Antioxidant activities and total phenolic contents of Thai curry pastes.
Thai food has been popularized and Thai curry is one of the most important kinds of foods. It is a favorite dish in Thai restaurants throughout the world due to its tasted, smell, color and health benefits. In Thai, curry paste is called "gaeng phet." The various gaeng phet dishes are based on blend of fresh and dried herbs and well known as Thai curry or "kreung gaeng" or "nam-prik-gaeng" in Thai. Thai curry paste is made from herbs and spices, sources of phytochemicals possess significant effect on human health. Apart from their major ingredients that are various bioactives, for example, antioxidants, antibacterial, antidiabetic, hypocholestero lemic and cancer preventive agent. [1-2] These biologically active components include garlic, chili (pepper) and shallot (red onion). Garlic (Allium sativum L.) and onion (Allium cepa L.) are the most important and widely used in Thai curry. They have also been reported recently of two main classes of antioxidant components, flavonoids , and sulfur-containing compounds such as dially sulfide, trisulfide, and allyl-cysteine. [4-8] Garlic can reduce the cholesterol and sugar level including, blood pressure. The therapeutic uses are as an antimicrobial, antifungal and antiviral activities as well as diaphoretic, diuretic, expectorant, antiflatulence and cholesterol lowering agents. [4-5,9-10] Shallot is normally recognized for its hypocholesterol emic and antimicrobial effects. [11-14] Chili (genus Capsicum) or pepper (Capsicum annuum) are one of the most important spices cultivated widely used as food ingredients because of their strong pungency and aroma flavour. They are an important source of P-carotene which have antimutagenic or anticarcinogenic properties.  Among these activities, capsaicinoids present anti-mutagenic and anti-tumoral properties, function as topical analgesics against pain, anti-inflammatory properties and stimulate the cardiovascular and respiratory systems. [16-17] In addition, the other ingredients of Thai curry include lemongrass, galangal, and kaffir lime leaves, which are effective in inhibiting tumors in the digestive tract. [18-21] Lemongrass contains the volatile oil citral. It reduces uric acid along with blood pressure, cholesterol, excess fats and other toxins. Lemongrass (Cympopogon citratus L.) is used in treatment of candiurectic, emmenagogue, antiflatulence, antiflu and antimicrobial agents, [22-25] while galangal root can be purchased fresh or dried. They have therapeulic uses as carminative, stomachic, antimicrobial, antibacterial and antispasmodic properties. Kaffir lime leaves have antioxidant properties and contain volatile oil. Kaffir leaves clean the blood and maintains healthy teeth and gums. Kaffir can be applied to the hair and scalp, and even prevents hair loss. The material of Thai curry paste has different type of ingredients. Red curry included red chili, shallot, garlic, lemon grass, galangal, kaffir lime leave, shrimp paste and salt. Massamun curry included dried red chili, shallot, garlic, lemon grass, galangal, kaffir lime leave, cumin, white pepper, shrimp paste and salt. Green curry included green chili, shallot, garlic, lemon grass, galangal, and kaffir lime leave. Tom-yum included lemon grass, galanga, some red chili, shallot, kaffir lime leave. Kaeng-Som included dried red chili, shallot, garlic, lemon grass, shrimp paste and salt. Thai curry is one of the most important materials of different type of Thai food, such as tom-yum, red curry, green curry, panang curry, and massaman curry.
Thus, this research aimed to study the antioxidative activity and total phenolic contents of 5 Thai curry paste (Green curry, Tom-Yum, Kaeng-Som, Massamun curry, and Red curry) extracts using a number of methods with different mechanisms. The total phenolic content (TPC) of each Thai curries which was measured. The total phenolic content of each Thai curry was measured according to the Folin-Ciocalteu method using gallic acid as a standard and the antioxidant activity was determined by a DPPH assay, inhibition of linoleic acid (LA) peroxidation and metal ion chelating.
Materials and methods
Plant Materials. Thai curry paste, 5 cultivars of Thai curry paste; Green curry, Tom-Yum, Kaeng-Som, Massamun curry, and Red curry were obtain from the local market, Mahasarakham province, Thailand, on 2009.
Reagents and Chemicals. All reagents include Folin-Ciocalteu reagent, 2,2-diphenyl-1-picrylhydrazyl, gallic acid, sodium carbonate, vitamin E, butylated hydroxyanisole, EDTA and all solvents (HPLC grade) were obtained from Fluka (Switzerland).
Extraction of crude antioxidants. The Thai curry pastes (25.0 g) were extracted with 80% ethanol (3x100 mL) for 30 min with intermittent shaking at room temperature. The extracts were combined and filtered through a 0.45 um Nylon membrane filter. After which, the extracts were then slowly concentrated under reduced pressure, at temperature below 40 [degrees]C, on a rotary evaporator to yield the crude extracts. The crude samples were used for the determination of antioxidant activity.
Total phenolic compounds. The total phenolic contents of crude Thai curry extracts were determined by spectrophotometric method using Folin-Ciocalteu's phenol reagent.  The crude extract in ethanol (0.5 mL) was placed in a test tube and was diluted to 5.0 mL with a glass of distilled water. Folin-Ciocalteu's phenol reagent (5.0 mL) was added, and the contents of the test tube were mixed thoroughly. After 3.0 min, 5 mL of 10% sodium carbonate solution was added, and the mixture was allowed to stand for 1 h with intermittent shaking. The absorbance of the blue color was measured in a Lamda 25 UV-VIS spectrophotometer (PerkinElmer, USA) at 750 nm. The concentration of total phenolic compounds was determined using the gallic acid equation (mg/mg extract) obtained from the standard gallic acid calibration curve. This experiment was carried out three times, and the results were averaged for the different fractions in the Thai curry varieties.
Antioxidative assay by the thiocyanate method. Antioxidative activity was carried out by using the linoleic acid system.  In a well-stopped Erlenmeyer flask containing linoleic acid (0.13 mL) in a 0.2 M NaOH-phosphate buffer (10 mL, pH 7), the crude antioxidants in ethanol (1 mg) from the different fractions of Thai curry were added, and the volume increased to 25 mL with a glass of distilled water. The flasks were incubated at 40 [degrees]C for a two-week period, and the degree of oxidative was measured according to the thiocyanate method. The incubation mixture (0.2 mL) was reacted with [NH.sub.4]SCN (30%, 0.2 mL), 9.4 mL of 75% EtOH, and 0.2 mL of Fe[Cl.sub.2] (2.53x[10.sup.-2] g/10 mL 3.5 % HCl) solution. The absorbance of the blue color (peroxide value) was measured in a Lamda 25 UV-VIS spectrophotometer (PerkinElmer, USA) at 500 nm. The control solution was prepared in a similar manner without the addition of any antioxidant, while a-tocopherol and butylated hydroxyanisole (BHA) at 200 ug per flask was used as a standard for comparison. This experiment was performed three times, and the results were averaged for the different fractions in the Thai curry varieties. The percentage of inhibition of lipid peroxidation was calculated using the following equation:
Inhibition (%) = [[[([A.sub.c] - [A.sub.s])]/[A.sub.c]]] x 100,
where [A.sub.c] is the absorbance of the control solution and [A.sub.s] is the absorbance in the presence of the black rice bran extracts
DPPH free radical-scavenging activity. The radical scavenging activity of crude extracts was measured using the method of Yamaguchi et al.  The crude extract and a-tocopherol (5-40 mg/mL) were added to 1.5 mL of 0.1 mM DPPH (2,2-diphenyl-1picrylhydrazyl) in ethanol. The mixture was shaken vigorously and was left to stand for 20 min at room temperature in the dark. The absorbance was measured in a Lamda 25 UV-VIS spectrophotometer (PerkinElmer, USA) at 517 nm. The control reaction contained all reagents except for the crude samples.
The radical scavenging effect was calculated by the following equation: Scavenging effect (%) = [[[([A.sub.c] - [A.sub.s])]/[A.sub.c]]] x 100,
where [A.sub.c] is the absorbance of the control at 517 nm, and [A.sub.s] is the absorbance of the extract/standard at 517nm. This experiment was repeated thrice, and the results were averaged for the different fractions in the Thai curry varieties.
Metal ion chelating activity. The chelating of ferrous ion was measured using the method of Dinis et al.  The extracts (5-25 mg/mL) were reacted with 0.05 mL of 2.0 mM FeCl2. The mixture was then added with 0.2 mL of 5.0 mM ferrozine. After which, the reaction was shaken and incubated at room temperature for 10 min. The absorbance of the red color was measured in a Lamda 25 UV-VIS spectrophotometer (PerkinElmer, USA) at 562 nm. This experiment was carried out three times, and the results were averaged for the different fractions in the Thai curry varieties. The percentage of metal chelating activity was calculated by the following equation:
% Metal chelating activity = [[[([A.sub.c] - [A.sub.s])]/[A.sub.c]]]] x 100,
where [A.sub.c] is the absorbance of the control at 562 nm, and [A.sub.s] is the absorbance of the extract/standard at 562nm. This method was performed three times, and the results were averaged for the different fractions in Thai curry cultivars. EDTA was used as a positive control.
Total phenolic compounds. The total phenolic content was measured by the Folin-Ciocalteu reagent method using gallic acid as the standard. A linear calibration curve of gallic acid resulted with a correlation coefficient of [R.sup.2] = 0.9990 over the concentration range 20-120 [micro]g/mL. This linear equation was used to determine the total phenolic compounds in Thai curry extracts. The average quantity of the total phenolic compounds found in Thai curry extracts is shown in Table 1. The amount of total phenolics content of Thai curry extract were in the range of 0.885-1.302 mg GAE/mg of extract.
Antioxidative assay of Thai curry paste extracts using the thiocyanate method. The results of antioxidative assays by the thiocyanate method compared with [alpha]-tocopherol and BHA are shown in Figure 1. All of the Thai curry samples show strong antioxidant activities which are higher than a-tocopherol but lower than BHA. The order of the inhibiting percentage is BHA (86%)>Red curry (82%)> Kaeng-Som (79%)> Green curry (77%)> Massamun curry (75%)> Tom-Yum (73%).
DPPH scavenging activity. The radical-scavenging activity of each crude Thai curry paste extract was measured by using the DPPH assay. In this study, the scavenging activity was amplified with the increased concentration of all Thai curry paste antioxidants in the range of 0.05-10 mg/mL, and it was constant at concentration above 1.0 mg/mL as shown in Figure 2. In the DPPH radical-scavenging assay, Table 1 shows the concentrations of Butylated hydroxyanisole (BHA) and the Thai curry paste extracts, at which the DPPH radicals were scavenged by 50 % ([IC.sub.50]), the lower the [IC.sub.50], the higher the antioxidant activity. The red curry extract showed the highest activity ([IC.sub.50] = 0.085 mg/mL) and the Tom-Yum showed the lowest ([IC.sub.50] = 0.202 mg/mL).
Metal chelating activity. The metal chelating activity of the Thai curry extracts was estimated by the ferrozine assay. The results indicate that the chelating ability increased with the increased concentration of all Thai curry antioxidants in the range of 0-15 mg/mL and it was constant at concentration above 10.0 mg/mL as shown in Figure 3. EDTA was used as a reference chelating agent. The order of the inhibiting percentage is EDTA>Red curry > Kaeng-Som > Green curry > Massamun curry > Tom-Yum
[FIGURE 1 OMITTED]
[FIGURE 2 OMITTED]
[FIGURE 3 OMITTED]
The present showed that Thai curry paste possess a relatively strong antioxidant activity and total phenolic contents of the Thai curry paste samples. Thai curry paste varied with different colored peppers. The phenolic compounds, carotenoids and vitamin C were observed in peppers. The phenolic extracts, the red curry showed a higher amount of phenolic components than green curry, massamun curry, kaengsom, and tom-yum, respectively. The results suggest that the level of phenolic compounds varies within different colored and fresh peppers. The red peppers contained a higher level of beta-carotene, capsanthin, quercetin, and luteolin. The green pepper had undetectable capsanthin and the lowest content of luteolin.  In the DPPH assay, radical scavenging activity in the phenolic extracts was related to the level of phenolic compounds. The red curry showed a higher level of DPPH radical scavenging activity. Therefore, we suggest that the radical scavenging activity in the peppers was from different antioxidants, mainly from vitamin C and phenolic compounds. The possible mechanisms suggest that the radical-scavenging effects of curry paste might be due to the hydroxyl groups in the antioxidants of the extracts. The antioxidative assay of Thai curry paste extracts using the thiocyanate method. The red curry extract showed the highest inhibition of lipid peroxidation and the TomYum curry extract showed the lowest inhibition of lipid peroxidation. It is suggested that the antioxidant activity in Thai curry paste comes from free radicals which promote chain reactions during the linoleic acid peroxidation system. and the chelating activity against [Fe.sup.2+] was examined. The red curry variety showed the highest percentage of metal chelating, which was reduced the concentration of the catalyzing transition metal in lipid peroxidation, which form [sigma]-bonds with a metal, are effective as secondary antioxidants because they reduce the redox potential.
The results presented in this study showed that Thai curry paste possess a relatively strong antioxidant activity of the Thai curry paste sample. Therefore, the Thai curry paste cultivar is a potential source of antioxidative phytochemicals and is a useful ingredient for nutraceutical or functional food products.
The authors wish to thanks to Department of Chemistry, Faculty of Science, Mahasarakham University and Center of Excellence for Innovation in Chemistry for a partial support of some chemicals. Financial support from Faculty of Science, Mahasarakham University is acknowledged.
 Murakami, A., Kondo, A., Nakamura, Y., Ohigashi, H., and Koshimizu, K., 1995, "Glycerogly colipids from Citrus hystrix, a traditional herb in Thailand, potenly inhibit the tumor promoting activity of 12-O-tetradecanoyl phorbol 13-actate in mouse skin," J. Agric. Food Chem., 43, pp. 2779-2783.
 Christine, M. K., and John, A. M.,2008, "The role of herbs and spices in cancer prevention," J. Nutri. Biochem., 19, pp. 347-61.
 Harborne, J. B., and Williams, C. A., 1996, Notes on flavonoid survey. In B. Mathew (Ed.): A review of Allium section Allium Kew, Royal Botanic Garden, UK, pp. 41-44.
 Agarwal, K. C., "Therapeutic actions of garlic constituents," 1996, Med. Res. Revl, 16, pp. 111-124.
 Banerjee, S. K., Mukherjee, P. K., and Maulik, S. K., 2003, "Garlic as an antioxidant: The good, the bad and the ugly," Phytother Res., 17 pp. 97-106.
 Cho, B. H. S., and Xu, S., 2000, "Effects of allyl mercaptan and various allium-derived compounds on cholesterol synthesis and secretion in Hep-G2 cells," Comp. Biochem. Physiol. C, 126, pp. 195-201.
 Rabinkov, A.., Miron, T., Konstantinovski, L., Wilchek, M., Mirelman, D., and Weiner, L., 1998, "The mode of action of allicin: Trapping of radicals and interaction with thiol containing proteins," Biochem, Biophys, Acta., 1379, pp. 233-244.
 Wang, E. J., Li, Y., Lin, M., Chen, L., Stein, A. P., Reuhl, K. R., and Yang, C. S., 1996, "Protective effects of garlic and related organosulfur compounds on acetaminophen-induced hepatotoxicity in mice," Toxicol. Appl. Pharmacol., 136, pp. 146-154.
 Amagase, H., 2006, "Clarifying the real bioactive constituents of garlic," J, Nutr., 136, pp. 716S-725S.
 Thomson, M., and Ali, M., 2003, "Garlic (Allium sativum): A review of its potential use as an anti-cancer agent," Curr. Cancer Drug Targets., 3, pp. 6781.
 Dankert, J. T. F., Tromp, H. de Vries., and Klasen, H. J., 1979, "Antimicrobial activity of crude juices of Allium ascalonicum, Allium cepa and Allium sativum,'" Zentralbl. Bakteriol., 245, pp. 229-239.
 Tappayuthpijarn, P., Dejatiwongse, Q., Hincherana, T., and Suriyant, P., 1989, "Effect of Allium ascalonicum on erythrocyte shape in induced hypercholesterolemia rabbits," J. Med. Assoc. Thai., 72, pp. 448-451.
 Adeniyi, B. A., and Anyiam, F. M., 2004, "In vitro anti-Helicobacter pylori potential of methanol extract of Allium ascalonicum Linn. (Liliaceae) leaf: susceptibility and effect on urease activity," Phytother Res., 18, pp. 358-361.
 Amin, M., and Kapadnis, B. P., 2005, "Heat stable antimicrobial activity of Allium ascalonicum against bacteria and fungi," Ind. J. Exp. Biol., 43, pp. 751-754.
 Monsereenusorn, Y., Kongsamut, S., and Pezalla, P. D., 1982, "Capsaicin in literature survey," CRC. Crit. Rev. Toxicol., 10, pp. 321-339.
 Surh, Y. J., Lee, C. R. J., Park, K. K., Mayne, S. T., Liem, A., and Miller, J. A., 1995, "Chemoprotective effects of capsaicin and diallyl sulfide against mutagenesis or tumorigenesis by vinyl carbamate and Nnitrosodimethylamine," Carcinogenesis., 16, pp. 2467.
 Sancho, R., Lucena, C., Macho, A., Calzado, M. A., Blanco-Molina, M., Min'ais, A., Appendino, G., and Munoz, E., 2002, "Immuno suppressive activity of capsaicinoids: capsiate derived from sweet peppers inhibits NFkappaB activation and is a potent antiinflammatory compound in vivo," Eur. J. Immunol., 32, pp. 1753 A.
 Siripongvutikorn, S., Thummaratwasik, P., and Huang, Y. W., 2005, "Antimicrobial and antioxidation effects of Thai seasoning. Tom-Yum," Lebensm, Wiss, Technol,, 38, pp. 347-352.
 Adegoke, G. O., and Odesola, B. A., 1996, "Storage of maize and cowpea and inhibitory of microbial agents of biodeterioration using the powder and essential oil of lemon grass (Cymbepogon citralis)," Int. Biodeter. Biodegrad., 37, pp. 81-84.
 Yasukawa, K., Sun, Y., Kitanaka, S., Tomizawa, N., Miura, M., and Motohashi, S., 2008, "Inhibitory effect of the rhizomes of Alpinia officinarum on TPA-induced inflammation and tumor promotion in two-stage carcinogenesis in mouse skin," J. Nat. Med., 62, pp. 374-378.
 Appendini, P., and Hotchkiss, J. H., 2002, "Reviewof antimicrobial food packaging," Innova. Food Science Emerging Technol., 3, pp. 113-126.
 Daferera, D., J., Ziogas, B. N., and Polissiou, M. G., 2003, "The effectiveness of plant essential oils on the growth of Botrytis cinerea, Fusarium sp. and Clavibacter michiganensis subsp," Michiganensis Crop. Prot., 22, pp. 39-44.
 Hammer, K. A., Carson, C. F., and Riley, T. V., 1999, "Antimicrobial activity of essential oils and other plant extracts," J. Appl. Microbiol., 86, pp. 985-990.
 Plotto, A., Roberts, D. D., and Roberts, R. G., 2003, "Evaluation of plant essential oils as natural postharvest disease control of tomato (Lycopersicon esculentum)," Acta Hort., 628, pp. 737-745.
 Saikia, D., Khanuja, S. P. S., Kahol, A. P., Gurta, A. P., and Kumar, S., 2001, "Comparative antifungal activity of essential oils and constituents from three distinct genotypes of Cymbopogon spp," Current Science., 80, pp. 264-266.
 Osawa, T., and Namiki, M. A., 1981, "Novel type of antioxidant isolated from leaf wax of Eucalyptus leaves," Agric. Biol. Chem., 45, pp. 735-739.
 Tsuda, T., Makino, Y., Kato, H., and Osawa, T., 1993, "Screening for antioxidative of edible pulses," Biosci. Biotech. Biochem., 57, pp 1606-1608.
 Yamaguchi, T., Takamura, H., Matobe, T., and Terao, J., 1998, "HPLC method for evaluation of the free radical-scavenging activity of food by using 1,1-diphenyl-2-picrylhydrazyl," Biosci. Biotechnol. Biochem., 62, pp. 12011204.
 Dinis, T. C. P., Madeira, V. M. C., and Almeida, L. M., 1994, "Action of phenolic derivates (acetoaminophen, salycilate and s-aminosalycilate) as inhibitors of membrane lipid peroxidation and as peroxyl radical scavengers," Arch. Biochem. Biophys., 315, pp.161-169.
 Sun, T., Xu, Z., Wu, C. T., Janes, J. M., Prinyawiwatkul, W., and No, H. K., 2007, "Antioxidant Activities of Different Colored Sweet Bell Peppers (Capsicum annuum L.)," J. Food Sci., 72, pp. S98-S102.
Muntana Nakornriab (1) and Darunee Puangpronpitag (2)
(1) Department of Chemistry and Center of Excellence for Innovation in Chemistry, Faculty of Science, Mahasarakham University, Mahasarakham 44150, Thailand.
(2) Faculty of Medicine, Mahasarakham University, Mahasarakham 44000, Thailand
Table 1: Total phenolic content and radical-scavenging activity expressed by [IC.sub.50] (mg/mL) of Thai curry extracts. Thai curry Total phenolic content [IC.sub.50] (mg GAE/mg of extract) (mg/mL) Red curry 1.302 0.085 Green curry 1.121 0.121 Kaeng-Som 0.908 0.145 Massamun curry 0.945 0.168 Tom-Yum 0.885 0.202
|Printer friendly Cite/link Email Feedback|
|Author:||Nakornriab, Muntana; Puangpronpitag, Darunee|
|Publication:||International Journal of Applied Chemistry|
|Date:||Jan 1, 2011|
|Previous Article:||Carbon nanotubes synthesis by catalytic decomposition of ethyne using Fe/Ni catalyst on aluminium oxide support.|
|Next Article:||Review of affinity precipitation methods.|