Anti-Inflammatory Activity of Babassu Oil and Development ...downloads.hindawi.com/journals/ecam/2017/3647801.pdf · ResearchArticle Anti-Inflammatory Activity of Babassu Oil and

Research ArticleAnti-Inflammatory Activity of Babassu Oil and Development ofa Microemulsion System for Topical Delivery

Mysrayn Y F A Reis1 Simone M dos Santos2 Danielle R Silva2 Maacutercia V Silva3

Maria Tereza S Correia3 Daniela M A Ferraz Navarro4 Geanne K N Santos4

Fernando Hallwass4 Otaacutevio Bianchi5 Alexandre G Silva6 Janaiacutena V Melo7

Alessandra B Mattos8 Rafael M Ximenes6 GiovannaMachado8 and Karina L A Saraiva9

1Programa de Pos-Graduacao em Ciencias Farmaceuticas Universidade Estadual da Paraıba Rua Juvencio Arruda SNBairro Universitario 58429-600 Campina Grande PB Brazil2Programa de Pos-Graduacao em Ciencias Farmaceuticas Universidade Federal de Pernambuco Av Prof Moraes RegoNo 1235 Cidade Universitaria 50670-901 Recife PE Brazil3Departamento de Bioquımica Universidade Federal de Pernambuco Av Prof Moraes Rego No 1235 Cidade Universitaria50670-901 Recife PE Brazil4Departamento de Quımica Fundamental Universidade Federal de Pernambuco Av Prof Moraes Rego No 1235Cidade Universitaria 50670-901 Recife PE Brazil5Programas de Pos-Graduacao em Engenharia e Ciencia dos Materiais e Ciencias da Saude Universidade de Caxias do SulRua Francisco Getulio Vargas No 1130 Petropolis 95070-560 Caxias do Sul RS Brazil6Departamento de Antibioticos Universidade Federal de Pernambuco Av Prof Moraes Rego No 1235 Cidade Universitaria50670-901 Recife PE Brazil7Faculdade Boa Viagem DeVry Av Jean Emile Favre No 422 Imbiribeira 61200-060 Recife PE Brazil8Laboratorio de Microscopia e Microanalise Centro de Tecnologias Estrategicas do Nordeste Av Prof Luıs Freire No 1Cidade Universitaria 50740-540 Recife PE Brazil9Nucleo de Plataformas Tecnologicas Instituto Aggeu Magalhaes FIOCRUZ Av Prof Moraes Rego SN Cidade Universitaria50740-465 Recife PE Brazil

Correspondence should be addressed to Rafael M Ximenes rafaelximenesufpebr

Received 18 September 2017 Revised 20 November 2017 Accepted 29 November 2017 Published 21 December 2017

Academic Editor Rocıo De la Puerta

Copyright copy 2017 Mysrayn Y F A Reis et al This is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properlycited

Babassu oil extraction is the main income source in nut breakers communities in northeast of Brazil Among these communitiesbabassu oil is used for cooking but also medically to treat skin wounds and inflammation and vulvovaginitis This study aimed toevaluate the anti-inflammatory activity of babassu oil and develop a microemulsion system with babassu oil for topical deliveryTopical anti-inflammatory activity was evaluated in mice ear edema using PMA arachidonic acid ethyl phenylpropiolate phenoland capsaicin as phlogistic agents A microemulsion system was successfully developed using a Span 80Kolliphor EL ratio of6 4 as the surfactant system (S) propylene glycol andwater (3 1) as the aqueous phase (A) and babassu oil as the oil phase (O) andanalyzed through conductivity SAXS DSC TEM and rheological assays Babassu oil and lauric acid showed anti-inflammatoryactivity in mice ear edema through inhibition of eicosanoid pathway and bioactive amines The developed formulation (39A 122 O and 488 S) was classified as a bicontinuous to ow transition microemulsion that showed a Newtonian profileThe topical anti-inflammatory activity of microemulsified babassu oil was markedly increased A new delivery system of babassumicroemulsion droplet clusters was designed to enhance the therapeutic efficacy of vegetable oil

1 Introduction

Babassu palm tree (Attalea speciosa Mart ex Spreng Are-caceae) is commonly found in North and Northeast regions

of Brazil mainly in the phytogeographic domain of ldquoMatados Cocaisrdquo (Port Lit Palm trees forest) In its areas ofoccurrence it represents the main income source for about300000 people in nut breakers communities in northeast

HindawiEvidence-Based Complementary and Alternative MedicineVolume 2017 Article ID 3647801 14 pageshttpsdoiorg10115520173647801

2 Evidence-Based Complementary and Alternative Medicine

region of Brazil most of them in state of Maranhao but alsoin Piauı and inChapada doAraripeCEGathering of fruits toproduce oil or to sell the nuts is activities predominantlymadeby women who are called ldquoquebradeiras de coco babacurdquo (PorLit Bassabu nut breaker women) [1ndash3] These communitiesdeveloped autochthonous knowledge about babassu usesas source of human and animal food utensils and toolsfuel construction and soil fertilizers and as cosmetic andmedicines [3ndash6]

Among the parts used for medicinal purposes leavesroots and fruits should be highlighted Leaves and roots areused as tea for pain and wound healing while fruits are usedin a much bigger scale mesocarp floor and milk are usedfor the treatment of gastritis hepatitis osteoporosis skinwounds and leukorrhea liquid albumen is used as eyedropsto treat conjunctivitis and the seed oil is used as laxativevermifuge and anti-inflammatory and for the treatment ofmyiasis mycosis skin wounds hemorrhoids leukorrhea andfemale genital inflammation and spider bites [3ndash5 7]

Vegetable oils such as olive palm and coconut oils arewell known for their anti-inflammatory properties Olive oilis rich in anti-inflammatory phenolics like oleocanthal andoleuropein glycosides [8 9] while the effect of palm oil isattributed to its high tocotrienol content [10] Like babassuoil coconut oil is poor in phenolics and tocopherols Its anti-inflammatory effect is assigned to the presence of lauric acidand glyceryl laurates [11]

The availability of babassu oil and its easy handling haveenabled development of several types of formulations toenhance the therapeutic efficacy of the biologically activecomponents Targeted delivery can be achieved by apply-ing pharmaceutical nanotechnology which is based on thesynthesis application and characterization of nanoscaletherapeutic systems to provide a more controlled drugrelease In this context nanostructured systems for examplemicroemulsions containing babassu oil may act as new andpotentially efficient therapies for benign prostatic hyperpla-sia due to their antiproliferative and apoptotic effects [12]and improve human immune system function by increas-ing superoxide anion release phagocytosis of mononuclearphagocytes and antimicrobial activities [13]

Microemulsion is a system of two immiscible fluids thatis stabilized by an interfacial film of surfactants It offersthe advantages of spontaneous formation thermodynamicstability manufacturing simplicity solubilization capacity oflipophilic andor hydrophilic solutes a large area per volumeratio for mass transfer and the potential for permeationenhancement [14ndash17] Thus the aim of this work was toevaluate the topical anti-inflammatory activity of babassu oilobtained from Chapada do Araripe in Ceara (Brazil) andsynthesize and characterize a microemulsion system aimingto enhance babassu oil topical anti-inflammatory activity

2 Material and Methods

21 Chemicals Phorbol 12-myristate 13-acetate (PMA) ar-achidonic acid (AA) ethyl phenylpropiolate (EPP) phenoland capsaicin were purchased from Sigma Chemical Co(St Louis MO USA) The surfactants Span 80 (sorbitan

mono-oleate) and Tween 80 (polyoxyethylene sorbitanmonooleate) ethanol 999 deuterated CDCl3 and Supel-co 37 Component FAME Mix were also obtained fromSigma Chemical Co Kolliphor EL (polyoxyl castor oil) wasfrom BASF SE (Ludwigshafen Germany) and propyleneglycol PA was purchased from Neon Comercial Ltda (SaoPaulo Brazil)

22 Plant Material and Oil Extraction Babassu fruits werecollected from one palm tree in Araripe located in the Stateof Ceara (Brazil) in July 2013 Botanical authentication wasconducted by Olıvia O Cano and a voucher specimen wasdeposited at the Herbarium of the Agronomic Institute ofPernambuco under number 90472

The oil was extracted by a well-established method thatis practiced by farmers in forested areas After harvestingthe seeds were ground with a stone grinder and the obtainedpaste was mixed well and allowed to stand overnight On thenext day cold water was added to the paste and the upperpart was collected and placed on a fire to heat it until itstarted boiling Subsequently the liquid phase was separatedfrom the oil meal using a tissue and heated until the watercompletely evaporated Finally the oil was filtered and used[18]

23 Physicochemical Characterization and Fatty Acid ProfileTraditionally extracted babassu oil was immediately analysedfor some physicochemical properties as described by AdolfoLutz Institute [19] as follows relative density was determinedin a 10mL glass hydrometer (pycnometer) at 25∘C refractiveindex at 40∘C was performed using a Abbe refractometerperoxide values were calculated from the iodine release frompotassium iodide and expressed as milliequivalents of activeoxygenkg of oil rancidity (lipid oxidation) was determinedby Kreis reaction using phloroglucinol in acid medium acidvalues were expressed as milligrams of KOH required toneutralize the free acids of 1 g of oil which was determinedby titration in methanolic solution

The transesterification procedure of the oil was realized inaccord with Metcalfe et al [20] The oil (150mg) was mixedwith 40mL of NaOH in 050M methanol and incubatedat 100∘C for 5 minutes Next 50mL solution of BF3 (12)in methanol was added in the mixture and was heated for2 minutes After cooling to room temperature 50mL of asaturated sodium chloride solution was added The mixturewas transferred to a separation funnel containing 200mL ofpetroleum ether and vigorously stirred After a period of restthe aqueous phase was discarded and the ether phase wasfiltered The solvent was evaporated in a rotary evaporatorat 60∘C and the residual solvent was removed with nitrogenflow Methyl esters were solubilized in n-heptane beforeinjection into the gas chromatographer [21]

The samples were analyzed using an Agilent Technolo-gies (Palo Alto CA USA) 5975C single quadrupole GC-MS equipped with a nonpolar HP-5MS (Agilent) fusedsilica capillary column (30m times 025mm id film thickness025mm) The oven was initially held at 150∘C for 2minincreased to 230∘C by 5∘Cmin (held for 7min) and finallyincreased to 260∘C by 4∘Cmin The final temperature was

Evidence-Based Complementary and Alternative Medicine 3

maintained for 75min The carrier gas was helium suppliedat a constant flow of 1mLmin and the splitsplitless injectorwas maintained at 230∘C The applied ionization potentialwas 70 eV the scan range was from 35 to 450119898119911 withscan rate of 05 scanss A standard fatty acid methyl estermixture (Supelco 37 Component FAMEMix USA) was usedto identify the fatty acid methyl esters Fatty acid data wereexpressed as percentage of total peak area

24 NMR Analysis The extracted oil (approximately 50mg)was dissolved in 06mL of CDCl3 and placed in a 5mmNMRtube The NMR analyses were performed on a Varian 400-VNMRS (Agilent Technologies CA USA) at 26∘C operatingat the frequencies of 39974 and 10051MHz for 1H and13C respectively 1H NMR spectra were recorded with 25 sacquisition times sweep widths of 64 kHz 45∘ pulse anglesand 1 s delay times 13C1H NMR spectra were recordedwith 304 s acquisition times sweep widths of 215 kHz 45∘pulse angles and 1 s delay times For processing 13C spectrazero filling and line broadening of 1Hz were applied priorto Fourier transformation The chemical shift scale wasindirectly referred to tetramethylsilane (TMS) by the residualCHCl3 signal at 726 ppm for 1H spectra and the signal at770 ppm for 13C spectra

25 Topical Anti-Inflammatory Activity

251 Animals Male Swiss and BALBcmice (25ndash30 g 119899 = 6)were provided by the Animal Facility of Federal Universityof Pernambuco The animals were housed and kept in aroom with controlled temperature (23 plusmn 2∘C) under a 1212 hlightdark cycle with food and water ad libitum Experimentswere conducted according to the Guide for the Care andUse of Laboratory Animals from the US Department ofHealth andHuman ServicesThe project had been previouslyapproved by the Animalrsquos Ethics Committee of FederalUniversity of Pernambuco (number 230760398872014-51)

252 Ear EdemaMeasurement For evaluation of ear weightmice were euthanized and 6mm diameter samples weretaken from both ears using a biopsy punch (Richter Brazil)Each biopsy was weighed on a semi-micro analytical bal-ance (AUW-D 220 Shimadzu Japan) Ear edema (EE) wasexpressed as the increase in ear sample weight using thefollowing formula

EE (mg) = 119908RE minus 119908LE (1)

where 119908RE is the weight obtained from the right ear sample(inflamed ear) and 119908LE is the weight obtained from the leftear sample (noninflamed ear)

For all treatments animals were anesthetized with 1halothane Right ears were then challenged with differentphlogistic agents diluted in acetone (20120583L) Babassu oil (13 and 10 120583Lear) was applied topically in 20120583L acetone whilemicroemulsionwas appliedwith nodilutionDexamethasoneor indomethacin (01 and 05mgear resp) was used topicallyas a positive control Ruthenium red (3mgkg sc) was usedas a positive control for capsaicin-induced ear edema

253 PMA-Induced Ear Edema Ear edema was induced bytopical application of 20120583L of PMA (25 120583gear) in acetoneon both sides of the right ear Left ears received onlyacetone as a control After 15min 10 120583L of babassu oil (100and 122) babassu microemulsion microemulsion vehicle(488 surfactants 39 aqueous phase and 122 water)05 dexamethasone or acetone were applied to both sidesof the right and left ears Six hours after PMA administrationmice were euthanized for ear edema measurement [22 23]To avoid misinterpretation of the results due to a possiblebarrier effect of babassu oil on ear surfaces an oral treatmentwith babassu oil was also evaluated against PMA-induced earedema Shortly mice were treated with babassu oil (100 300and 1000mgkg) or indomethacin (10mgkg po) 60minbefore PMA administration [23]

254 Investigation of Babassu Oil Mode of Action on EarEdema For elucidation of the mechanisms underlying thetopical anti-inflammatory activity of babassu oil on PMA-induced mice ear edema different phlogistic agents wereused to induce ear edema arachidonic acid (2mgear) ethylphenylpropiolate 5 (20120583Lear) phenol 10 (20 120583Lear)and capsaicin (025mgear) Immediately after acetone evap-oration babassu oil (10 120583Lear) or indomethacin (05mgear)dexamethasone (01mgear) were topically applied onboth sides of the right ear while the left ear received 20120583Lof the vehicle Ruthenium red (3mgkg sc) was used aspositive control for capsaicin-induced ear edema and it wasadministered 30min before capsaicin

After 1 h mice were euthanized for ear edema measure-ment Mice challenged with capsaicin were euthanized after30min of exposure AA-induced ear edema was performedin BALBc mice [23ndash25]

26 Preparation of Emulsions A series of oil-in-water emul-sions with HLB values ranging from 45 to 15 was preparedwith the surfactants Span 80 and Tween 80 at a 2 total blendconcentration wv 93 water and 5 babassu oil using theGriffin equation [26] The aqueous phase (water plus Tween80) and the oil phase (oil plus Span 80)were heated to 70plusmn5∘Cseparately Both phases were mixed by the inversion methodwith mechanical stirring (9000 rpm) for 5min in ULTRA-TURRAX homogenizer equipment (UniqueDES500 SaoPaulo Brazil) The samples were characterized 24 h afterpreparation

27 Droplet Size Analysis by Dynamic Light Scattering (DLS)The droplet size distribution of the dispersed phase of theemulsions was determined by DLS using the Nanotrac Waveequipment (Microtrac Inc Montgomeryville PA USA) withmeasurement capability from 08 to 6500 nanometers Thedata were calculated using the manufacturerrsquos software Eachemulsion was diluted with aqueous propylene glycol (1 100)and analyzed in triplicate

28 Turbidimetric Method The emulsions were diluted 1 25with aqueous propylene glycol and the percentage of trans-mission ( 119879) was measured at 600 nm using a spectropho-tometer (UV-mini 1240 Shimadzu Japan) With the blank


control set at 100 transmission the turbidity of the dilutedemulsion was calculated as follows turbidity = 100 minus119879 [27]The reported results were the average of three determina-tions

29 Construction of the Pseudo-Ternary Phase DiagramBabassu oil Span 80 andKolliphor ELwere selected as the oilphase and surfactants respectively Surfactant mixtures weretested at a ratio of 6 4 (ww) as defined by the Griffin equa-tion The pseudo-ternary phase diagrams were constructedusing the water titration method at room temperature andthe results were plotted using Software Origin Pro 80For each phase diagram mixtures of surfactants and oilwere prepared at weight ratios of 1 9 2 8 3 7 4 6 5 56 4 7 3 8 2 and 9 1 (ww) These mixtures were titrateddropwise with water-propylene glycol (1 3) under mechanicstirring After equilibration the systemswere visually charac-terized Single-phase transparent mixtures were designatedas microemulsions [28]

After data analyses of the pseudo-ternary phase diagramsthe microemulsion system was prepared by mixing babassuoil (122) with the surfactants (488) before adding theaqueous phase (39) undermagnetic stirring After two daysthe system was evaluated by complementary techniques asdescribed below

210 Characterization of the Microemulsion

2101 Electrical Conductivity (EC) EC was evaluated usinga digital conductivity meter (mCA 150P Tecnopon SaoPaulo Brazil) previously calibratedwith a calibration solution(1469 120583Scm)The electrode was inserted directly into 10mLof the formulation The analysis was performed in triplicateat a temperature of 25 plusmn 05∘C

2102 TEM The sample was dropped onto a 300 meshcarbon-coated cooper grid and negatively stained with 2phosphotungstic acidThe grid was analyzed in an FEI TecnaiSpirit Biotwin G2 microscope (Hillsboro Oregon USA)operated with 80KV of accelerating voltage

2103 SAXS SAXS experiments were performed on theSAXS1 beamline of the Brazilian Synchrotron Light Labo-ratory (LNLS Sao Paulo Brazil) monitored with a photo-multiplier and detected on a Pilatus detector (300k Dectris)positioned at 836mm that generated scattering wave vectors(119902) from 013 to 25 nmminus1 The wavelength of the incidentX-ray beam (120582) was 0155 nmminus1 Background and parasiticscattering were determined by separate measurements on anempty holder and subtracted from the scattering data of thesample The X-ray scattering was experimentally determinedas a function of the scattering vector 119902 whose modulus isgiven as follows

119902 =(4120587 sen 120579)120582 (2)

where 120579 is half of the scattering angle (2120579)

Table 1 Physicochemical parameters of babassu oil from Chapadado Araripe Brazil

Physicochemicalparameters

Babassu oil(unrefined)

Reference value(refined oil)

Relative density(gmL) 09210 09140ndash09170

Refractive index at40∘C 1458 1448ndash1451

Acid value(mgKOHg) 013 Max 4

Peroxide value(meqkg) nd Max 15

Rancidity Absent Absentnd not detected

2104 DSC DSC analysis was performed using a ShimadzuDSC 50 (Kyoto Japan) with 9-10mg of sample under a nitro-gen atmosphere with a flow rate of 50mLmin The meltingtemperature and enthalpy were calibrated with indium andzinc standards The samples were analyzed from minus50 to 60∘Cwith a heating rate of 10∘Cmin

2105 Rheological Analysis The rheological behavior of theformulation was investigated on an Anton Paar PhysicaMCR502 oscillatory rheometer (Ashland VA USA) withcone and plate geometry (50mm diameter)The gap betweenthe cone and plate was set at 005mm The measurementswere taken at 25∘C over a shear rate (120574) range from 01 to1000s

211 Statistical Analysis Results were expressed as mean plusmnSEM and normal distributions were checked by Shapiro-Wilktest Then data were analyzed by one-way ANOVA followedby Tukeyrsquos test using GraphPad Prism 50 with significanceset at 119901 lt 005

3 Results and Discussion

31 Physicochemical Characterization GC-MS and NMRAnalyses of Babassu Oil Physicochemical parameters ofbabassu oil and reference values are placed in Table 1while fatty acid composition obtained by GC-MS is showedin Table 2 The main fatty acids found were dodecanoicacid (lauric acid) (4078) 9-octadecenoic acid (oleic acid)(2135) tetradecanoic acid (myristic acid) (2005) andhexadecanoic acid (palmitic acid) (1226) (see SM Figure2) These data are in accordance with previous publisheddata and with reference standards of Brazilian regulatoryagencies [29ndash31] Differences in relative density and refractiveindex from reference values can be explained since referencevalues are derived from refined oils Nevertheless thoseparameters indicative of oil degradation as acidity rancidityand peroxide value are well below the established limits ofquality1H NMR and 13C NMR analyses were performed to

characterize the components of the sample and determine thedistribution of fatty acids in the glycerol backbone of babassu


Table 2 Fatty acid composition of babassu oil from Chapada doAraripe Brazil

Skeleton Compound Area () plusmn St DevC120 Dodecanoic acid 4078 plusmn 156C130 Tridecanoic acid 003 plusmn 001C140 Tetradecanoic acid 2005 plusmn 027C160 Hexadecanoic acid 1226 plusmn 059C182n6c (ZZ)-912-Octadecadienoic acid 239 plusmn 029C181n9c (Z)-9-Octadecenoic acid 2135 plusmn 036C180 Octadecanoic acid 264 plusmn 009

Table 3 1H and 13C NMR chemical shift (120575) data of babassu oil inCDCl3

Hydrogen 120575 (ppm) Carbon 120575 (ppm)CH=CH 533 C=O 1732 and 1728CH-O 522 CH=CH 1299 and 1296CH2-O 429 CH2-O 621CH2-O 415 CH-O 688C=C-CH2-C=C 274 C=C-CH2-C=C 272CH2-C=O 230 CH2-C=O 341CH2-C=C 201 CH2-C=C 318CH2-CH2-C=O 161 CH2-CH2-C=O 248(CH2)119899 128 CH2-CH3 226CH3 087 (CH2)119899 297ndash288

CH3 140

oil (Table 3) (see SM Figures 3 and 4) NMR signals wereassigned according to the literature [31ndash35] There are someadvantages to using NMR spectroscopy to study fatty acidcompositions (a) the ability to directly operate on the oilsample without any chemical manipulation and (b) NMR isthe only direct instrumental method by which the positionaldistribution of fatty acids on glycerol can be specificallyidentified [36]

The signals at 533 201 and 274 ppm were assigned tothe olefinic hydrogens the protons attached to the allyliccarbons and the protons attached to the bis-allylic carbonsrespectively (Table 3) These signals revealed the presence ofunsaturated fatty acids such as oleic acid in the oil mixtureMoreover the last signal indicated that a small amount oflinoleic acid was present Linolenic acid was not detected inthis mixture because no signal was observed at 098 ppmwhich corresponds to the terminal methyl group of the fattyacid [32] These results agree with those obtained in the GC-MS spectra (Table 2) The full identification of every fattyacid component was not possible due to signal overlap in thespectra13C NMR spectra provide information regarding the

positional isomerism of fatty acids in the glycerol back-bones of triacylglycerols (TAG) diacylglycerols (DAG) andmonoacylglycerols (MAG) [35] When analyzing the spectralregion of the carbonyl group only two signals at 1732 and1728 ppm were observed (Table 3) This pattern indicatedthat the oilmixture includes only TAG a fatty acid distributed

at the sn-13 and sn-2 positions Moreover the spectralregion of the acyl carbons between 60 and 73 ppm showsonly two peaks (6885 and 6207 ppm) (Table 3) supportingthat the sample only contains TAG isomers The presenceof unsaturated fatty acids that was observed in the 1Hspectrum was confirmed in the 13C spectrum by the signalsbetween 12800 and 13000 ppm This region is characteristicof ethylenic carbons The chemical shifts at 12966 and12999 ppm indicated that the unsaturated fatty acids in theoil mixture were majority oleic acid in agreement with theGC-MS results

32 Topical Anti-Inflammatory Activity PMA-induced earedema is a useful model for screening topical anti-in-flammatory compounds andor plant extracts that act ata variety of levels Skin inflammation induced by topicalPMA administration is mediated trough protein kinaseC (PKC) activation of NF-120581B with production of tumornecrosis factor-120572 (TNF-120572) cyclooxygenase-2 (COX-2) andprostaglandin E2 (PGE2) [37ndash39] PMA given to mouse earsinduces mast cell infiltration with release of mediators thatincrease vascular permeability and promote neutrophil influx[40 41] Anti-inflammatory efficacy of evaluated compoundsimplies mainly interference of arachidonic acid metabolismsince phospholipase A2 (PLA2) COX and lipoxygenase(LOX) inhibitors have been shown to inhibit PMA-inducedinflammation [23 42]

Babassu oil (3 and 10 120583Lear) was able to inhibit PMA-induced ear edema in 191 (119901 lt 005) and 541 (119901 lt 0001)respectively Lauric acid (4mgear) the major fatty acid ofbabassu oil showed 903 of inhibition (119901 lt 0001) whiledexamethasone inhibited the ear edema in 797 (119901 lt 0001)(Figure 1(a)) To avoid misinterpretation of the results dueto a possible barrier effect of babassu oil on ear surfaces anoral treatment with babassu oil was also evaluated againstPMA-induced ear edema When given orally babassu oil(100 300 and 1000mgkg) showed 235 (119901 lt 005) 397 and519 of edema inhibition (119901 lt 0001) while indomethacin(10mgkg) inhibited the edema in 620 (119901 lt 0001)(Figure 1(b))

It is worth to comment that the high content of lauric acidin babassu oil defines the importance of this study since itsantibacterial and anti-inflammatory activities are describedin the literature [43ndash45] through inhibition of MAPK path-way and subsequent NF-120581B activation which are prominentcontributors to the production of proinflammatory cytokinesand chronic inflammatory responses [46] including in PMA-induced ear edema model After topical administrationlauric acid penetrates the skin and accumulates in the dermis[47]

To further elucidate the topical anti-inflammatory activ-ity of babassu oil different phlogistic agents-induced earedema was performed Arachidonic acid-induced ear edemais a useful tool to identify compounds that interfere in eicos-anoid pathway such as COX and LOX inhibitors Howeverthis model is not sensitive to PLA2 inhibitors such as glu-cocorticoids [23 48ndash50] Here both babassu oil (10 120583Lear)and lauric acid (4mgear) inhibited AA-induced ear edema


10

AA

B

C

DD

1 3 01 4

Dexa LA

minus

Babassu oil

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)

(a)

101000100 300

A

B

CBC

C

minus

Babassu oil

0

5

10

15

Ear e

dem

a (m

g)

PMA (25gear)

Indo

(b)

Figure 1 (a) Topical and (b) systemic anti-inflammatory activity of babassu oil in PMA-induced ear edema Results are expressed as themeanplusmn SEM and analyzed by ANOVA followed by Tukeyrsquos test with 119901 set at 005 Different letters show statistical differences between groups (119901 lt005) (minus) Negative control (acetone) babassu oil (120583Lear in (a)) and (mgkg po in (b)) (LA) lauric acid (mgear) (Dexa) dexamethasone(mgear) and (Indo) indomethacin (mgkg po)

by 785 and 616 respectively (119901 lt 0001) Indomethacin(05mgear) a nonselective inhibitor of COX-12 decreasedthe ear edema by 654 (119901 lt 0001) (Figure 2(a)) Short-chain fatty acids (C10ndashC12) which include lauric acid arereported as mid-range inhibitors of COX-1 (about 50ndash60of inhibition) and COX-2 (about 25ndash30 inhibition) in vitro[51]

Barbosa et al [52] demonstrated that babassu oil de-creased ischemia-induced macromolecular leakage in post-capillary venules and inhibited histamine-induced microvas-cular permeability increase in hamster cheek pouch Toverify if the same occurs in skin instead of mucosa ethylphenylpropiolate-induced ear edema was performed EPPadministration immediately increases vascular permeabilitywith blood flow increasing somewhat slowly Histamineserotonin (5-HT) kinins and prostaglandins combinedactions on vascular permeability seemed to be involved inearly EPP response [53] Babassu oil (10 120583Lear) and lauricacid (4mgear) inhibited EPP-induced ear edema by 820and 718 (119901 lt 0001) respectively in accordance with theresults of Barbosa et al [52] Dexamethasone (01mgear)used as positive control inhibited edema formation by 656(119901 lt 0001) (Figure 2(b))

Phenol-induced skin inflammation is very like humancontact dermatitis processes [54] Phenol disrupts the plasmamembranes of keratinocytes in the skin resulting in releaseof preformed cytokines (IL-1120572 IL-8 and TNF-120572) Thesecytokines stimulate the production of reactive oxygen species(ROS) and AA metabolites amplifying the inflammatoryprocess [54ndash56] Herein babassu oil (10 120583Lear) and lauricacid (4mgear) inhibited phenol-induced ear edema by898 and 900 (119901 lt 0001) respectively Dexamethasone(01mgear) used as positive control due to its membranestabilizing effect inhibited edema formation by 910 (119901 lt0001) (Figure 2(c))

Topical administration of capsaicin releases proinflam-matory mediators such as substance P and histamine dueto TRPV-1 activation which result in an immediate vasodi-lation and erythema followed by edema Maximum edemais achieved within 30 minutes after capsaicin administration[57 58] Neither babassu oil (10 120583Lear) nor lauric acid(4mgear) was able to inhibit capsaicin-induced ear edemaRuthenium red (3mgkg sc) given 30min before capsaicinadministration reduced the ear edema by 753 (119901 lt 001)(data not shown)

33 HLB Value and Microemulsion Formulation The HLBnumber is a semiempirical scale for selecting surfactants [26]The corrected HLB of the selected surfactant or blend ofsurfactants thatmatch theHLBof the selected oil provides thelowest interface tension between the oil and water phases andgives the system stability [28] In this context the most stableemulsion is obtained in system with the smallest droplet sizeAnalysis by DLS showed the smallest mean droplet diametersin babassu oil emulsions at an HLB of 80 (23882120583m)

The maximum turbidity values are the same HLB valueat which the mean droplet diameter is minimal [27] Herewe found that the highest mean turbidity values for babassuoil emulsions were obtained at HLB values from 80 to 100(08ndash106) The correlation coefficient (Pearson 119903) betweenthe turbidity values and the mean droplet size for the emul-sion with an HLB value of 80 was 0275 (1199032 = 0075624)showing a positive correlation (119901 lt 005 ANOVA followedby Tukeyrsquos test)

Using the HLB value required for the babassu oil (HLB80) Span 80 and Kolliphor EL were selected as the surfac-tants at a ratio of 6 4 respectively because this blend had anHLB value equal to that determined for the oil The aqueousphase was composed of water and propylene glycol (1 3) andthe oil phase was babassu oil In most cases single-chain


410 05

BA Indo LA

A

B

CBC

minus

0

2

4

6

8

10Ea

r ede

ma (

mg)

Arachidonic acid (2 mgear)

(a)

10 01 4

Ethyl phenylpropiolate 5

BA Dexa LA

A

BBB

minus

0

1

2

3

4

5

Ear e

dem

a (m

g)

(b)

4

Phenol 10

BA Dexa LA

A

BB

B

10 01minus0

5

10

15

Ear e

dem

a (m

g)

(c)

Figure 2 Topical anti-inflammatory activity of babassu oil and lauric acid in (a) arachidonic acid (b) ethyl phenylpropiolate and (c) phenol-induced ear edema Results are expressed as the mean plusmn SEM and analyzed by ANOVA followed by Tukeyrsquos test with 119901 set at 005 Differentletters show statistical differences between groups (119901 lt 005) (minus) Negative control (acetone) (BA) babassu oil (120583Lear) (LA) lauric acid(mgear) (Indo) indomethacin (mgear) and (Dexa) dexamethasone (mgear)

surfactants alone are unable to reduce the interfacial tensionsufficiently to form a microemulsion Propylene glycol wasadded as a cosurfactant to decrease the interfacial tensionand increase the fluidity of the interface In this case partof the propylene glycol content was incorporated into thesurfactant layer and the other part decreased the polarityof the water by dissolving in the water However a higheramount of propylene glycol molecules favors formation ofa bicontinuous microemulsion and avoids rigid structuressuch as gels and liquid-crystals [59]

The transparent liquid systems formed by the pseudo-ternary phase diagram can be used to obtain concentrationranges of babassu oil emulsifiers and the aqueous phasefor microemulsion formulations A system composed of39 aqueous phase 122 oil phase and 488 surfactants(Figure 3 white dot) was selected for physical and morpho-logical characterization The selected system exhibited the

typical characteristics of a microemulsion system such asslight viscosity transparency and stability In the diagramthis formulation is found near the borderline of ow andbicontinuous microemulsions which may indicate a tran-sitional phase structure In addition the ratio of systemcomponents favors the formation of oil droplets surroundedby water suitable for biological applications

34 Electrical Conductivity Electrical conductivity is com-monly used to characterize the microstructure transitionsthat occur in microemulsions that is transformation fromwater-entrapped systems to intermediate structures and thento water continuous microstructures Conductivity is low forreverse structures in nonconducting oil media that have littleinteractions with each other When more water is added tothe system the conductive droplets begin to contact oneanother and form other structures resulting in increased


075

075

100

100

000

050050

025

025

000

OLE

OLE

MLE

ME

Wate

rpro

pylen

e glyc

ol Span 80Kolliphor EL

075050025000 100

Babassu oil

Figure 3 Pseudo-ternary phase diagram (8 2) of the babassuoil microemulsion The black region includes the microemulsionsystems (ME) and the selected formulation (white dot) OLEopaque liquid emulsions MLE milk liquid emulsions

100 nm

Figure 4 The ultrastructure of the babassu microemulsion thatshows clusters of nanodroplets filled with oil some of which havemerged with one another and are surrounded by the surfactantinterface and aqueous phase

EC [60] This phenomenon may have occurred in the ba-bassu microemulsion because its conductivity value was2096 120583Scm

35 Morphological Analysis The ultrastructure of the ba-bassu microemulsion was investigated using transmissionelectronmicroscopy (Figure 4) We observed clusters of nan-odroplets filled with oil that were surrounded by an electrondense material composed of the surfactant interface bondedto the aqueous phase Most of the oil droplets were connectedside by side but some of them were merging indicatinga transition to an ow microemulsion In addition thickperipheral layers surrounding the nanodroplet network weresometimes observed which could be related to displacementof the water and emulsifiers from the inner phase duringsystem transformation

The microemulsion composition and individual charac-teristics of components define the phase morphology of thesystem As the concentration of water increases the dropletsincrease in size and eventually form a cluster that is consid-ered infinite At this stage the microemulsion possesses abicontinuous structure Further addition of the water phasetransforms the bicontinuous system into an ow microemul-sion where the droplets of the organic phase are surroundedby the water bath and the interface is composed of surfactantspecies During each of these transition steps morphologicalphases in intermediate regions may be formed and thestability of the systemdepends on thermodynamic conditions[60ndash62]

36 SAXS Measurements SAXS is a well-established tech-nique used to investigate the morphology shape and size ofa multiphase sample namely aggregates dispersed in liquidsto obtain structural information regarding inhomogeneitiesbased on the difference of electron density in the samplesThis technique provides them with a characteristic length onthe order of tens to hundreds of Angstroms (A) [63 64]

Figure 5(a) shows the experimental data simulatedaccording to (3) (see SM) The proposed model fits theexperimental data which present droplets sizes from 5 to15 nm with the majority size of approximately 87 nm

The scattering length density difference between the shelland matrix (Δ120578) for the microemulsion was 85 plusmn 003 nmand the ] value was 04 plusmn 001 The SAXS results showedthat the structure had a thin transition layer between the oiland aqueous phases In this case the surfactant hydrophobicchains interacted with babassu oil which was observed as asmall transition region that surrounded the oil droplets in theTEMmicrograph (see Figure 4) 119878(119902) is the Fourier transformof the pair correlation function which in turn depends onthe pair potential between micelles However the Percus-Yevick approximation provides an analytical expression for119878(119902) for monodisperse particles acting as hard spheres [65]The structure factor was estimated using the monodisperseapproachThismethod onlymultiplies the size averaged formfactor with the structure factor The interaction potentialsbetween the particles are assumed to be spherical symmetricand independent of the particle size [66 67] Figure 5(b)shows the average structure factor for the microemulsionsample The peak at approximately 119902 sim 025 nmminus1 in the119878(119902) curve indicates the correlation distance between theoil droplets Besides the other peaks at around 119902 sim 050and 078 nmminus1 show that the average interaction potentialdistance between the oil droplets was approximately 12 plusmn001 nm

37 DSC Analysis The thermal behaviors of water in themicroemulsion system were investigated by DSC and com-pared with that of pure water (Figure 6) Initially the sam-ples were cooled to minus50∘C and then heated to 60∘C at arate of 10∘Cmin The thermal transition for the propyleneglycol sample was not observed in the temperature rangePure water showed an endothermic transition near 0∘C andenthalpy of fusion of approximately 330 Jg [68] Pure babassu


0200

0 5

04060810

10 15 20 25

101

Proposed model (Equation (S1))Experimental data

01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)

Figure 5 (a) Scattering intensity 119868 of the SAXS measurements as a function of the scattering vector 119902 for the babassu oil microemulsionThesolid line is the fit according to supplementary file procedure (Equation (S1))The spherical shell had an average outer radius of119877 = 87 nmandinner radius of V119877 = 348 plusmn 002 nmThe normalized size distribution 119875(119903) as a function of the overall radius (119877) is also shown (b) Structurefactor 119878(119902) for a hard sphere interaction potential of the babassu microemulsion with an average correlation distance (119877HS) of 12 plusmn 001 nmand a volume fraction of 014 plusmn 0001

Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm

minus20 40 60minus40 200

Temperature (∘C)

Figure 6 DSC thermographs for the babassu microemulsionshowing a broad endothermic melt transition in two temperatureregions (inset)

oil showed two endothermic peaks at 7 and 25∘C that wererelated to the melting temperatures of fatty acid components[69]Themultiple peaks that were observed for the oil sampleare due to the polymorphism of different fatty acid Mostvegetable oils can be found in at least three crystalline formsdesignated as 120572 1205731015840 and 120573 [70 71] The enthalpy of fusion

of pure oil is approximately 100 plusmn 002 Jg whereas themicroemulsion sample showed a broad endothermic melt(fusion) transition in two temperature regions at 11 and 18∘C(Figure 6 inset) It is essential to note that the presenceof a nearby surface alters the thermodynamic propertiessuch as freezing point melting point and enthalpy [72 73]The endothermic transition was not observed at the sameposition as pure water This effect is related to the partialmiscibility of the surfactants and propylene glycol with water[74] The literature has shown that the melting temperaturedeviation in relation to pure water depends on the nature ofthe interaction between the surfactant and water [72 73 75]When the surfactant concentration is low the transition ofpure water is observed at approximately 0∘CTherefore whenwe are dealing with systems in which the interactions dependon the concentrations the surfactant properties can stronglycontribute to the melting temperature and fusion enthalpyFor this reason the microemulsion enthalpy decreased by95 plusmn 02 Jg when compared with water and pure oil

The presence of two melting peaks is a characteristicbehavior of a bicontinuous microemulsion in situationswhere the oil phase is composed of a single fatty acid [60 76ndash78] However when the oil phase consists of a triglyceridemolecule which is composed of various fatty acids thisbehavior depends on the interactions of each fatty acid withthe surfactants speciesThefirst peak in theDSC thermogramof the microemulsion (sim11∘C) is related to the amount of freewater The second peak (sim18∘C) refers to the fusion of the oilphase in microemulsion This thermal transition is shifted tolower temperatures because the fatty acids interact with thesurfactant through the nonpolar head


1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)

Figure 7 Topical anti-inflammatory activity of babassu oil andmicroemulsion in PMA-induced ear edema Results are expressedas the mean plusmn SEM and analyzed by ANOVA followed by Tukeyrsquostest with 119901 set at 005 Different letters show statistical differencesbetween groups (119901 lt 005) (minus) Negative control (acetone) (V)microemulsion vehicle (488 surfactants 39 aqueous phase and122 water) babassu oil (120583Lear) (ME) babassu microemulsion(120583Lear) and (Dexa) dexamethasone (mgear)

38 Rheological Behavior of the System The rheologicalbehavior of the babassu microemulsion is demonstrated inSM (Figure S5) The shear stress is proportional to increasedshear rate which is characteristic of Newtonian behaviorThe profile of shear stress versus shear rate for Newtonianliquids yields a straight line which is the expected behaviorofmicroemulsions [79ndash81]The average viscosity value for thebabassu microemulsion was approximately 029 plusmn 01Pas

39 Topical Anti-Inflammatory Activity of Babassu Microe-mulsion The topical anti-inflammatory activity of the ba-bassu microemulsion compared with pure babassu oil isshown in Figure 7 Pure babassu oil and 122 babassu oildiluted in acetone inhibited the PMA-induced ear edema by580 (119901 lt 0001) and 175 (not significant) respectivelyThe babassu microemulsion inhibited ear edema by 662(119901 lt 0001) showing an enhanced activity promoted by themicroemulsification of babassu oil since babassu oil at 122(final concentration in themicroemulsion) did not have a sig-nificant effect on this experimental model Dexamethasone(01mgear) inhibited PMA-induced ear edema by 784(119901 lt 0001)

Skin permeation enhancement by microemulsions hasbeen widely studied for several anti-inflammatory drugsfor example indomethacin aspirin and rofecoxib [82ndash84]Microemulsification of babassu oil seems to enhance skinpermeation of anti-inflammatory active compounds found inthe oil reaching the same percentages of edema inhibition aspure babassu oil with a much lower oil concentration

4 Conclusions

In this work we determined the chemical composition of themain fatty acids of babassu oil using GC-MS and the results

were confirmed by 1H and 13C NMR spectra Moreover13C NMR provided additional information showing thattriacylglycerol was the only positional fatty acid isomer in theglycerol backbone

Babassu oil and lauric showed topical anti-inflammatoryactivity in different phlogistic agents-induced ear edema inmice probably due to inhibition of AA metabolism andprostaglandin biosynthesis andor action release of his-tamine and serotonin and inhibition of preformed cytokinerelease

The developed babassu nanosystem was characterized asa phase transition microemulsion which can be consideredsimilarly bicontinuous as an ow phase in contrast withclassical microemulsion systems Addition of more aqueousphase should form a well-defined ow microemulsion

The babassu oil microemulsion obtained here displayedadvantages due to the combined features of bicontinuous andow microemulsions including very low interfacial tensionhigh fluctuating interface and high solubilizing propertiesThis system may have the ability to incorporate hydrophilicandor lipophilic drugs that could be released faster thanglobular microemulsions with superior stability against theaqueous biological environment

Topical anti-inflammatory activity of babassu oil wasenhanced by microemulsification reaching the same earedema inhibition as pure babassu oil at a much lowerconcentrationThis synthesized nanocarrier represents a newpromising strategy for diseases treatment because babassuoil contains fatty acids with important biological proper-ties such as antioxidant anti-inflammatory antitumor andantimicrobial

Abbreviations

AA Arachidonic acidBA Babassu oilCOX CyclooxygenaseDAG DiacylglycerolsDexa DexamethasoneDLS Dynamic light scatteringDSC Differential scanning calorimetryEC Electrical conductivityEPP Ethyl phenylpropiolateGC-MS Gas chromatography-mass spectrometryHLB Hydrophilic-lipophilic balanceIL InterleukinIndo IndomethacinLA Lauric acidLNLS Brazilian synchrotron light laboratoryMAG MonoacylglycerolsME MicroemulsionNMR Nuclear magnetic resonanceOW Oil-in-waterPG ProstaglandinPLA2 Phospholipase A2PMA Phorbol 12-myristate 13-acetateROS Reactive oxygen speciesSAXS Small-angle X-ray scatteringTAG Triacylglycerols


TEM Transmission electron microscopyTMS TetramethylsilaneTNF Tumor necrosis factorWO Water-in-oil

Conflicts of Interest

The authors declare no conflicts of interest

Authorsrsquo Contributions

Mysrayn Y F A Reis Simone M dos Santos Rafael MXimenes Giovanna Machado and Karina L A Saraivadesigned the research Mysrayn Y F A Reis Simone Mdos Santos Danielle R Silva Marcia V Silva Maria TerezaS Correia Daniela M A Ferraz Navarro Geanne K NSantos Fernando Hallwass Otavio Bianchi Alexandre GSilva Janaına V Melo and Alessandra B Mattos performedthe research Mysrayn Y F A Reis Simone M dos SantosDaniela M A Ferraz Navarro Fernando Hallwass OtavioBianchi Rafael M Ximenes Giovanna Machado and KarinaL A Saraiva analyzed the data Mysrayn Y F A Reis SimoneM dos Santos Rafael M Ximenes Giovanna Machadoand Karina L A Saraiva wrote the manuscript All authorsreviewed and edited the manuscript Mysrayn Y F A Reisand Simone M dos Santos contributed equally to this workRafael M Ximenes Giovanna Machado and Karina L ASaraiva also contributed equally to this work

Acknowledgments

This work was supported by FACEPE (Grant no APQ-1067-40315) CAPES and CNPq Maria Tereza S CorreiaDaniela M A Ferraz Navarro Otavio Bianchi and GiovannaMachado are grateful for CNPq Productivity Research Fel-lowships The authors also thank the Brazilian SynchrotronLight Laboratory (LNLS) for the use of scientific installations(SAXS1 beamline)

Supplementary Materials

As supplementary material one can find the GC-MS chro-matogram and also H1 and C13 MNR spectra of babassuoil Data regarding droplet size distribution shear stressand stability studies can also be found SAXS equationsused in the measurement of babassu microemulsion aredescribed in detail in this file Fig S1 droplet size andturbidity as a function of HLB The values from 8 to 10showed the smallest droplet size and the highest turbiditywhich is indicative of more stable emulsions Fig S2 GC-MS chromatogram of the babassu oil fatty acids Fig S3 1HNMR spectrum of the babassu oil sample in CDCl3 Fig S413C NMR spectrum of the babassu oil sample in CDCl3 FigS5 microemulsion rheological curve showing Newtonianbehavior Table S1 centrifugation study to investigate thestability of Babassu microemulsion Table S2 heating stressapplied to babassu microemulsion to check stability in thetemperature range 40ndash80∘C Table S3 heating-cooling cycles

to check the effect of temperature variations on the stabilityof babassu microemulsion (Supplementary Materials)

References

[1] GMA deAlmeidaMA Ramos E L Araujo C Baldauf andU P Albuquerque ldquoHuman perceptions of landscape changeThe case of a monodominant forest of Attalea speciosa Mart exSpreng (Northeast Brazil)rdquo AMBIO vol 45 no 4 pp 458ndash4672016

[2] N Porro I Veiga andDMota ldquoTraditional communities in theBrazilianAmazon and the emergence of new political identitiesThe struggle of the quebradeiras de coco babacu-babassubreaker womenrdquo Journal of Cultural Geography vol 28 no 1pp 123ndash146 2011

[3] M H S L Souza C A Monteiro P M S Figueredo F RF Nascimento and R N M Guerra ldquoEthnopharmacologicaluse of babassu (Orbignya phalerata Mart) in communitiesof babassu nut breakers in Maranhao Brazilrdquo Journal ofEthnopharmacology vol 133 no 1 pp 1ndash5 2011

[4] F R Araujo S E Gonzalez-Perez M A Lopes and I D J MViegas ldquoEthnobotany of babassu palm (Attalea speciosa Mart)in the Tucuruı Lake Protected AreasMosaic - Eastern AmazonrdquoActa Botanica Brasilica vol 30 no 2 pp 193ndash204 2016

[5] J L Almeida Campos T L L da Silva U P Albuquerque NPeroni andE LimaAraujo ldquoKnowledgeUse andManagementof the Babassu Palm (Attalea speciosa Mart ex Spreng) in theAraripe Region (Northeastern Brazil)rdquo Economic Botany vol69 no 3 pp 240ndash250 2015

[6] W Mors C T Rizzini and N A Pereira Medicinal Plants ofBrazil Mich USA 1st edition 2000

[7] I G C Bieski M Leonti J T Arnason et al ldquoEthnobotanicalstudy of medicinal plants by population of Valley of Juru-ena Region Legal Amazon Mato Grosso Brazilrdquo Journal ofEthnopharmacology vol 173 pp 383ndash423 2015

[8] E A Miles P Zoubouli and P C Calder ldquoDifferential anti-inflammatory effects of phenolic compounds from extra virginolive oil identified in human whole blood culturesrdquo NutritionJournal vol 21 no 3 pp 389ndash394 2005

[9] S Cicerale L J Lucas and R S J Keast ldquoAntimicrobial antiox-idant and anti-inflammatory phenolic activities in extra virginolive oilrdquo Current Opinion in Biotechnology vol 23 no 2 pp129ndash135 2012

[10] S-J Wu P-L Liu and L-T Ng ldquoTocotrienol-rich fraction ofpalm oil exhibits anti-inflammatory property by suppressingthe expression of inflammatorymediators in humanmonocyticcellsrdquo Molecular Nutrition amp Food Research vol 52 no 8 pp921ndash929 2008

[11] S Intahphuak P Khonsung andA Panthong ldquoAnti-inflamma-tory analgesic and antipyretic activities of virgin coconut oilrdquoPharmaceutical Biology vol 48 no 2 pp 151ndash157 2010

[12] V P de Sousa J Crean V R D A Borges et al ldquoNanostruc-tured systems containing babassu (Orbignya speciosa) oil as apotential alternative therapy for benign prostatic hyperplasiardquoInternational Journal of Nanomedicine vol 8 pp 3129ndash31392013

[13] R S Pessoa E L Franca E B Ribeiro et al ldquoMicroemulsionof babassu oil as a natural product to improve human immunesystem functionrdquo Drug Design Development and Therapy vol9 pp 21ndash31 2014


[14] M J Lawrence andG D Rees ldquoMicroemulsion-basedmedia asnovel drug delivery systemsrdquo Advanced Drug Delivery Reviewsvol 64 pp 175ndash193 2012

[15] M Fanun ldquoMicroemulsions as delivery systemsrdquoCurrent Opin-ion in Colloid amp Interface Science vol 17 no 5 pp 306ndash3132012

[16] W Naoui M-A Bolzinger B Fenet et al ldquoMicroemulsionmicrostructure influences the skin delivery of an hydrophilicdrugrdquo Pharmaceutical Research vol 28 no 7 pp 1683ndash16952011

[17] F H Xavier-Junior C Vauthier A R V Morais E N Alencarand E S T Egito ldquoMicroemulsion systems containing bioactivenatural oils an overview on the state of the artrdquo Drug Develop-ment and Industrial Pharmacy vol 43 no 5 pp 700ndash714 2017

[18] F Mezni A Maaroufi M Msallem and et al ldquoFatty acid com-position antioxidant and antibacterial activities of Pistacialentiscus L fruit oilsrdquo Journal of Medicinal Plants Research vol6 no 39 pp 5266ndash5271 2012

[19] American Oil Chemistsrsquo Society Official Methods and Rec-ommended Practices of the American Oil Chemists AOCSChampaign Ill USA 4th edition 1990

[20] LDMetcalfe J R Pelka andAA Schmitz ldquoRapid preparationof fatty acid esters from lipids for gas chromatographic analysisrdquoAnalytical Chemistry vol 38 no 3 pp 514-515 1966

[21] M C Milinsk M Matsushita J V Visentainer C C DeOliveira and N E De Souza ldquoComparative analysis of eightesterification methods in the quantitative determination ofvegetable oil fatty acid methyl esters (FAME)rdquo Journal of theBrazilian Chemical Society vol 19 no 8 pp 1475ndash1483 2008

[22] A Tubaro P Dri G Delbello C Zilli and R D Loggia ldquoTheCroton oil ear test revisitedrdquo Agents and Actions Supplementsvol 17 no 3-4 pp 347ndash349 1986

[23] R P Carlson L OrsquoNeill-Davis J Chang and A J Lewis ldquoMod-ulation of mouse ear edema by cyclooxygenase and lipoxyge-nase inhibitors and other pharmacologic agentsrdquo Agents andActions Supplements vol 17 no 2 pp 197ndash204 1985

[24] R Brattsand A Thalen K Roempke L Kallstrom and EGruvstad ldquoInfluence of 1612057217120572-acetal substitution and steroidnucleus fluorination on the topical to systemic activity ratioof glucocorticoidsrdquo The Journal of Steroid Biochemistry andMolecular Biology vol 16 no 6 pp 779ndash786 1982

[25] M Gabor and Z Razga ldquoDevelopment and inhibition of mouseear oedema induced with capsaicinrdquoAgents and Actions Supple-ments vol 36 no 1-2 pp 83ndash86 1992

[26] W C Griffin ldquoClassification of Surface Active Agents by HLBrdquoJournal of the Society of Cosmetic Chemists vol 1 no 5 pp 311ndash326 1949

[27] L O Orafidiya and F A Oladimeji ldquoDetermination of therequired HLB values of some essential oilsrdquo International Jour-nal of Pharmaceutics vol 237 no 1-2 pp 241ndash249 2002

[28] E S Mahdi M H Sakeena M F Abdulkarim G Z AbdullahMA Sattar andAMNoor ldquoEffect of surfactant and surfactantblends on pseudoternary phase diagram behavior of newlysynthesized palm kernel oil estersrdquo Drug Design DevelopmentandTherapy vol 5 pp 311ndash323 2011

[29] R A Ferrari and M P Soler ldquoObtention and characterizationof coconut babassu derivativesrdquo Scientia Agricola vol 72 no 4pp 291ndash296 2015

[30] F L Jackson and H E Longenecker ldquoThe fatty acids and glyc-erides of babassu oilrdquo Oil amp Soap vol 21 no 3 pp 73ndash75 1944

[31] B S Ferreira L P Faza and M Le Hyaric ldquoA comparison ofthe physicochemical properties and fatty acid composition ofindaia (Attalea dubia) and Babassu (Orbignya phalerata) oilsrdquoThe Scientific World Journal vol 2012 Article ID 532374 4pages 2012

[32] A Barison C W P da Silva F R Campos F Simonelli C ALenz and A G Ferreira ldquoA simple methodology for the deter-mination of fatty acid composition in edible oils through 1HNMR spectroscopyrdquoMagnetic Resonance in Chemistry vol 48no 8 pp 642ndash650 2010

[33] F D Gunstone Advances in Lipid Methodology Oily PressDundee Scotland 2nd edition 1993

[34] G Knothe and J A Kenar ldquoDetermination of the fatty acidprofile by 1H-NMR spectroscopyrdquo European Journal of LipidScience and Technology vol 106 no 2 pp 88ndash96 2004

[35] A Rosa A Rescigno A Piras et al ldquoChemical compositionand effect on intestinal Caco-2 cell viability and lipid profile offixed oil from Cynomorium coccineum Lrdquo Food and ChemicalToxicology vol 50 no 10 pp 3799ndash3807 2012

[36] R Sacchi F Addeo I Giudicianni et al ldquoAnalysis of thepositional distribution of fatty acids in olive oil triacylglycerolsby high resolution 13C-NMR of the carnonyl regionrdquo ItalianJournal of Food Science vol 4 pp 117ndash123 1992

[37] R Medeiros M F Otuki M C W Avellar and J B Cal-ixto ldquoMechanisms underlying the inhibitory actions of thepentacyclic triterpene 120572-amyrin in the mouse skin inflamma-tion induced by phorbol ester 12-O-tetradecanoylphorbol-13-acetaterdquo European Journal of Pharmacology vol 559 no 2-3 pp227ndash235 2007

[38] MMurakawa K Yamaoka Y Tanaka and Y Fukuda ldquoInvolve-ment of tumor necrosis factor (TNF)-120572 in phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced skin edema inmicerdquo Biochemical Pharmacology vol 71 no 9 pp 1331ndash13362006

[39] Y H Shin S-H Yoon E-Y Choe et al ldquoPMA-induced up-regulation of MMP-9 is regulated by a PKCa-NF-B cascade inhuman lung epithelial cellsrdquo Experimental Molecular Medicinevol 39 no 1 pp 97ndash105 2007

[40] I A B S Alves S M Santos R F V Mendes et al ldquoChemicalcomposition antioxidant and topical anti-inflammatory activ-ities of Croton cordiifolius Baill (Euphorbiaceae)rdquo Journal ofMedicinal Plant Research vol 11 no 2 pp 22ndash33 2017

[41] E E Bralley P Greenspan J L Hargrove L Wicker and D KHartle ldquoTopical anti-inflammatory activity of Polygonum cusp-idatum extract in the TPA model of mouse ear inflammationrdquoJournal of Inflammation vol 5 article 1 2008

[42] T S Rao J L Currie A F Shaffer and P C Isakson ldquoCom-parative evaluation of arachidonic acid (AA)- and tetrade-canoylphorbol acetate (TPA)-induced dermal inflammationrdquoInflammation vol 17 no 6 pp 723ndash741 1993

[43] C L Fischer D R Blanchette K A Brogden et al ldquoThe roles ofcutaneous lipids in host defenserdquo Biochimica et Biophysica Acta(BBA) - Molecular and Cell Biology of Lipids vol 1841 no 3 pp319ndash322 2014

[44] TNakatsujiM C Kao J-Y Fang et al ldquoAntimicrobial propertyof lauric acid against propionibacterium acnes Its therapeuticpotential for inflammatory acne vulgarisrdquo Journal of Investiga-tive Dermatology vol 129 no 10 pp 2480ndash2488 2009

[45] D Yang D Pornpattananangkul T Nakatsuji et al ldquoThe an-timicrobial activity of liposomal lauric acids against Propioni-bacterium acnesrdquo Biomaterials vol 30 no 30 pp 6035ndash60402009


[46] W-CHuang T-H Tsai L-T Chuang Y-Y Li C C Zouboulisand P-J Tsai ldquoAnti-bacterial and anti-inflammatory propertiesof capric acid against Propionibacterium acnes A comparativestudy with lauric acidrdquo Journal of Dermatological Science vol73 no 3 pp 232ndash240 2014

[47] T Kezutyte NDesbenoit A Brunelle andV Briedis ldquoStudyingthe penetration of fatty acids into human skin by ex vivo TOF-SIMS imagingrdquo Biointerphases vol 8 no 1 pp 1ndash8 2013

[48] A Crummey G P Harper E A Boyle and F R ManganldquoInhibition of arachidonic acid-induced ear oedema as a modelfor assessing topical anti-inflammatory compoundsrdquo Agentsand Actions Supplements vol 20 no 1-2 pp 69ndash76 1987

[49] E E Opas R J Bonney and J L Humes ldquoProstaglandin andleukotriene synthesis in mouse ears inflamed by arachidonicacidrdquo Journal of Investigative Dermatology vol 84 no 4 pp253ndash256 1985

[50] J L Humes E E Opas M Galavage D Soderman and R JBonney ldquoRegulation of macrophage eicosanoid production byhydroperoxy- and hydroxy-eicosatetraenoic acidsrdquo BiochemicalJournal vol 233 no 1 pp 199ndash206 1986

[51] G E Henry R A Momin M G Nair and D L DewittldquoAntioxidant and cyclooxygenase activities of fatty acids foundin foodrdquo Journal of Agricultural and Food Chemistry vol 50 no8 pp 2231ndash2234 2002

[52] M D C L Barbosa E Bouskela F Z Cyrino et al ldquoEffectsof babassu nut oil on ischemiareperfusion-induced leukocyteadhesion and macromolecular leakage in the microcirculationObservation in the hamster cheek pouchrdquo Lipids in Health andDisease vol 11 article no 158 2012

[53] E Patrick A Burkhalter and H I Maibach ldquoRecent Investiga-tions of Mechanisms of Chemically Induced Skin Irritation inLaboratory Micerdquo Journal of Investigative Dermatology vol 88no s3 pp 24sndash31s 1987

[54] H LimH Park andH P Kim ldquoInhibition of contact dermatitisin animal models and suppression of proinflammatory geneexpression by topically applied flavonoid wogoninrdquo Archives ofPharmacal Research vol 27 no 4 pp 442ndash448 2004

[55] A R Murray E Kisin V Castranova C Kommineni MR Gunther and A A Shvedova ldquoPhenol-induced in vivooxidative stress in skin Evidence for enhanced free radical gen-eration thiol oxidation and antioxidant depletionrdquo ChemicalResearch in Toxicology vol 20 no 12 pp 1769ndash1777 2007

[56] J L Wilmer F G Burleson F Kayama J Kanno and M ILuster ldquoCytokine induction in human epidermal keratinocytesexposed to contact irritants and its relation to chemical-induced inflammation in mouse skinrdquo Journal of InvestigativeDermatology vol 102 no 6 pp 915ndash922 1994

[57] M Gabor Mouse Ear Inflammation Models and their Pharma-cological Applications Akademiai Kiado Budapeste Hungary2000

[58] B Zegarska A Lelinska and T Tyrakowsk ldquoClinical and exper-imental aspects of cutaneous neurogenic inflammationrdquo Phar-macological Reports vol 58 no 1 pp 13ndash21 2006

[59] P Szumała ldquoStructure of Microemulsion Formulated with Mo-noacylglycerols in the Presence of Polyols and Ethanolrdquo Journalof Surfactants and Detergents vol 18 no 1 pp 97ndash106 2015

[60] A Kogan D E Shalev U Raviv A Aserin and N GartildquoFormation and characterization of ordered bicontinuousmicroemulsionsrdquo The Journal of Physical Chemistry B vol 113no 31 pp 10669ndash10678 2009

[61] A E Giannakas T C Vaimakis A K Ladavos P N Trikalitisand P J Pomonis ldquoVariation of surface properties and textural

features of spinel ZnAl2O4 and perovskite LaMnO3 nanoparti-cles prepared viaCTAB-butanol-octane-nitrate saltmicroemul-sions in the reverse and bicontinuous statesrdquo Journal of Colloidand Interface Science vol 259 no 2 pp 244ndash253 2003

[62] D G Hayes J A Gomez Del Rio R Ye V S Urban S VPingali and H M OrsquoNeill ldquoEffect of protein incorporation onthe nanostructure of the bicontinuous microemulsion phaseof winsor-III systems A small-angle neutron scattering studyrdquoLangmuir vol 31 no 6 pp 1901ndash1910 2015

[63] O Bianchi L G Barbosa G MacHado L B Canto R SMauler and R V B Oliveira ldquoReactive melt blending of PS-POSS hybrid nanocompositesrdquo Journal of Applied PolymerScience vol 128 no 1 pp 811ndash827 2013

[64] AA RMota C CGattoGMachado et al ldquoStructural organi-zation and supramolecular interactions of the task-specific ionicliquid 1-methyl-3-carboxymethylimidazolium chloride Solidsolution and gas phase structuresrdquo The Journal of PhysicalChemistry C vol 118 no 31 pp 17878ndash17889 2014

[65] J K Percus and G J Yevick ldquoAnalysis of Classical Statisti-cal Mechanics by Means of Collective Coordinatesrdquo PhysicalReview A Atomic Molecular and Optical Physics vol 110 no1 pp 1ndash13 1958

[66] O Glatter and O Kratky Small Angle X-Ray Scattering firstEd Academic Press Inc London UK 1982

[67] D I Svergun Structure Analysis by Small-Angle X-Ray andNeu-tron Scattering Plenum Press New York NY USA 1st edition1987

[68] P Atkins and J Paula Elements of Physical Chemistry OxfordUniversity Press Oxford UK 6th edition 2012

[69] R T Nassu and L A G Goncalves ldquoDetermination of melt-ing point of vegetable oils and fats by differential scanningcalorimetry (DSC) techniquerdquo Grasas y Aceites vol 50 no 1pp 16ndash22 1999

[70] R D OrsquoBrien Fats and Oils Formulating and Processing forApplications Taylor amp Francis Boca Raton FL USA 3rdedition 2008

[71] T Wang and J L Briggs ldquoRheological and thermal propertiesof soybean oils with modified FA compositionsrdquo Journal of theAmerican Oil Chemistsrsquo Society vol 79 no 8 pp 831ndash836 2002

[72] N Garti A Aserin I Tiunova and M Fanun ldquoA DSC studyof water behavior in water-in-oil microemulsions stabilized bysucrose esters and butanolrdquo Colloids and Surfaces A Physico-chemical and Engineering Aspects vol 170 no 1 pp 1ndash18 2000

[73] F Podlogar M Gasperlin M Tomsic A Jamnik and M BRogac ldquoStructural characterisation of water-Tween 40Imwitor308-isopropyl myristate microemulsions using different exper-imental methodsrdquo International Journal of Pharmaceutics vol276 no 1-2 pp 115ndash128 2004

[74] F Sardari and A Jouyban ldquoSolubility of nifedipine in ethanol+ water and propylene glycol + water mixtures at 2932 to 3132Krdquo Industrial amp Engineering Chemistry Research vol 52 no 40pp 14353ndash14358 2013

[75] P Boonme K Krauel A Graf T Rades and V B JunyaprasertldquoCharacterization of microemulsion structures in the pseu-doternary phase diagramof isopropyl palmitatewaterBrij 971-butanolrdquo AAPS PharmSciTech vol 7 no 2 article E45 pp 1ndash62006

[76] S Fisher E J Wachtel A Aserin and N Garti ldquoSolubilizationof simvastatin and phytosterols in a dilutable microemulsionsystemrdquo Colloids and Surfaces B Biointerfaces vol 107 pp 35ndash42 2013


[77] R M Hathout T J Woodman S Mansour N D Mortada AS Geneidi and R H Guy ldquoMicroemulsion formulations forthe transdermal delivery of testosteronerdquo European Journal ofPharmaceutical Sciences vol 40 no 3 pp 188ndash196 2010

[78] C Note J Koetz and S Kosmella ldquoStructural changes inpoly(ethyleneimine) modified microemulsionrdquo Journal of Col-loid and Interface Science vol 302 no 2 pp 662ndash668 2006

[79] H A Barnes J F Hutton and K Walters An Introductionto Rheology Elsevier Science Publishers Amsterdam Nether-lands 1st edition 1989

[80] C W Macosko Rheology principles measurements and appli-cations Wiley-VCH New York NY USA 1st edition 1994

[81] A Mouri O Diat A El Ghzaoui et al ldquoPhase behavior ofreverse microemulsions based on Peceolrdquo Journal of Colloidand Interface Science vol 416 pp 139ndash146 2014

[82] N Barakat E Fouad and A Elmedany ldquoEnhancement ofskin permeation and anti-inflammatory effect of indomethacinusing microemulsionrdquo Asian Journal of Pharmaceutics vol 5no 3 pp 141ndash149 2011

[83] K G H Desai ldquoEnhanced skin permeation of rofecoxib usingtopicalmicroemulsion gelrdquoDrugDevelopment Research vol 63no 1 pp 33ndash40 2004

[84] B Subramanian F Kuo E Ada et al ldquoEnhancement of anti-inflammatory property of aspirin in mice by a nano-emulsionpreparationrdquo International Immunopharmacology vol 8 no 11pp 1533ndash1539 2008

Submit your manuscripts athttpswwwhindawicom

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014


MEDIATORSINFLAMMATION

of


Behavioural Neurology

EndocrinologyInternational Journal of



Disease Markers


BioMed Research International

OncologyJournal of



Oxidative Medicine and Cellular Longevity


PPAR Research

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Immunology ResearchHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

ObesityJournal of



Computational and Mathematical Methods in Medicine

OphthalmologyJournal of


Diabetes ResearchJournal of



Research and TreatmentAIDS


Gastroenterology Research and Practice


Parkinsonrsquos Disease

Evidence-Based Complementary and Alternative Medicine

Volume 2014Hindawi Publishing Corporationhttpwwwhindawicom


region of Brazil most of them in state of Maranhao but alsoin Piauı and inChapada doAraripeCEGathering of fruits toproduce oil or to sell the nuts is activities predominantlymadeby women who are called ldquoquebradeiras de coco babacurdquo (PorLit Bassabu nut breaker women) [1ndash3] These communitiesdeveloped autochthonous knowledge about babassu usesas source of human and animal food utensils and toolsfuel construction and soil fertilizers and as cosmetic andmedicines [3ndash6]

Among the parts used for medicinal purposes leavesroots and fruits should be highlighted Leaves and roots areused as tea for pain and wound healing while fruits are usedin a much bigger scale mesocarp floor and milk are usedfor the treatment of gastritis hepatitis osteoporosis skinwounds and leukorrhea liquid albumen is used as eyedropsto treat conjunctivitis and the seed oil is used as laxativevermifuge and anti-inflammatory and for the treatment ofmyiasis mycosis skin wounds hemorrhoids leukorrhea andfemale genital inflammation and spider bites [3ndash5 7]

Vegetable oils such as olive palm and coconut oils arewell known for their anti-inflammatory properties Olive oilis rich in anti-inflammatory phenolics like oleocanthal andoleuropein glycosides [8 9] while the effect of palm oil isattributed to its high tocotrienol content [10] Like babassuoil coconut oil is poor in phenolics and tocopherols Its anti-inflammatory effect is assigned to the presence of lauric acidand glyceryl laurates [11]

The availability of babassu oil and its easy handling haveenabled development of several types of formulations toenhance the therapeutic efficacy of the biologically activecomponents Targeted delivery can be achieved by apply-ing pharmaceutical nanotechnology which is based on thesynthesis application and characterization of nanoscaletherapeutic systems to provide a more controlled drugrelease In this context nanostructured systems for examplemicroemulsions containing babassu oil may act as new andpotentially efficient therapies for benign prostatic hyperpla-sia due to their antiproliferative and apoptotic effects [12]and improve human immune system function by increas-ing superoxide anion release phagocytosis of mononuclearphagocytes and antimicrobial activities [13]

Microemulsion is a system of two immiscible fluids thatis stabilized by an interfacial film of surfactants It offersthe advantages of spontaneous formation thermodynamicstability manufacturing simplicity solubilization capacity oflipophilic andor hydrophilic solutes a large area per volumeratio for mass transfer and the potential for permeationenhancement [14ndash17] Thus the aim of this work was toevaluate the topical anti-inflammatory activity of babassu oilobtained from Chapada do Araripe in Ceara (Brazil) andsynthesize and characterize a microemulsion system aimingto enhance babassu oil topical anti-inflammatory activity

2 Material and Methods

21 Chemicals Phorbol 12-myristate 13-acetate (PMA) ar-achidonic acid (AA) ethyl phenylpropiolate (EPP) phenoland capsaicin were purchased from Sigma Chemical Co(St Louis MO USA) The surfactants Span 80 (sorbitan

mono-oleate) and Tween 80 (polyoxyethylene sorbitanmonooleate) ethanol 999 deuterated CDCl3 and Supel-co 37 Component FAME Mix were also obtained fromSigma Chemical Co Kolliphor EL (polyoxyl castor oil) wasfrom BASF SE (Ludwigshafen Germany) and propyleneglycol PA was purchased from Neon Comercial Ltda (SaoPaulo Brazil)

22 Plant Material and Oil Extraction Babassu fruits werecollected from one palm tree in Araripe located in the Stateof Ceara (Brazil) in July 2013 Botanical authentication wasconducted by Olıvia O Cano and a voucher specimen wasdeposited at the Herbarium of the Agronomic Institute ofPernambuco under number 90472

The oil was extracted by a well-established method thatis practiced by farmers in forested areas After harvestingthe seeds were ground with a stone grinder and the obtainedpaste was mixed well and allowed to stand overnight On thenext day cold water was added to the paste and the upperpart was collected and placed on a fire to heat it until itstarted boiling Subsequently the liquid phase was separatedfrom the oil meal using a tissue and heated until the watercompletely evaporated Finally the oil was filtered and used[18]

23 Physicochemical Characterization and Fatty Acid ProfileTraditionally extracted babassu oil was immediately analysedfor some physicochemical properties as described by AdolfoLutz Institute [19] as follows relative density was determinedin a 10mL glass hydrometer (pycnometer) at 25∘C refractiveindex at 40∘C was performed using a Abbe refractometerperoxide values were calculated from the iodine release frompotassium iodide and expressed as milliequivalents of activeoxygenkg of oil rancidity (lipid oxidation) was determinedby Kreis reaction using phloroglucinol in acid medium acidvalues were expressed as milligrams of KOH required toneutralize the free acids of 1 g of oil which was determinedby titration in methanolic solution

The transesterification procedure of the oil was realized inaccord with Metcalfe et al [20] The oil (150mg) was mixedwith 40mL of NaOH in 050M methanol and incubatedat 100∘C for 5 minutes Next 50mL solution of BF3 (12)in methanol was added in the mixture and was heated for2 minutes After cooling to room temperature 50mL of asaturated sodium chloride solution was added The mixturewas transferred to a separation funnel containing 200mL ofpetroleum ether and vigorously stirred After a period of restthe aqueous phase was discarded and the ether phase wasfiltered The solvent was evaporated in a rotary evaporatorat 60∘C and the residual solvent was removed with nitrogenflow Methyl esters were solubilized in n-heptane beforeinjection into the gas chromatographer [21]

The samples were analyzed using an Agilent Technolo-gies (Palo Alto CA USA) 5975C single quadrupole GC-MS equipped with a nonpolar HP-5MS (Agilent) fusedsilica capillary column (30m times 025mm id film thickness025mm) The oven was initially held at 150∘C for 2minincreased to 230∘C by 5∘Cmin (held for 7min) and finallyincreased to 260∘C by 4∘Cmin The final temperature was







EE (mg) = 119908RE minus 119908LE (1)

















119902 =(4120587 sen 120579)120582 (2)






















CH3 140










10

AA

B

C

DD

1 3 01 4

Dexa LA

minus

Babassu oil

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)

(a)

101000100 300

A

B

CBC

C

minus

Babassu oil

0

5

10

15

Ear e

dem

a (m

g)

PMA (25gear)

Indo

(b)










410 05

BA Indo LA

A

B

CBC

minus

0

2

4

6

8

10Ea

r ede

ma (

mg)


(a)

10 01 4


BA Dexa LA

A

BBB

minus

0

1

2

3

4

5

Ear e

dem

a (m

g)

(b)

4

Phenol 10

BA Dexa LA

A

BB

B

10 01minus0

5

10

15

Ear e

dem

a (m

g)

(c)







075

075

100

100

000

050050

025

025

000

OLE

OLE

MLE

ME

Wate

rpro

pylen

e glyc


075050025000 100

Babassu oil


100 nm










0200

0 5

04060810

10 15 20 25

101


01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)


Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm


Temperature (∘C)






1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of






















EE (mg) = 119908RE minus 119908LE (1)

















119902 =(4120587 sen 120579)120582 (2)






















CH3 140










10

AA

B

C

DD

1 3 01 4

Dexa LA

minus

Babassu oil

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)

(a)

101000100 300

A

B

CBC

C

minus

Babassu oil

0

5

10

15

Ear e

dem

a (m

g)

PMA (25gear)

Indo

(b)










410 05

BA Indo LA

A

B

CBC

minus

0

2

4

6

8

10Ea

r ede

ma (

mg)


(a)

10 01 4


BA Dexa LA

A

BBB

minus

0

1

2

3

4

5

Ear e

dem

a (m

g)

(b)

4

Phenol 10

BA Dexa LA

A

BB

B

10 01minus0

5

10

15

Ear e

dem

a (m

g)

(c)







075

075

100

100

000

050050

025

025

000

OLE

OLE

MLE

ME

Wate

rpro

pylen

e glyc


075050025000 100

Babassu oil


100 nm










0200

0 5

04060810

10 15 20 25

101


01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)


Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm


Temperature (∘C)






1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
























119902 =(4120587 sen 120579)120582 (2)






















CH3 140










10

AA

B

C

DD

1 3 01 4

Dexa LA

minus

Babassu oil

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)

(a)

101000100 300

A

B

CBC

C

minus

Babassu oil

0

5

10

15

Ear e

dem

a (m

g)

PMA (25gear)

Indo

(b)










410 05

BA Indo LA

A

B

CBC

minus

0

2

4

6

8

10Ea

r ede

ma (

mg)


(a)

10 01 4


BA Dexa LA

A

BBB

minus

0

1

2

3

4

5

Ear e

dem

a (m

g)

(b)

4

Phenol 10

BA Dexa LA

A

BB

B

10 01minus0

5

10

15

Ear e

dem

a (m

g)

(c)







075

075

100

100

000

050050

025

025

000

OLE

OLE

MLE

ME

Wate

rpro

pylen

e glyc


075050025000 100

Babassu oil


100 nm










0200

0 5

04060810

10 15 20 25

101


01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)


Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm


Temperature (∘C)






1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of





















CH3 140










10

AA

B

C

DD

1 3 01 4

Dexa LA

minus

Babassu oil

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)

(a)

101000100 300

A

B

CBC

C

minus

Babassu oil

0

5

10

15

Ear e

dem

a (m

g)

PMA (25gear)

Indo

(b)










410 05

BA Indo LA

A

B

CBC

minus

0

2

4

6

8

10Ea

r ede

ma (

mg)


(a)

10 01 4


BA Dexa LA

A

BBB

minus

0

1

2

3

4

5

Ear e

dem

a (m

g)

(b)

4

Phenol 10

BA Dexa LA

A

BB

B

10 01minus0

5

10

15

Ear e

dem

a (m

g)

(c)







075

075

100

100

000

050050

025

025

000

OLE

OLE

MLE

ME

Wate

rpro

pylen

e glyc


075050025000 100

Babassu oil


100 nm










0200

0 5

04060810

10 15 20 25

101


01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)


Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm


Temperature (∘C)






1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















10

AA

B

C

DD

1 3 01 4

Dexa LA

minus

Babassu oil

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)

(a)

101000100 300

A

B

CBC

C

minus

Babassu oil

0

5

10

15

Ear e

dem

a (m

g)

PMA (25gear)

Indo

(b)










410 05

BA Indo LA

A

B

CBC

minus

0

2

4

6

8

10Ea

r ede

ma (

mg)


(a)

10 01 4


BA Dexa LA

A

BBB

minus

0

1

2

3

4

5

Ear e

dem

a (m

g)

(b)

4

Phenol 10

BA Dexa LA

A

BB

B

10 01minus0

5

10

15

Ear e

dem

a (m

g)

(c)







075

075

100

100

000

050050

025

025

000

OLE

OLE

MLE

ME

Wate

rpro

pylen

e glyc


075050025000 100

Babassu oil


100 nm










0200

0 5

04060810

10 15 20 25

101


01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)


Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm


Temperature (∘C)






1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















410 05

BA Indo LA

A

B

CBC

minus

0

2

4

6

8

10Ea

r ede

ma (

mg)


(a)

10 01 4


BA Dexa LA

A

BBB

minus

0

1

2

3

4

5

Ear e

dem

a (m

g)

(b)

4

Phenol 10

BA Dexa LA

A

BB

B

10 01minus0

5

10

15

Ear e

dem

a (m

g)

(c)







075

075

100

100

000

050050

025

025

000

OLE

OLE

MLE

ME

Wate

rpro

pylen

e glyc


075050025000 100

Babassu oil


100 nm










0200

0 5

04060810

10 15 20 25

101


01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)


Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm


Temperature (∘C)






1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















075

075

100

100

000

050050

025

025

000

OLE

OLE

MLE

ME

Wate

rpro

pylen

e glyc


075050025000 100

Babassu oil


100 nm










0200

0 5

04060810

10 15 20 25

101


01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)


Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm


Temperature (∘C)






1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















0200

0 5

04060810

10 15 20 25

101


01

001

q (ＨＧminus1)

1E minus 3

I(q)

P(r)

(a u

)

R (nm)

]R

Δ

R

Δ

(a)

25

20

15

05001 101

10

S(q)

q (ＨＧminus1)

(b)


Propylene glycol

Kolliphor EL

Span 80

Babassu oil

ME 25201550 10

Temperature (∘C)

（2

Tm

Tm


Temperature (∘C)






1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















1010

A A

B

A

BC

V 01122

DexaME

minus

Babassu oil

C

0

5

10

15Ea

r ede

ma (

mg)

PMA (25gear)





4 Conclusions







Abbreviations








Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of






















Acknowledgments





References






























































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
































































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of































































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Documents

Anti-Inflammatory Activity of Babassu Oil and Development ...downloads.hindawi.com/journals/ecam/2017/3647801.pdf · ResearchArticle Anti-Inflammatory Activity of Babassu Oil and