Antioxidant Activity of Phenolic Compounds in ... - ACS Publications

Chapter 17

Antioxidant Activity of Phenolic Compounds in Meat Model Systems Fereidoon Shahidi, P. K. J. P. D. Wanasundara, and C. Hong

Downloaded by COLUMBIA UNIV on June 2, 2013 | http://pubs.acs.org Publication Date: October 1, 1992 | doi: 10.1021/bk-1992-0506.ch017

Department of Biochemistry, Memorial University of Newfoundland, St. John's, Newfoundland A1B 3 X 9 , Canada Antioxidant activity of a number of phenolic compounds of plant origin was evaluated in meat model systems. Amongst the phenolic compounds tested quercetin, morin, kaempferol, myricetin, tannic acid, eugenol, isoeugenol and ellagic acid were most effective when applied to meat at 30 or 200 ppm levels. Addition of 0.5 to 2.0% of deheated mustard flour (DMF), available commercially, or its alkanolic extracts, effectively retarded lipid oxidation and off-flavor development in meat emulsion systems. The activity of all phenolics tested and that of DMF or its extracts was superior to that of α -tocopherol at a 200 ppm addition level. Furthermore, DMF had a favorable effect on water binding characteristics of meat model systems studied.

The deleterious effects of oxidation in meat products are numerous. Oxidative reactions are responsible for changes inflavor,color, texture and nutritional value, due to the destruction of fat-soluble vitamins and essential fatty acids such as linoleic acid, in both fresh and cooked muscle foods (7). Lipids in cooked meats, particularly their phospholipids, are susceptible to autoxidation. These reactions are generally catalyzed by a number of factors such as presence of oxygen, light, heat, heavy metals, pigments, alkaline conditions and also depend on the degree of unsaturation of lipid fatty acids. Lipid oxidation, defined as oxidative deterioration of unsaturated fatty acids, is a free radical-mediated phenomenon involving a chain reaction mechanism. The relative rate of oxidation of lipid fatty acids of meats, mainly C18, depends on their degree of unsaturation as given below: C78:3o3 (2500) > C18:2 C78:1G>9 (100) > C18.0 (1)

Antioxidants are added to lipid-containing foods to prevent the formation of various off-flavors and development of rancidity. In meats, nitrite curing has 0097-6156/92/0506-0214$06.00/0 © 1992 American Chemical Society In Phenolic Compounds in Food and Their Effects on Health I; Ho, C., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.


17. SHAHIDI ET AL.

Antioxidant Activity of Phenolic Compounds in Meat

shown to be an effective means of extending the shelf-life of cooked products against autoxidation. Commonly used synthetic antioxidants have also proven effective in retarding lipid oxidation in meat products. However, the present trend in food processing is to use natural ingredients (2). Thus, use of naturally-occurring antioxidants for retarding meat flavor deterioration (MFD)/warmed-over flavor (WOF) development is highly desirable. Pratt and Watts (3) reported that hot-water extracts of many vegetables were effective in retarding MFD in cooked products. These authors attributed the effectiveness of vegetable extracts to their content of flavonoids. Flavonoids are widely distributed in plant materials and as such have considerable practical impact on the flavor retention and wholesomeness of raw and cooked meats (3). Antioxygenic activity of herbs and spices has also been attributed to the presence and content of flavonoids and/or flavonoid-related compounds (4). The relationship between flavone structures and their antioxidant activity has been thoroughly investigated. Effectiveness of flavonoids in retarding lipid oxidation in fat-containing foods is apparently related to their ability to act as free radical acceptors (5-10) as well as to their chelating capacity for metal ions. Metal chelation by flavonols is due to ortho-dihydroxy (3',4'-dihydroxy) grouping on the Β ring and to the ketol structure in the C ring in their chemical structures (7,77,72). Lack of at least one of these groups may reduce or even delete the chelating capacity of flavonoids. In addition to flavonoids, many plant protein extenders used in meat emulsion systems contain large quantities of phenolic acids. Phenolic compounds isolated from rapeseed processing in the preparation of protein concentrates contained a large percentage of phenolic acids. The extracted phenolics in ethanol, after fractionation, possessed strong antioxidant effects in linoleate/0-carotene systems which were, in some cases, comparable to the effectiveness of approved food phenolics (13). Application of some natural phenolics to meat was previously reported from our laboratories (14). The present paper summarizes the results of studies on the efficiency of individual phenolic compounds of plant origin in meat model systems. Use of deheated mustard flour (DMF), as an example of a non-meat ingredient, a seasoning, and as a protein extender in comminuted meat products, and its effectiveness in retarding MFD was also investigated. Materials and Methods Materials. All flavonoids and phenolic compounds, as well as tannic acid, sesemol and other related compounds were obtained from the Sigma Chemical Company (St. Louis, Missouri) or the Aldrich Chemical Company (Milwaukee, Wisconsin). Deheated mustard flour (DMF) was a commercial product from UFL Foods Inc. (Mississauga, Ontario). Pork loins with their subcutaneous fat removed were obtained from the Newfoundland Farm Products Corporation (St. John's, Newfoundland). Extracts of DMF were prepared by extraction with water or 85% (v/v) methanol-water. The content of total phenolics in DMF extracts was determined using the Folin-Denis reagent, as described by Rhee et al. (15,16).

In Phenolic Compounds in Food and Their Effects on Health I; Ho, C., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.

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PHENOLIC COMPOUNDS IN FOOD AND THEIR EFFECTS ON HEALTH I


Sample Preparation. In each case, 80 g comminuted (4.8 mm grind plate) pork, less the amount of additives (if > 0.5%), was mixed with 20 g of distilled water. Selected additives were introduced, generally at a 30 or 200 ppm level of addition, as such, in aqueous, or in a 50% (v/v) ethanolic solution to meat prior to heat processing. Meat systems were then cooked, to an internal temperature of 75 ± 1°C, in Mason jars. Cooked meat samples were cooled to room temperature, homogenized in a Waring blender and then stored in Nasco Whirl-Pak bags at 4°C until used. The percent inhibition of malonaldehyde (MA) production in meat samples was quantified after their steam distillation using the classical 2-thiobarbituric acid test (77) as given in the following equation: 4

c o n t e n t

o f t r e a t e d

% inhibiion of MA production = ( I - * * ""^frlOO MA content of control

Results and Discussion Inhibition of malonaldehyde (MA) production, deduced from 2-thiobarbituric acid (TBA) values during a 3-week storage period of treated meats at 4°C, is given in Table I. The antioxidant activity of flavonoids arises (Table I) from their action as primary antioxidants by a free-radical scavenging mechanism and to a lesser extent by their chelating ability (results not shown) through the presence of free ortho-hydroxyl groups at 3' and 4' on the Β ring. Due to the limited number of flavonoids tested in this study, no other structural effects were evident. In all cases, antioxidant activity of flavonoids tested was superior at a 200 ppm, as opposed to a 30 ppm level of addition to comminuted pork. Furthermore, it was evident that the antioxidant activity of flavonoids was generally governed by their chemical structural attributes. Presence of a double bond at the C2-C3 position and a free hydroxyl group at the C position of ring C had a marked influence on the antioxidative activity of flavonoids. Hence, naringin and naringenin, which did not possess these features, were least effective in preventing oxidation of meat lipids. In addition, presence of a free hydroxyl group at the C position of ring A had no effect on the activity of flavonoids as no difference in the action of naringenin and naringen was observed. Rutin with a bound (etherified) hydroxyl group at the C position was much less effective as an antioxidant than its analogue with a free hydroxyl group. Generally, presence of ortho-hydroxyl groups at the 3' and 4' positions of ring Β contributed to the enhancement of flavonoids' antioxidative activity. Thus, quercetin was found to have a superior activity as compared with morin. Myricetin, with the highest number of hydroxyl groups in ring B, was found to be the most active flavonoid (at 200 ppm) in this study. The activity of benzoic acid derivatives as antioxidants depended on their chemical structure, especially the number of free hydroxyl groups found in each compound (Table II). Of the benzoic acid derivatives tested, the activity of gallic acid was superior to that of syringic acid which in turn was better than that of 3

7

3



17. SHAHIDI ET AL.

Table I

Inhibition of malonaldehyde formation by flavonoids in cooked, stored meat model systems 3'


Ring A: 5,7-di-OH Ring C

Ring B Compound

2'

3'

4'

5'

3

Kaempferol Morin Myricetin Naringenin Naringin Quercetin Rutin

H H OH H H OH H H H H H OH H OH

OH OH OH OH OH OH OH

H H OH H H H H

OH OH OH H,H H,H OH O-rutinose

b

bc

Inhibition, %' 40,96 29,97 -,99 4,7 4,7 96,98 28,37

a

At 30 and 200 ppm level of addition to meat, respectively. Saturated C -C bond. 7-0-Rhamnoglucose.

b

2

3

c

vanillic acid. This latter trend seems to be related to the number of hydroxyl groups present, and to a lesser extent on the number of oxygen-containing substituents. Thus, ellagic acid, a dimer of gallic acid, was determined to be the most effective antioxidant at a 30 ppm level of addition. In view of its desirable pharmacological and anticarcinogenic activity (19) application of ellagic acid to meat products may have practical significance. The activity of tannic acid, a compound consisting of glucose and three molecules of gallic acid, was moderate at a 30 ppm and excellent at a 200 ppm level of addition. Of the cinnamic acid derivatives generally found in oilseeds, the activity of caffeic acid with two free hydroxyl group was superior to others. Activity of chlorogenic acid, an adduct of quinic acid and with two hydroxyl groups, was pronounced at the 200 ppm level (Table III). Other phenolic acids with only one free OH group were less active (Table III). Of the other natural antioxidants examined, the activity of eugenol, isoeugenol and sesamol (Figure 1) was 95-99% at the 200 ppm level of addition. However, they were less effective at the 30 ppm level (65-79%). These activities are higher than those of ascorbic acid, α-tocopherol and butylated hydroxytoluene, BHT. The activity of BHT, a known synthetic antioxidant, was 13 and 88% at 30 and 200 ppm level of addition to meat, respectively (Figure 1).


217


218


Thus, naturally-occurring antioxidants, found in different plant materials, lend themselves to exploitation in different processing applications (18). Presence of flavonoid and flavonoid-related compounds in vegetables, fruits, oilseeds and other foods contributes to our intake at least 1 g/day (20). Thus, their possible use in meat formulations, as such or in the form of protein extenders and binders as well as spices and condiments may present an effective and attractive means of retarding lipid oxidation in muscle foods. Deheated mustard flour (DMF) is an example of an economical meat adjunct which may be used as a spice for flavor and could also improve technological properties of wieners and bologna-type products. Its inclusion in emulsified pork model systems inhibited lipid oxidation. The percent inhibition of MA production was 34% for 0.5%, 75% for 1.0%, 95% for 1.5%, and 96% for 2.0% DMF addition after a 3-week of storage at 4°C. Based on these results, inclusion of 1.0 to 2.0% DMF in meats not only offers its intended flavor effects but it also effectively retards MFD (See Figure 2). Extracts of DMF were also found to possess antioxidant activity. Both aqueous and 85% (v/v) methanoic solutions were used for extraction preparation. However, the antioxidant effect of 85% (v/v) methanoic extracts was superior to that of aqueous extracts (Figure 2). The antioxidant properties of extracts so obtained might in part be due to the content of their phenolic compounds (13). Thus, 85% (v/v) methanoic extracts of DMF contained 1.56% total phenolics, while the aqueous extracts contained only 1.08 %. The chemical nature of the antioxidant in DMF is currently under investigation in our laboratories. In addition to its strong antioxidant properties, DMF was also effective in reducing the cooking loss and improving the juiciness of products. The potency Table II

Inhibition of malonaldehyde formation by benzoic acid derivatives in cooked, stored meat model systems 3

a

Compound

Substituent

Gallic Acid Vanillic Acid Syringic Acid Ellagic Acid Tannic Acid

3,4,5-triOH 4-OH, 3-OMe 4-OH, 3,5-diOMe Dilactone of 2 gallic acids Ester of 3 to 5 gallic acids with glucose

Inhibition, %* 44,73 21,28 39,57 98,99 56,99

At 30 and 200 ppm level of addition to meat, respectively.


17. SHAHIDI ET AL.

Antioxidant Activity ofPhenolic Compounds in Meat

HO

α-Tocopherol (11,56)


Ο

OH Sesamol (70,99)

Ascorbic acid (50)

Eugenol (65,95)

Isoeugenol (79,99)

OH

Butylated Hydroxytoluene, BHT (13,88)

Figure 1. Some naturally occurring antioxidants and their efficiency as percent inhibition of malonaldehyde formation, as given in parentheses, at 30 and 200 ppm levels of addition, respectively; ascorbic acid was used at 500 ppm level.


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PHENOLIC COMPOUNDS IN FOOD AND THEIR EFFECTS ON H E A L T H I

Figure 2. Inhibition of malonaldehyde (MA) formation (%) by 0-2% deheated mustard flour (DMF) at 0.5%, 1.0%, 0 ; 1.5%, • ; and 2.0%, and with D M F extracts with water (XDMF1) and 85% methanol (XDMF2) from 0.6%, and 1.5%, meal. Values are compared with butylated hydroxytoluene at 30 ppm, S and 200 ppm, =, level in cooked, stored meat model systems.


17.

SHAHIDI ET AL.

Table III


Inhibition of malonaldehyde formation by cinnamic acid derivatives in cooked, stored meat model systems 3


Ο

a

Compound

Substituent

Coumaric Acid Caffeic Acid Ferulic Acid Sinapic Acid Chlorogenic Acid

4-OH 3,5-diOH 4-OH; 3-OMe 4-OH; 3,5-diOMe 3,4-diOH, ester with quinic acid

Inhibition, %* 24,39 57,70 34,51 27,46 8,53

At 30 and 200 ppm level of addition to meat, respectively.

of DMF at a 1-2% addition level was equivalent to that of sodium tripolyphosphate (STPP) in pork at 3000-5000 ppm, as it has been shown elsewhere (27). In mechanically deboned chicken meat, its effect was close to that of 1500-3000 ppm of STPP (results not shown). Conclusions and Future Research Needs Phenolic compounds of natural origin and related compounds, as such or as spices or protein extenders, may offer an alternative method of meat flavor preservation. Extending of the shelf-life of products would have a definite positive effect in the operation of institutional food services, fast food chains and pre-cooked meat markets. Work exploring natural antioxidants and the test of their efficiency is deemed necessary. Furthermore, effectiveness of novel protein extenders, spices and their extracts as well as selected amino acids/peptide combinations may not only confer expected properties but may also function as natural antioxidant substitutes for incorporation in food systems. Further research in this area may prove beneficial. Furthermore, application of such natural antioxidants in muscle foods, particularly restructured and emulsified systems, may have practical significance. Acknowledgments Financial support from the Natural Sciences and Engineering Research Council (NSERC) of Canada is acknowledged. Literature Cited 1.

Dziezak, J.D. FoodTechnol.1986. 40, 94-97, 101-102.


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2. 3. 4.


5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21.


Bailey, M.E. FoodTechnol.1988. 42(6), 123-126. Pratt, D.E.; Watts, B.M. J. FoodSci.1964. 29, 27-33. Pruthi, J.S. In: Spices and Condiments: Chemistry, Microbiology and Technology. Academic Press, New York. 1980. pp. 17-24. Heimann, W.; Reiff, F. Fette U. Seifen 1953. 5, 451-456. Simpson, T.H.; Uri, N . Chem. Ind. 1956. 956-957. Mehta, A.C.; Seshadri, R.T. J. Sci. Ind. Res. 1959. 18B, 24-28. Crawford, D.L.; Sinnhuber, R.O.; Aft, A. J. FoodSci.1961. 26, 139-142. Das, N.P.; Pereira, T.A. J. Amer. Oil. Chem. Soc. 1990. 67, 255-258. Pratt, D.E.; Hudson, B.J.F. In: Food Antioxidants. Hudson, B.J.F., ed. Elsevier Applied Science. London and New York. 1990. pp. 171-191. Lewis, E.J.; Watts, B.M. Food Res. 1958. 23, 274-279. Kelley, G.G.; Watts, B.M. Food Res. 1957. 22, 308-312. Zadernowski, R.; Nowak, H.; Kozlowska, H. 1991. Abstract No. C-40. Presented at the 8th International Rapeseed Congress. Saskatoon, Canada. July 9-11. Shahidi, F. Bull. Liason Groupe Pholphenols. 1988. 14, 361-362. Rhee, K.S.; Ziprin, Y.A.; Rhee, K.C. J. Food Sci. 1979. 44, 1132-1135. Rhee, K.S.; Ziprin, Y.A.; Rhee, K.C. J. Food Sci. 1981. 46, 75-77. Shahidi, F.; Rubin, L.J.; Wood, D.F. J. FoodSci.1987. 52, 564-567. Hayes, R.B.; Bookwalter, G.N.; Bagley, E.B. J. Food Sci. 1977. 42, 15271532. Castonguay, Α.; Lui, L.; Stoner, G.D. Bull. Liason Groupe Polyphenols. 1990. 15, 153-157. Kuhnau, J. Wld. Rev. Nutr. Diet. 1976. 24, 117-191. Saleemi, Z.O.; Wanasundara, P.K.J.P.D.; Shahidi, F. J. Agric. Food Chem. 1991. Submitted.

RECEIVED February 11,

1992


Antioxidant Activity of Phenolic Compounds in ... - ACS Publications

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