Subscribe Now Subscribe Today
Research Article
 

Isoflavones and Anti-oxidant Activities of Soybeans in Thailand



Ratana Sapbamrer, Komsak Pinta and Payungsak Tantipaiboonwong
 
Facebook Twitter Digg Reddit Linkedin StumbleUpon E-mail
ABSTRACT

Soybeans are industrial crops and widely cultivated in central and northern Thailand. Isoflavones have considerable attention because of their beneficial health effects and anti-oxidant properties. The present study aimed to investigate isoflavone levels in form of aglycones (genistein and daidzein) and anti-oxidant activities among 13 soybean cultivars in central and northern Thailand. Three soybean cultivars in central region consisted of Sukhothai 1 (SK1), Sukhothai 2 (SK2) and Srisamrong 1 (SS1) and ten soybean cultivars in northern Thailand were Chiang Mai 1 (CM1), Chiang Mai 2 (CM2), Chiang Mai 3 (CM3), Chiang Mai 6(CM6), Chiang Mai 60 (CM60), SOR JOR 1 (SJ1), SOR JOR 2 (SJ2), SOR JOR 3 (SJ3), SOR JOR 4 (SJ4) and SOR JOR 5 (SJ5). Isoflavones were analysed by solid phase extraction method and anti-oxidant activities were determined using DPPH scavenging assay. The results were found that genistein was the predominant isoflavones in soybeans. Isoflavone levels and % DPPH scavenging activities among soybeans in northern cultivars were significantly higher than those in central cultivars (p=0.000). Soybean cultivars in northern region had an average of 104.08±45.25 μg g-1 for genistein, 76.78±36.91 μg g-1 for daidzein and 25.72±3.67% for DPPH scavenging activities. Soybean cultivars in central region had an average of 18.81±9.10 μg g-1 for genistein, 17.46±12.35 μg g-1 for daidzein and 14.07±1.10% for DPPH scavenging activities. The remarkable findings were that the correlation coefficient of % DPPH scavenging activities with genistein (r = 0.706, p = 0.000) was higher than those with daidzein (r = 0.497, p = 0.000). Present results therefore suggested that genistein showed the most potent anti-oxidant and high bio-availability as a promising candidate for the prevention of cancers and other diseases.

Services
Related Articles in ASCI
Search in Google Scholar
View Citation
Report Citation

 
  How to cite this article:

Ratana Sapbamrer, Komsak Pinta and Payungsak Tantipaiboonwong, 2012. Isoflavones and Anti-oxidant Activities of Soybeans in Thailand. Research Journal of Phytochemistry, 6: 113-119.

URL: https://scialert.net/abstract/?doi=rjphyto.2012.113.119
 
Received: July 16, 2012; Accepted: September 19, 2012; Published: December 03, 2012



INTRODUCTION

Soybeans (Glycine max) are industrial crops and widely cultivated in Thailand with 25,257,600 square meters cultivated area. They are composed of macronutrients such as protein (36-46%), carbohydrate (30%) and lipid (18%) which mainly consist of polyunsaturated fatty acids such as linoleic acid (Kim and Kim, 2005; Cederroth and Nef, 2009). They also contain micronutrients which include isoflavones, saponins, phytates, vitamins and minerals. Among soybean micronutrients, isoflavones have considerable attention because of their biological properties including estrogenic and anti-oxidant activities (Mitchell et al., 1998; Patel et al., 2001; Magee et al., 2004; Lee et al., 2005; Cederroth and Nef, 2009). Epidemiological and clinical studies have shown that dietary soy isoflavones is associated with beneficial health effects, such as reduction of menopause symptom, prevention of coronary heart disease and prevention of breast and reproductive cancers form (Ozasa et al., 2004; Hedelin et al., 2006; Bandera et al., 2011; Bolanos-Diaz et al., 2011; Setchell et al., 2011; Ollberding et al., 2012). It have been suggested that some of the effect were related to the anti-oxidant activities of isoflavones.

Although, there are various soybean cultivars cultivated in Thailand for long times, there is little data regarding their isoflavone contents and anti-oxidant properties in each cultivar. Therefore, the aim of the present study was to investigate isoflavone levels in aglycone forms (genistein and daidzein) and anti-oxidant activities among 13 soybean cultivars in Central and Northern Thailand.

MATERIALS AND METHODS

Cultivars of soybeans and sampling: Between December 2011 and May 2012, thirteen soybean cultivars were took a random for analyzing isoflavones and anti-oxidant activities. Cultivar name, locality of origin and breed certification of 13 soybean cultivars are shown in Table 1. Three soybean cultivars in central Thailand were Sukhothai 1 (SK1), Sukhothai 2 (SK2) and Srisamrong 1 (SS1). Ten soy bean cultivars in northern Thailand were Chiang Mai 1(CM1), Chiang Mai 2 (CM2), Chiang Mai 3 (CM3), Chiang Mai 6(CM6), Chiang Mai 60 (CM60), SOR JOR 1 (SJ1), SOR JOR 2 (SJ2), SOR JOR 3 (SJ3), SOR JOR 4 (SJ4) and SOR JOR 5 (SJ5). One kilogram of each foundation seed was brought from Crops Research Center in Chiang Mai and Sukhothai province. Each cultivar was stepwise sampling until the final sample as 250 g and then ground into powder. The powdered were mixed and took a random 5 samples in each cultivar for analysis. All samples were stored in freezer at -20°C throughout the period of the experiment.

Analysis of isoflavones: Standards of genistein and daidzein were obtained from Sigma-Aldrich, USA. All solvents for extraction were HPLC grades (J.T. Baker, USA). Sample extraction and analysis was modified from the method of Nakamura et al. (2000). Recovery was 118.9% for genistein and 110% for daidzein. The quantitation limit was 0.95 μg g-1 for genistein and 0.88 μg g-1 for daidzein. The intrabatch coefficient of variation (% CV) was 10.68% for genistein and 10.06% for daidzein and interbatch % CV was 10.18% for genistein and 17.78% for daidzein.

Table 1: Cultivar name, locality of origin, and breed certification of 13 soybean cultivars
Image for - Isoflavones and Anti-oxidant Activities of Soybeans in Thailand
Chiang Mai Field Crop Research Center, online: http://www.chiangmai2010.com/cmfcrc/2012

Determination of anti-oxidant activities: DPPH scavenging assay was measured by modified method of Katsube et al. (2004). One gram of the powdered were mixed with 10 mL of 70%(v/v) ethanol and extracted for 12 h. The extracts were then filtered with filter paper (Whatman no.1) and stored at 4°C until analysis. The extract was diluted with water stepwise and 10 μL of dilution was pipetted into a 96-well plate. One hundred and eighty-five microliters of DPPH (2,2-diphenyl-2-picrylhydrazyl) solution dissolved in a 50% ethanol solution was added to each well and the plate was shaken for 5 min at room temperature. The absorbance was read at 550 nm by Microplate Reader (MultiRead 400, Anthos). The experiment was carried out in triplicate:

Image for - Isoflavones and Anti-oxidant Activities of Soybeans in Thailand

Statistical analysis: All results were expressed as Mean±standard deviation (SD). Data were analyzed by two independent sample test (Mann-Whitney U test) and Spearman Rank correlation coefficient (r) using SPSS 11.5. Statistical significance was set at p<0.05.

RESULTS

The highest level of genistein was found in CM1 with mean of 159.14±60.71 μg g-1. The second highest level was that in CM2 (150.85±38.56 μg g-1), followed by SJ3 (128.21±18.39 μg g-1). The highest level of daidzein was found in SJ4 with mean of 97.12±10.08 μg g-1. The second highest level was that in SJ1 (96.40±53.86 μg g-1), followed by CM2 (89.89±12.99 μg g-1). Genistein levels in most strain samples (except CM3, CM6, SJ1 and SJ5) were higher than daidzein. The mean ratio between genistein and daidzein was 1.49 (ranged from 0.31-8.96) (Table 2). Comparing with isoflavones of soybean cultivars in central and northern regions, the levels in northern cultivars (104.08±45.25 μg g-1 for genistein and 76.78±36.91 μg g-1 for daidzein) were significantly higher than those in central cultivars (18.81±9.10 μg g-1 for genistein and 17.46±12.35 μg g-1 for daidzein) (p = 0.000) (Table 3).

The highest % scavenging activities was found in CM1 with mean of 29.4±1.95%. The second highest scavenging activities was that in SJ2 (29.2±1.3%), followed by SJ3 and CM2 (27.6±1.82 and 27.6±2.7), respectively (Table 2).

Table 2: Isoflavone levels (μg g-1) and DPPH scavenging activities in 13 soybean cultivars
Image for - Isoflavones and Anti-oxidant Activities of Soybeans in Thailand
Values are Mean±SD, n = 5

Table 3: Isoflavone levels (μg g-1) and DPPH scavenging activities, compared between soybeans of central and northern regions of Thailand
Image for - Isoflavones and Anti-oxidant Activities of Soybeans in Thailand
**Statistically different at 0.01

Table 4: Spearman Rank correlation coefficient between isoflavones and % DPPH scavenging activities in soybeans
Image for - Isoflavones and Anti-oxidant Activities of Soybeans in Thailand
**Statistical correlation significant at 0.01, n = 65

Comparing with % scavenging activities of soybeans cultivated in central and northern regions, % scavenging activities in northern cultivars (25.72±3.67%) were significantly higher than those in central cultivars (14.07±1.10%) (p = 0.000) (Table 3).

Genistein levels were positively associated with daidzein (r = 0.666, p = 0.000) and positively associated with % scavenging activities (r = 0.706, p = 0.000). Daidzein levels were also positively associated with % scavenging activities (r = 0.497, p = 0.000) (Table 4).

DISCUSSION

In our study, genistein was the predominant isoflavones in soybeans and the results were in agreement with several studies (Franke et al., 1994, 1999; Shao et al., 2009).

The remarkable findings were that isoflavones and anti-oxidant activities in northern cultivars were significantly higher than those in central cultivars. Especially, CM1 from northern cultivar had the highest genistein and anti-oxidant activities. The results therefore suggest to cultivating CM1 for more production of isoflavones and anti-oxidant activities.

Several studies have reported that isoflavone contents were affected by genetic factor, geographical location and environmental conditions during seed development (Wang and Murphy, 1994; Tsukamoto et al., 1995; Seguin et al., 2004; Vamerali et al., 2012). Central and northern Thailand are differences in geographical and environmental conditions which include temperature, precipitation, soil fertility, soil moisture and light level. Temperature seems to be a major factor affecting isoflavone synthesis. In Thailand, an average temperature in northern region (26°C) is lower than in central region (28°C). Soybeans are subtropical plants that require root zone temperature in range of 25-30°C for optimal cultivation (Jones and Tisdale, 1921). Several studies have reported that low temperature increased the activity of enzymes of phenylpropanoid and flavonoid pathways and temperature higher than 24°C during seed development reduced isoflavone contents (Tsukamoto et al., 1995; Carrao-Panizzi et al., 1999; Janas et al., 2002; Posmyk et al., 2005). Water supply is also expected to be increased isoflavone contents (Caldwell et al., 2005). However, the study of Vamerali et al. (2012) reported that water supply increased protein and yield but effect on isoflavones was negligible.

Genistein and daidzein levels were positively associated with % scavenging activities (r = 0.706, r = 0.497, p = 0.000, respectively). It was due to their ability to reduce free radical formation and to scavenge free radicals (Pietta, 2000; Djuric et al., 2001). The remarkable findings were that the correlation coefficient of % scavenging activities with genistein was higher than those with daidzein. A number of studies have reported that genistein showed the most potent anti-oxidant (Wei et al., 1995; Ruiz-Larrea et al., 1997; Rimbach et al., 2003). It was also mentioned that the number and position of hydroxyl groups were factors for anti-oxidants activities and the c-4’ position was crucial (Wei et al., 1995; Arora et al., 1998). Therefore, it could be hypothesized that high amounts and relatively high anti-oxidants of genistein is a promising candidate for the prevention of human cancers and other diseases.

In conclusion, the important factors affecting isoflavone contents in soybeans were geographical and environmental conditions. The high association between genistein and anti-oxidant activities in soybean indicated that genistein showed the most potent anti-oxidant and high bio-availability as a promising candidate for the prevention of cancers and other diseases.

ACKNOWLEDGMENTS

The study was funded by University of Phayao Grant 2011. We are grateful to Crops Research Centers in Chiang Mai and Sukhothai provinces for giving foundation soybeans and thankful to Emeritus Prof. Dr. Maitree Suttajit and Assoc. Prof. Dr. Nuwat Visavarungroj for their guidance. We are also grateful to School of Science, School of Medical Science and School of Medicine, University of Phayao for assisting laboratories.

REFERENCES

  1. Arora, A., M.G. Nair and G.M. Strasburg, 1998. Antioxidant activities of isoflavones and their biological metabolites in a liposomal system. Arch. Biochem. Biophys., 356: 133-141.
    CrossRef  |  Direct Link  |  


  2. Bandera, E.V., M. King, U. Chandran, L.E. Paddock, L. Rodriguez-Rodriguez and S.H. Olson, 2011. Phytoestrogen consumption from foods and supplements and epithelial ovarian cancer risk: A population-based case control study. BMC. Womens Health, 11: 40-40.
    CrossRef  |  Direct Link  |  


  3. Bolanos-Diaz, R., J.C. Zavala-Gonzales, E. Mezones-Holguin and J. Francia-Romero, 2011. Soy extracts versus hormone therapy for reduction of menopausal hot flushes: Indirect comparison. Menopause, 18: 825-829.
    PubMed  |  


  4. Caldwell, C.R., S.J. Britz and M. Mirecki, 2005. Effect of temperature, elevated carbon dioxide and drought during seed development on the isoflavone content of dwarf soybean [Glycine max (L.) Merrill] grown in controlled environments. J. Agric. Food Chem., 53: 1125-1129.
    CrossRef  |  


  5. Cederroth, C.R. and S. Nef, 2009. Soy, phytoestrogens and metabolism: A review. Mol. Cell. Endocrinol., 304: 30-42.
    CrossRef  |  


  6. Djuric, Z., G. Chen, D.R. Doerge, L.K. Heilbrun and O. Kucuk, 2001. Effect of soy isoflavone supplementation on markers of oxidative stress in men and women. Cancer Lett., 172: 1-6.
    CrossRef  |  


  7. Franke, A.A., J.H. Hankin, M.C. Yu, G. Maskarinec, S.H. Low and L.J. Custer, 1999. Isoflavone levels in soy foods consumed by multiethnic populations in Singapore and Hawaii. J. Agric. Food Chem., 47: 977-986.
    CrossRef  |  


  8. Franke, A.A., L.J. Custer, C.M. Cerna and K.K. Narala, 1994. Quantitation of phytoestrogens in legumes by HPLC. J. Agric. Food Chem., 9: 1905-1913.
    Direct Link  |  


  9. Hedelin, M., K.A. Balter, E.T. Chang, R. Bellocco and A. Klint et al., 2006. Dietary intake of phytoestrogens, estrogen receptor-beta polymorphisms and the risk of prostate cancer. Prostate, 66: 1512-1520.
    PubMed  |  


  10. Janas, K.M., M. Cvikrova, A. Palagiewicz, K. Szafranska and M.M. Posmyk, 2002. Constitutive elevated accumulation of phenylpropanoids in soybean roots at low temperature. Plant Sci., 163: 369-373.
    CrossRef  |  


  11. Jones, F.R. and W.B. Tisdale, 1921. Effect of soil temperature upon the development of nodules on the roots of certain legumes. J. Agri. Res., 22: 17-37.


  12. Katsube, T., H. Tabata, Y. Ohta, Y. Yamasaki, E. Anuurad, K. Shiwaku and Y. Yamane, 2004. Screening for antioxidant activity in edible plant products:‚ÄČ Comparison of low-density lipoprotein oxidation assay, DPPH radical scavenging assay and Folin-Ciocalteu assay. J. Agric. Food Chem., 52: 2391-2396.
    CrossRef  |  PubMed  |  Direct Link  |  


  13. Kim, M.J. and K.S. Kim, 2005. Functional and chemical composition of Hwang gumkong, Yakong and Huktae. Korean J. Food Cook Sci., 21: 844-850.


  14. Lee, C.H., L. Yang, J.Z. Xu, S.Y.V. Yeung, Y. Huang and Z.Y. Chen, 2005. Relative antioxidant activity of soybean isoflavones and their glycosides. Food Chem., 90: 735-741.
    Direct Link  |  


  15. Magee, P.J. and I.R. Rowland, 2004. Phyto-oestrogens, their mechanism of action: Current evidence for a role in breast and prostate cancer. Br. J. Nutr., 91: 513-531.
    PubMed  |  


  16. Mitchell, J.H., P.T. Gardner, D.B. McPhail, P.C. Morrice, A.R. Collins and G.G. Duthie, 1998. Antioxidant efficacy of phytoestrogens in chemical and biological model systems. Arch. Biochem. Biophys., 360: 142-148.
    CrossRef  |  


  17. Nakamura, Y., S. Tsuji and Y. Tonogai, 2000. Determination of the levels of isoflavonoids in soybeans and soy-derived foods and estimation of isoflavonoids in the Japanese daily intake. J. AOAC Int., 83: 635-650.
    PubMed  |  


  18. Ollberding, N.J., U. Lim, L.R. Wilkens, V.W. Setiawan and Y.B. Shvetsov et al., 2012. Legume, soy, tofu and isoflavone intake and endometrial cancer risk in postmenopausal women in the multiethnic cohort study. J. Natl. Cancer Inst., 104: 67-76.
    PubMed  |  


  19. Ozasa, K., M. Nakao, Y. Watanabe, K. Hayashi and T. Miki et al., 2004. Serum phytoestrogens and prostate cancer risk in a nested case-control study among Japanese men. Cancer Sci., 95: 65-71.
    PubMed  |  


  20. Patel, R.P., B.J. Boersma, J.H. Crawford, N. Hogg and M. Kirk, et al., 2001. Antioxidant mechanism of isoflavones in lipid systems: Paradoxical effects of peroxyl radical scavenging. Free Radic. Biol. Med., 31: 1570-1581.
    CrossRef  |  


  21. Pietta, P.G., 2000. Flavonoids as antioxidants. J. Nat. Prod., 63: 1035-1042.
    CrossRef  |  PubMed  |  Direct Link  |  


  22. Rimbach, G., S. de Pascual-Teresa, B.A. Ewins, S. Matsugo and Y. Uchida et al., 2003. Antioxidant and free radical scavenging activity of isoflavone metabolites. Xenobiotica, 33: 913-925.
    PubMed  |  


  23. Posmyk, M.M., C. Bailly, K. Szafranska, K.M. Janas and F. Corbineau, 2005. Antioxidant enzymes and isoflavonoids in chilled soybean (Glycine max (L.) Merr.) seedlings. J. Plant Physiol., 162: 403-412.
    PubMed  |  


  24. Ruiz-Larrea, M.B., A.R. Mohan, G. Paganga, N.J. Miller, G.P. Bolwell and C.A. Rice-Evans, 1997. Antioxidant activity of phytoestrogenic isoflavones. Free Radic. Res., 26: 63-70.
    PubMed  |  Direct Link  |  


  25. Carrao-Panizzi, M.C., A.D.P. Beleia, K. Kitamura and M.C.N. Oliveira, 1999. Effects of genetics and environment on isoflavone content of soybean from different regions of Brazil. Pesq. Agropec. Brasileira, 34: 1781-1795.
    Direct Link  |  


  26. Setchell, K.D., N.M. Brown, X. Zhao, S.L. Lindley, J.E. Heubi, E.C. King and M.J. Messina, 2011. Soy isoflavone phase II metabolism differs between rodents and humans: Implications for the effect on breast cancer risk. Am. J. Clin. Nutr., 94: 1284-1294.
    CrossRef  |  


  27. Shao, S., A.M. Duncan, R. Yang, M.F. Marcone, I. Rajcan and R. Tsao, 2009. Tracking isoflavones: From soybean to soy flour, soy protein isolates to functional soy bread. J. Functional Foods, 1: 119-127.
    CrossRef  |  


  28. Tsukamoto, C., S. Shimada, K. Igita, S. Kudou, M. Kokubun, K. Okubo and K. Kitamura, 1995. Factors affecting isoflavone content in soybean seeds: Changes in isoflavones, saponins and composition of fatty acids at different temperatures during seed development. J. Agric. Food Chem., 43: 1184-1192.
    CrossRef  |  


  29. Vamerali, T., G. Barion, M. Hewidy and G. Mosca, 2012. Soybean isoflavone patterns in main stem and branches as affected by water and nitrogen supply. Europ. J. Agronomy, 41: 1-10.
    CrossRef  |  


  30. Wang, H. and P.A. Murphy, 1994. Isoflavone composition of American and Japanese soybeans in iowa: Effects of variety, crop year and location. J. Agric. Food Chem., 42: 1674-1677.
    CrossRef  |  


  31. Wei, H., R. Bowen, Q. Cai, S. Barnes and Y. Wang, 1995. Antioxidant and antipromotional effects of the soybean isoflavone genistein. Proc. Soc. Exp. Biol. Med., 208: 124-130.
    PubMed  |  


  32. Seguin, P., W. Zheng, D.L. Smith and W. Deng, 2004. Isoflavone content of soybean cultivars grown in Eastern Canada. J. Sci. Food Agric., 84: 1327-1332.
    CrossRef  |  Direct Link  |  


©  2022 Science Alert. All Rights Reserved