Actividad de Enzima Peroxidasa de Algunos Vegetales

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J . Sci. Food Agr ic. 1985,36,877-880

The Peroxidase Enzyme Activity of someVegetables and its Resistance to HeatNezih MuftugilTUBITAK , Marmara Research Institute, Nutrition and Food Technology Department,Gebze, Kocaeli, Turkey(Manuscript received 12 April 1985)

The peroxidase enzyme activities of some fresh vegetables (cabbage, leeks, carrot,spinach, celery, squash, potatoes, onions and green beans) were determined. Theperoxidase activities of cabbage and green beans were high. Onions showed verylittle peroxidase activity. The vegetables were blanched in hot water at75", 5" and95C. Peroxidase inactivation was faster at the higher temperature blanch.Blanching of green beans, potatoes and squash at 75C for 30 min was not sufficientfor complete inactivation of peroxidase enzyme. Peroxidase enzyme inactivationwas also affected by the type of vegetable and the size of vegetable pieces.Keywords: Peroxidase; vegetables; blanching; heat treatment; enzyme activity.

1. IntroductionBlanching is an important process in the preparation of vegetables for canning, freezing anddehydration. It consists of a mild heat treatment accomplished by exposing the vegetables to hotwater or steam at 60-100C for several minutes. This treatment is quite acceptable since mostvegetables are cooked prior to consumption.

Raw and underblanched vegetables discolour, lose flavour and develop off-odours duringfrozen and dried storage. It is now widely recognised that the loss of quality that unblanchedvegetables experience during frozen and dried storage is caused by enzymic action.'-3 Despite theefforts of many investigators over the past 30 years the enzymes responsible for qualitydeterioration have not been positively identified.4 There is a general agreement that whereflavour is concerned several enzymic systems may be involved, working in sequence orsimultaneously. Peroxidase, a member of a large group of enzymes called the oxidoreductases, isconsidered to have an empirical relationship to off-flavours and off-colours in raw andunblanched vegetables, although the reactions involved have not been conclusively identified.5 Ithas been found that enzyme peroxidase present in all vegetables is the most heat stable enzymein plank5 For this reason peroxidase activity is widely used as an index of blanching.

In this study the peroxidase enzyme activity of some vegetables was determined and the effectof blanching temperature on the inactivation of peroxidase enzyme was investigated.

2. Experimental2.1 MaterialsIn this study nine different kinds of vegetables were used as the raw materials (Table 1).Vegetables were size graded, trimmed, washed and some of them were peeled. Cabbage (cut5mm), leek (cut lOmm), carrot (cut lOmm), spinach (leaves), celery (cut lOmm), squash (cutlOmm), potato (10mm cubes), onion (cut 5mm) and green beans (cut 20mm) were used, onecommercial variety of each blanched in water at three different temperatures (75, 5 and 95C).

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Table 1. Th e peroxidase activity of some fresh vegetablesPeroxidaseenzyme activityVeeetable Botanical name (unit 100n")

CabbageLeeksCarrotsSpinachCelerySquashPotatoesOnionsGreen beans

Brassica oleraceaAll ium porrumDaucus carotaSpinacia oleraceaAp ium graveolensCucurbito p e p 0Solanum tuberosumAllium cepaPhaseolus vulgaris

56.203.731.2419.464.0536.104.020.8051.46

Blanching was performed in a blanching kettle with 8 parts of water to 1 part vegetable.Temperature of the blanching water was controlled by an immersed thermocouple connected to adigital thermometer. After blanching the vegetables were cooled in cold water.2.2 MethodThe quantitative peroxidase activities of raw and blanched vegetables were assayed according tothe method described by Harvey et aL6 Vegetable purees were made by blending 50g of thesample with 50ml of 0 . 2 ~cetate buffer (pH 5.6). A known weight of puree (approximately50mg) was weighed into each of two spectrophotometer cuvettes; 2ml of acetate buffer wasadded to the sample cuvette whilst 2.7ml of acetate buffer was added to the sample cuvettewhilst 2.7ml of the same buffer was added to the reference cuvette. The contents of the cuvettewere transferred to, and homogenised in, a small tissue grinder. After homogenisation thesuspended vegetable tissue was returned to the cuvettes; 0.7ml of an aqueous mixture (1:l) of0.5% guaicol and 0.1% hydrogen peroxidase was added to the sample cuvette. Both cuvette werethen shaken prior to determining the rate of formation of brown colour in the sample bymeasuring the absorbance of light at a wavelength of 420nm in a Varian Techtron Model 635spectrophotometer. This rate was generally linear with time in the early stages of the reactionand the gradient of the line was taken to be proportional to the concentration of peroxidase inthe sample.

3. Results and discussionPeroxidase was found in all samples of fresh vegetables investigated (Table 1). Cabbage andgreen beans had high enzyme activities whereas in ordon and carrots the peroxidase enzymeactivity was low.

Table 2. The inactivation of peroxidase during the blanching of some vegetables in water at 75CPeroxidase activity 01 fresh vegetable (W)

Blanching: tim e (s)Vegetable 0 15 30 60 90 120 180 240 300 510 600 900 1200 1500 18MCabbage 100 66.40 53.13 47.43 40.80 37.95 26.56 14.45 2.10Leeks 100 45.07 30.02 12.08 1.15Carrots 100 64.50 59.60 51.61 27.41 1.63Spinach 100 31.50 22.61 11.62 2.43Celery 100 62.46 42.72 6.41 0.32Squash 100 73.87 55.40 45.70 39.88 27.14Potatoes 100 59.70 57.96 61.44 64.17 41.10 20.0Onion 100 68.75 -Green beans 100 89.10 86.37 79.90 68.20 58.02 37.31 32.8


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Peroxidase enzyme activity of vegetables 879

Table 3. The inactivation of peroxidase during the blanching of some vegetables in water at 85C

VegetableCabbageLeeksCarrotsSpinachCelerySquashPotatoesOnionGreen beans

~~ ~Peroxidas e activity of fre sh vegetable (%)Blanching time (s )0 15 30 60 90 120 180 240 360 420 600 900 1080 1200

-100 18.02 11.62 2.88 1.79100 39.90 35.00 32.17 26.17 16.80 7.9 1 2.15100 69.35 40.30 14.19 6.13 2.57 0.57100 11.76 -100 43.90 38.02 2.46100 94.50 84.76 40.60 26.59 17.72 11.80100 72.13 59.70 13.18 14.42 9.27 3.40100 37.75 -100 10.72 7.77 2.8'

Table 4. The inactivation of peroxidase during the blanching of some vegetables in water at95C

Peroxidas e activity of fresh vegetable (%)Blanching time (s)

Vegetable 0 15 30 45 60 90 120 180 210 240CabbageLeeksCarrotsSpinachCelerySquashPotatoesOnionGreen beans

100 16.25 4.10 2.82 0.8 5100 32.97 24.93 21.85 9.18 0.92100 53.20 27.45 4.83 1.05100 0.20 -100 41.48 26.17 1.97100 47.92 33.14 23.62 6.26 1.46100 72.88 6.46 3.76 1.77100 26.25 -100 73.17 56.95 23.82 8.12 5.94 0.50

The effect of blancing temperature on the inactivation of peroxidase enzyme is given in Tables2, 3 and 4.The intensity of heat treatment applied during blanching caused differences in enzymeactivity of the vegetables. Peroxidase enzyme was most easily inactivated at the high temperatureblanch. Blanching of green beans, potatoes and cabbage at 75C for 30min was not sufficient forcomplete inactivation of peroxidase. A full account of the biochemistry of this enzyme has beengiven by Saunders et aL7 It has been stated that various types of peroxidase exist and theproperties depend to some extent on its source.8 Jansen reported that when peroxidase isconsidered responsible for changes in frozen vegetables, the locale and occurrence of isoenzymesshould also be considered.' If one or more isoenzyme is responsible for adverse changes infrozen vegetables, then the measurement of total peroxidase and of the loss therein could havelittle meaning.

During blanching, peroxidase inactivation was also affected by the size of vegetable pieces.The enzyme inactivation in spinach and cabbage samples which have large surface to volumeratios and comparably high enzyme activities was faster. Fawpenetration of heat to the centrescould explain fast enzyme inactivation during blanching.

Gelatinisation of starch in potatoes by heat caused formation of a thin film on the surface ofthe potatoe cubes and thereby decreased the inactivation rate of peroxidase enzyme.

4. ConclusionsPeroxidase enzyme inactivation time depended on the temperature of the blanching water, sizeof vegetables and the nature of the enzyme present. In industrial blanching processes theperoxidase enzyme test is commonly used to determine whether a sufficient blanch has beengiven. Attempts to inhibit all peroxidase activity is not strictly necessary; a significant proportion


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880 N. Miiftiigil

of active peroxidase can be left in many vegetables an d a long shelf life in frozen storage can stillbe achieved." Th ere is some concern tha t the hea t treatmen t required to give completeinactivation may represent an excessive blanch in many cases. However, for some vegetablescomplete inactivation of peroxidase enzyme is necessary in order to obtain good quality frozenstored products."Some observations support the view that the enzyme lipoxygenase in green beans, spinach,peas and cauliflowers is responsible for the development of off-flavour during frozen storage12but its low measurable activity in the fresh products makes this enzyme less suitable as ablanching pa rame ter.AcknowledgementThis work forms par t of a research project sponsored by th e Scientific Affairs of NATO, Sciencefor Stability Programme to whom thanks are due.References1.2.3 .4.

Hartz ler, E.R .; Gue rrant, N .B. Effect of blanching and of frozen storage of vegetables on ascorbic acid retention andthe concomitant activity of certain enzymes. Food Res. 1952 17, 15-29.Rhee , K.S.; Watts, B.M. Lipid oxidation in frozen vegetables in relation to flavour change. 1. Food Sci. 1966, 31,657-679.Chow, L.; W atts, B.M . Origin of off-odors in frozen green beans. Food Technol. 1969, 23 , 1 1 S1 1 4 .Adams, J.B. Thermal requirements for blanching of fruits and vegetables to be frozen. Proceedings of Internationa[Meeting Day of 'Blanching of Fruits and Vegetables to be Fro Zen-Present Knowledge and Future Perspectives'.C . O . S .T . 91, Paris 1981, 12-32.Burne tte, F .S. Peroxidase a nd its relationship to food flavour and quality. A review. J . Food Sci. 1977, 42, 1-5.Harvey, P.; Butler , R.C .; Powel, S.J .; Ongley, M.H .; A dams. M.J. Subjective and Objective Methods Used inBlanching Research in 1976177. Technical Memorandum. No. 178. The Campden Food Preservation ResearchAssociation, G loucestershire, 1977, pp. 1&11.Saunders, B.C.; Holmes-Siedle, A.G .; Stark, B.P. Peroxidase. Butlen vorths, K ent, 1964, pp. 2-32.Aylward, F.; Haisman, D.R. Oxidation systems in fruits and vegetables. Ad v . Food Res. 1969, 17, 1-76.Jansen, E .F. Quality related chemical and physical changes in frozen foods. In : Quality and Stabiliry of Frozen Foods.(Van Arsdel, W. B.; Copley, M.S.; Olson, R.L ., E ds), W iley, New York , 1964, pp. 19-42.Bottcher, H . The enzyme content an d the quality of frozen vegetables. I. Residual peroxidase activity. Nahrung. 1975,19, 1 7 S1 7 9 .Lindquist, F.E .; Dietrich, W .C .; Masu re, M .P. ; Boggs, M.M. Effect of enzyme inactivation on quality retention infrozen Brussel Sprouts. Food Technol. 1951, 5 , 198-199Baardseth, P. Quality chang es of frozen vegetables. Food Chem. 1978,3,271-282.

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Actividad de Enzima Peroxidasa de Algunos Vegetales