Bacterial Alginate Production From Azotobacter Vinelandii And Its Application In The Induction Of Phytochemical Glyceollin In Soybeans

Bacterial alginate produced by Azotobacter vinelandii is considered to be of good quality when compared with the alginate obtained from the seaweed source and it is capable of being used as an elicitor. Pathogenic effect and poor jellifying ability associated with species of Pseudomonas has made Azotobacter the preferred genus for bacterial alginate production. Azotobacter vinelandii is capable of producing bacterial alginate of high quality, through fermentation and precipitation processes. This bacterium(A. vinelandii) is able to adapt its metabolism to distinct sources of nutrients. Most studies regarding alginate have focused on its applications in the food industry, chemical characterization and physicochemical properties, such as its chemical structure, molecular weight and gelation behavior. However, no current studies focused on the induction effect of bacterial alginate in eliciting bioactive compounds such as phytochemicals in plants like the soybeans. One of the best and longest-studied defense responses of plants to infection is the induced accumulation of anti-microbial, low-molecular-weight secondary metabolites known as phytoalexins. The role of these phytoalexins compound in defense mechanism has been revealed through several experimental methods. Several isoflavonoid phytoalexins such as the glyceollins produced by soybeans are known to be estrogenic, with potential health effect in humans in reducing the risk of some diseases and cancer prevention. This present study reports the production and extraction process of bacterial alginate from A. vinelandii cultures and also investigates the induction effect of bacterial alginate in the elicitation of phytochemical glyceollin accumulations in treated soybean seeds as the main elicitor. The High performance liquid chromatography(HPLC) was used to detect and quantify extracted samples, Ultra performance liquid chromatography-mass spectrometry(UPLC-ESI-MS) was used to detect the molecular weight of the hydrolyzed bacterial alginate and induced glyceollin samples, and the thin layer chromatography(TLC) was used to detect the guluronic(G) and mannuronic(M) content of the hydrolyzed bacterial alginate and standard samples during this study. Optimal parameters were determined by investigating the effect of concentration of the elicitor(bacterial alginate), soak time, temperature, moisture and time course, and also a comparative study was carried out by using bacterial alginate extracted from A. vinelandii and A. oryzae respectively as elicitors to investigate effects of some parameters. The study recorded bacterial alginate production from A. vinelandii, while extracted quantity was(6.25 mg/mL for every 25 mL) from A. vinelandii production cultures and the determined relative molecular weight(Mw) of bacterial alginate obtained in this study is(Mw = 2.5 x 105 Da). TLC detection method was helpful in the identification of uronic acids(M and G) found present in the hydrolyzed bacterial alginate extracted samples. Bacterial alginate induced glyceollin synthesis in treated soybean seeds, and the optimal conditions developed for the elicitation process were as follows: concentration of bacterial alginate(0.24 mg/mL with a volume of 30 μL); soak time of soybean seeds in distilled water(5 h); controlled temperature(30 ℃); moisture(40 %) and incubation period(4 d). The optimal conditions developed during this study recorded quantitation of glyceollins as follows:(1.775 mg/g dry weight) for bacterial alginate treatment of soybean seeds,(0.012 mg/g) for alginate from seaweed source treatment in soybean seeds, while for the un-treated but wounded soybean seeds recorded(0.154 mg/g). The method adopted in this study for bacterial alginate production, extraction and its application in phytochemical glyceollin induction may be of practical use in the food and pharmaceutical industry to develop nutraceutical products. These findings identify the effects of bacterial alginate as an inducer of phytochemical glyceollin accumulation in soybean seeds following optimal condition protocol. However, phytochemical glyceollin merit further research as a potential therapeutic agent for human health and the introduction of such bioactive substances in food products beneficial to human health is strongly encouraged and recommended. These findings offer future and new prospects for wider application of bacterial alginate extracted from A. vinelandii as an inducer which could be utilized in the formulation of bioactive foods in the food industries or introduced in pharmaceutical products.