Study On Carotenoids Fermentation By Rhodotorula Mucilaginosa

In this study,a wild-type red yeast previously screened by this laboratory was used as the starting strain.Through morphological observation,26s rDNA sequencing,and physiological and biochemical tests,it was identified as Rhodotorula mucilaginosa,and was named as Rhodotorula mucilaginosa WZW003.Then the optimal solution for carotenoids extraction was investigated and employed for subsequent product analysis belonged to media optimization.The single-factor optimization of seed media and fermentation media and the RSM optimization of fermentation media was sequentially involved in the optimization of synthetic media,and the final optimized result was applied for scale-up experiment in a 5 L fermentor.Consequently,the feasibility of producing carotenoid-enriched feed-staff by Rhodotorula mucilaginosa WZW003 utilizing cheap media dominated by tomato pomace was investigated and optimized through comprehensive application of single-factor method and uniform design.The main results were as follows:1.The optimal abstraction method concerning the carotenoids inside the cells of Rhodotorula mucilaginosa WZW003 was:DMSO was to extract it under 60℃ and stirring,with material to solvent ratio of 1:60(m/V).The extraction duration was 1 h.After that,the extraction mixture was centrifuged and the supernatant was taken.Ethyl ether was added to it with a ratio of 1:8(V/V)to perform liquid phase extraction.Subsequently,the ethyl ether phase was taken and applied for calculation of total carotenoids content obeying the principle of molar extinction.2.The optimal seed media composition and culture conditions were:glucose 45 g/L,peptone 15 g/L,initial pH 5.0,loading volume in 250 mL shaking flask was 40 mL,culture temperature was 30℃,agitation speed of rotary shaker was 150 rpm.3.The optimal synthetic fermentation media composition and culture conditions were:glucose 45 g/L,peptone 9.75 g/L,NaH2PO4·2H2O 8.26 g/L,initial pH 4.11,loading volume in 250 mL shaking flask was 30 mL,culture temperature was 30℃,agitation speed of rotary shaker was 150 rpm,inoculum age of seed liquid was 48 h,inoculum size was 10%,fermentation period was 72 h.Under such condition,the total carotenoids concentration yielded in shaking flask fermentation was 103.02 mg/L.The maximum value of total carotenoids concentration yielded in the scale-up experiment in 5 L fermentor was 533.72 mg/L.The scale-up experiment of synthetic media indicated that the production of total carotenoids yielded by fermentation of Rhodotorula mucilaginosa WZW003 increased,mainly resulting from the increment of biomass concentration,however,the cellular total carotenoids content represented downward trend actually.This might illustrate that the biomass concentration concentrated very much to the concentration of target product,and had big leeway of increment.In the fermentation broth,the numerical variation of biomass concentration and total carotenoids concentration had huge relationship with residual sugar and pH.4.The optimal tomato pomace fermentation media composition and culture conditions were:cellulase additive amount 12 g/L,cellulase enzymolysis duration 54 h,tomato pomace additive amount 50 g/L,soybean meal additive amount 10 g/L,MgSO4·7H2O additive amount 8 g/L,initial pH 4.5,culture temperature was 30℃,inoculum age of seed liquid was 48 h,inoculum size was 10%.Under such condition,the maximum value of total carotenoids concentration yielded in scale-up experiment in 5 L fementor was 89.9 mg/L.The scale-up experiment of tomato pomace media indicated that the residual sugar in fermentation broth of Rhodotorula mucilaginosa WZW003 was nearly exhausted in the 48 h,however the total carotenoids concentration could still maintain increasing henceforward.This was clearly because of the media contained tomato pomace,whose enzymatically hydrolyzed products could be utilized by Rhodotorula mucilaginosa and thus keep its carotenoids biosynthesis approach.