طراحي و بهینه سازی فرآیند تولید و خالص سازی مانان از مخمر کلویورومایسس مارکسیانوس به منظور استفاده به عنوان ادجوانت واکسن

Mannan glycoprotein consist of a branched polymer of mannose six carbon sugar and is located on extra layer of cell wall that its sugar is linked to protein by conjunctional groups. Mannan is a natural polymer that can have adjuvant effect by depot impact by using several polymer figures (small particle, solutions and gel) and increasing stimulating signals for better activity. Studies showed that mannan is a NF-kB activator, 1β interleukin inducer, interleukin 6 and TNF-α. The cell wall of kluyveromyces Marxianus yeast can e a very good resource for extracting mannan with high efficiency. Background and Objectives: Mannan is a linear glycoprotein with β-1,4 bonds that binds mannose units to proteins. It contains natural amphiphiles and acts as a bio-emulsifier. Compounds with emulsifiers' types and structures can be used as adjuvants in AS03 (Influenza-pandemic) vaccines and facilitate the process of local production of cytokines and absorption of intrinsic cells. This study was performed to achieve these goals: optimizing the growth of the yeast Kluyveromyces marxianus (IBRC-M 30114) in the fermenter (bio-reactor) to its maximum extent, designing an optimized method for extracting mannan with three methods of homogenization, alkalinity and optimized bio-emulsifier, and data modeling at three levels based on three factors of acidity, temperature, and the mannan extraction method using QUALITEK-4 software and the Taguchi method in 3 runs with 9 parts derived from firmament biomass that could be used as a biological natural emulsifier as an adjuvant. Methodology: Homogenization, alkaline, and bio-emulsifier methods with various pHs and temperatures were performed to optimize the production and extraction processes of mannan glycoprotein. Mannan was extracted (in 9 runs with 3 parts) by performing the QUALITEK-4 software and the Taguchi method. In the designed stage, various temperatures (25 °C, 30 °C, and 35 °C) and pHs (2, 5, and 7) were used in the Homogenization, alkaline, and bio-emulsifier methods. Findings and Conclusion: Given the growth outcomes of the yeast Kluyveromyces marxianus in a fully optimized condition inside the Erlenmeyer (optimized factors for the yeast growth included 55.15 g of glucose/l, 9.35 g of yeast extract/l, pH 4.99, and fermentation time = 168 hours) after transferring to the fermenter, the highest biomass was obtained. The growth of the yeast kluyveromyces marxianus in a completely optimized condition in the fermenter showed that the maximum amount of the biomass yielded from the scale increase inside the reactor manifested that the maximum amount of the fermentation biomass extracted from the yeast Kluyveromyces marxianus was 34 (g.l-1), in 30 hours, pH 4.6, 29 °C, 500 rpm, oxygen 19.7, inlet air volume 1(vvm)3, and 36.6 DO. Also, the maximum amount of mannan was extracted by the alkaline method with pH 5, which was 8.243 mg (100 ml)-1 mannan from 10g yeast biomass. In the bio-emulsifier method, the maximum amount of mannan extraction occurred at pH 7. Mannan extraction by the alkaline method in pH 5, compared to pH 2 and 7, was significantly higher. The extraction and purification processes by the homogenization method in pH 7 produced the maximum amount of output. The maximum amount of purified mannan was taken by the alkaline method at pH 5 and 25 °C.