toxicology Program

G.P. Flake, M.D. National Toxicology Program R.A. Herbert, D.V.M., Ph.D. National Toxicology Program G.D. Hill, D.V.M., Ph.D. ILS, Inc. D.E. Malarkey, D.V.M., Ph.D. National Toxicology Program R.A. Miller, D.V.M., Ph.D. Experimental Pathology Laboratories, Inc. J.C. Peckham, D.V.M., M.S., Ph.D. Experimental Pathology Laboratories, Inc. N. Wakamatsu, D.V.M., Ph.D., Observer

NTP Pathology Working Group (continued) Constella Group, Inc. Provided statistical analyses P.W. Crockett, Ph.D., Principal Investigator L.J. Betz, M.S. K.P. McGowan, M.B.A. Biotechnical Services, Inc. Prepared Technical Report S.R. Gunnels, M.A., Principal Investigator

L.M. Harper, B.S. D.C. Serbus, Ph.D. G.E. Simmons, M.A.


LXXIV

Board Draft Evaluated slides and prepared pathology report on mice (January 24, 2006) A. Suttie, B.V.Sc., Ph.D., Coordinator ILS, Inc. C. Colleton, D.V.M. Battelle Columbus Operations S.A. Elmore, D.V.M., M.S. National Toxicology Program R.A. Herbert, D.V.M., Ph.D. National Toxicology Program D.E. Malarkey, D.V.M., Ph.D. National Toxicology Program R.A. Miller, D.V.M., Ph.D


xiii

ABSTRACT

5-(HYDROXYMETHYL)-2-FURFURAL

CAS No. 67-47-0

Chemical Formula: C6H6O3 Molecular Weight: 126.11

Synonyms:

5-Hydroxymethyl-2-formylfuran;

5-(hydroxymethyl)-2-furaldehyde;

5-(hydroxymethyl)-2-furancarbonal;

5-(hydroxymethyl)-2-furancarboxaldehyde (9CI);

Hydroxymethyl furfuraldehyde;

5-oxymethylfurfurole

3-WEEK STUDY IN RATS

Core study groups of five male and five female rats were administered 0, 94, 188, 375, 750, or

1,500 mg 5-(hydroxymethyl)-2-furfural/kg body weight in deionized water by gavage for a total

of 13 doses over a 22-day period. Special study groups of five male and five female rats

designated for neuropathology were administered 0 or 1,500 mg/kg on the same schedule.

Except for one 1,500 mg/kg core study male rat, all rats survived to the end of the study. The

final mean body weight of 1,500 mg/kg males was significantly less than that of the vehicle

control group. No chemical-related histopathologic lesions were observed in core or special

study animals.

3-WEEK STUDY IN MICE

Groups of five male and five female mice were administered 0, 94, 188, 375, 750, or 1,500 mg

5-(hydroxymethyl)-2-furfural/kg body weight in deionized water by gavage for a total of 13

doses over a 22-day period. Three male and three female mice administered 1,500 mg/kg died

before the end of the study. Mean body weights of 1,500 mg/kg males were significantly less

than those of the vehicle control group. Heart weights of 1,500 mg/kg females were significantly

greater than those of the vehicle controls. No chemical-related lesions were observed.


xiv

3-MONTH STUDY IN RATS

Core groups and special study groups (for clinical pathology and neuropathological evaluation)

of 10 male and 10 female rats were administered 0, 94, 188, 375, 750, or 1,500 mg 5-

(hydroxymethyl)-2-furfural/kg body weight in deionized water by gavage for 3 months. One

male and three female rats administered 1,500 mg/kg died before the end of the study; the male

died as a result of gavage trauma. Mean body weights of 750 and 1,500 mg/kg males were

significantly less than those of the vehicle control group. Female rats had elongated estrous

cycles; fewer 750 and 1,500 mg/kg females had regular cycles, and 350, 750, and 1,500 mg/kg

females had a significantly increased probability of extended diestrus. No chemical-related

lesions were observed in core or special study animals.

3-MONTH STUDY IN MICE

Groups of 10 male and 10 female mice were administered 0, 47, 94, 188, 375, or 750 mg 5-

(hydroxymethyl)-2-furfural/kg body weight in deionized water by gavage for 3 months. One 750

mg/kg male and one 375 mg/kg female died before the end of the study; the death of the female

was attributed to ovarian teratoma. The final mean body weight of 750 mg/kg males and body

weight gains of 750 mg/kg males and females were significantly less than those of the vehicle

controls. The incidences of minimal to mild cytoplasmic alteration of the kidney were

significantly increased in males administered 188 mg/kg or greater.

2-YEAR STUDY IN RATS

Groups of 50 male and 50 female rats were administered 0, 188, 375, or 750 mg

5-(hydroxymethyl)-2-furfural/kg body weight in deionized water by gavage for 104 weeks.

Survival of 188 and 750 mg/kg males was greater than that of the vehicle control group. Mean

body weights of dosed groups of males and females were generally similar to those of the vehicle

controls throughout the study.Incidences of olfactory epithelium degeneration were significantly

increased in 750 mg/kg males and 188 and 375 mg/kg females. Incidences of olfactory

epithelium respiratory metaplasia and respiratory epithelium squamous metaplasia were

significantly increased in 750 mg/kg males and females. Incidences of suppurative inflammation


xv

of the nose and chronic active inflammation of the nasolacrimal duct were significantly increased

in 750 mg/kg females.

2-YEAR STUDY IN MICE

Groups of 50 male and 50 female mice were administered 0, 188, 375, or 750 mg 5-

(hydroxymethyl)-2-furfural/kg body weight in deionized water by gavage for 104 weeks.

Survival of 750 mg/kg males and females was significantly less than that of the vehicle control

groups. Mean body weights of 750 mg/kg males were 14% less than those of the vehicle controls

after week 26. Mean body weights of 375 and 750 mg/kg females were 9% and 30% less,

respectively, than those of the vehicle controls after week 36. Beginning in month 8 and

continuing until the end of the study, 750 mg/kg males and females exhibited clinical signs

indicative of neurological effects of 5-(hydroxymethyl)-2-furfural administration. These signs

included decreased exploratory behavior, piloerection, salivation, straub tail, catatonia,

excitation, dyspnea, clonic-tonic seizures, and unconsciousness. Because of the reduced survival

of this group and the presence of the treatment-related clinical signs, groups of mice that

received 750 mg/kg were not included in the evaluation of carcinogenic potential. The incidences

of hepatocellular adenoma were significantly increased in 188 and 375 mg/kg females. In the

nose, the incidences of olfactory epithelium metaplasia, degeneration, and hyaline droplet

accumulation;chronic active inflammation; respiratory epithelium hyaline droplet accumulation;

and hyperplasia, dilatation, and chronic active inflammation of the glands were significantly

increased in 375 and 750 mg/kg males and females. Incidences of olfactory epithelium

hyperplasia were significantly increased in 375 and 750 mg/kg females.

GENETIC TOXICOLOGY

5-(Hydroxymethyl)-2-furfural was tested in two independent bacterial mutagenicity assays. In

the first study, the chemical was weakly mutagenic in Salmonella typhimurium strain TA100 in

the absence of exogenous metabolic activation; no mutagenic activity was detected in TA100

with activation or in strains TA97, TA98, TA102, or TA1535, with or without activation. In the

second study, no mutagenicity was detected, with or without activation, in TA98 or TA100 or

Escherichia coli WP2 uvrA/pKM101. No increases in the frequencies of micronucleated


xvi

erythrocytes were observed in peripheral blood of male or female mice administered

5(hydroxymethyl)-2-furfural by gavage for 3 months.

CONCLUSIONS

Under the conditions of these 2-year gavage studies, there was no evidence of

carcinogenic activity* of 5-(hydroxymethyl)-2-furfural in male or female F344/N rats

administered 188, 375, or 750 mg/kg. There was no evidence of carcinogenic activity of 5-

(hydroxymethyl)-2-furfural in male B6C3F1 mice administered 188 or 375 mg/kg. There was

some evidence of carcinogenic activity of 5-(hydroxymethyl)-2-furfural in female B6C3F1 mice

based on increased incidences of hepatocellular adenoma in the 188 and 375 mg/kg groups.

Administration of 5-(hydroxymethyl)-2-furfural was associated with increased incidences of

lesions of the olfactory and respiratory epithelium of the nose in male and female rats and mice.


xvii

Histo-Pathology Reports


xviii


xix


xx


xxi


xxii

Instruments

Incubator Measuring specific gravity in pyknometer

Hot air oven Digital weighing Balance

Dessicator Water filter


xxiii

Spectrophotometer Homogeniser

Microcentrifuge pH-Meter

Flask shaker Refrigerated centrifuge


xxiv

Each rats put in separate cages Labeling of each cages

Weighing machine for rats Collection of Left over diet

Blood Collection from Orbital sinus Dissection of rats to excise organs

Education

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