Quantitative analysis and metabonomic study of phytoestrogens in Africans

Ogegbo, Olumuyiwa Lateefah (2009) Quantitative analysis and metabonomic study of phytoestrogens in Africans. Doctoral thesis, London Metropolitan University.


Phytoestrogens have been widely studied in various parts of the world. However, little is known about the availability, absorption, metabolism and its biochemical effects in Africans. The principal aim of this research was to investigate the phytoestrogen content in six African staple foods, their metabolism and biochemical effects after soyintervention in Africans.

With respect to the food investigations, sensitive and reliable analytical methods for the simultaneous quantification of seven phytoestrogens using GC-MS, LC-MS, UPLC-MS were developed. The phytoestrogens analysed include four isoflavones (formoinonetin, biochanin A, daidzein, genistein), one coumestan (coumestrol), and two lignans (secoisolariciresinol and matairesinol). The methods were applied to quantitate the phytoestrogen content in six African staple foods: beans, cassava, plantain, pumpkin leaves 'ugu', rice and yam. The results showed that pumpkin leaves contained the highest amount of phytoestrogens analysed (5.66 - 12.68 ptg/g) whilst rice contaned the least amount of phytoestrogens (0.58 - 1.17 4g/g). In addition, three phytoestrogenic compounds: 3,5,7- trihydroxy-2-(4-hydroxyphenyl)-4H-chromen-4-one, 4-(hydroxyl(4-(4-hydroxy-3-methoxy-benzyl)tetrahydrofuran-3-yi)-2-methoxyphenoI and 3-(4-hydroxy-3-methoxybenzyl)-4-((4-hydroxy-3-methoxyphenyl)-(methoxy)methyl)dihydrofuran-2(3H)-one, were found in pumpkin leaves for the first time.

The second part of this research was based on a soy - intervention, where African subjects; 25 UK-based and 25 Nigerian - based were recruited and fed 200 mL Alpro soy-milk (~20 mg phytoestrogen) daily over a five - day period. The urine samples collected over 2-weeks (inclusive of the 5 consumption days) were used as biomarkers media in assessing the phytoestrogen ingestion and metabolism. The biomarkers monitored and analysed were daidzein, genistein and their metabolites: equol and odesmethylangolensin. The results showed that 35% of the selected population are 'good responders' to soy - phytoestrogens being equol and o-DMA producers. In addition, results showed metabolism was influenced by certain factors including location, gender and age.

1H NMR-metabonomics was performed on the collected bio-fluids to investigate soy - phytoestrogens' effects on specific endogenous metabolites involved in major metabolic processes. Twenty-two endogenous metabolites were quantified using Chenomx® NIVIR Suite and statistically analysed using multivariate analyses: Principal Component Analysis (PCA) and (orthogonal-) Partial-Least Square - Discriminant Analysis ((O-)PLS-DA). The results showed the levels of endogenous metabolites present in urine ranged from 5 ýM to 15 mM with huge inter - individuality variances in acetone, citrate, creatinine, glycine, hippurate and trimethylamine-N-oxide. With the MVA, results showed high inter - individuality and inter - sampling variances based on the PCA plots, however with a definitive MVA: (0)-PLS-DA, results clearly showed metabolism to be mainly influenced by location and gender.

From the exploration of the soy - challenge and metabonomics results, correlation between soy - phytoestrogen, their metabolites and endogenous metabolites were investigated using Pearson product - moment correlation. The results showed that there were significant correlations (p<0.05) between soy - phytoestrogens and their metabolites as well as between the endogenous metabolites. On the other hand, no distinct correlation was observed between either the soy - phytoestrogens or their metabolites and endogenous metabolites.

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