| Aldehydes belong to the carbonyl group and have a great influence on the flavor stability of beer.More than 200 carbonyl compounds have been found in beer,among which acetaldehyde is the most concentrated,accounting for about 60% of the total aldehyde content.It produces the flavor of freshly cut green apples,grass,or green leaves in beer.High concentrations of acetaldehyde can lead to a spicy,grassy taste in beer.The high content of acetaldehyde will lead to the rapid aging of beer,which will have a great impact on the quality of beer.Yeast metabolism is the main source of acetaldehyde.Therefore,screening brewer’s yeast with low acetaldehyde production is the most fundamental method to reduce the acetaldehyde content in finished beer.The main findings of this paper are as follows:(1)In this study,firstly,spectrophotometry and headspace injection gas chromatography were compared to determine the content of acetaldehyde in beer.The method of headspace injection gas chromatography was more accurate,with a detection limit of 0.5 mg/kg,R~2 =0.9995,greater than0.998,and a high correlation.The recovery rate was between 95% and 102%,and the overall reproducibility of the experiment was good.(2)Yeast strains were initially screened by using ARTP mutagenesis and malt juice AGAR plate containing fomeprazole.The selected strains were acclimated in the laboratory.According to the changes in the activities of alcohol dehydrogenase and acetaldehyde dehydrogenase,the mutant strain F1-2 with lower acetaldehyde production was obtained.Firstly,the growth performance and fermentation indexes of the initial strains F1-F5 were compared,and the best initial strain was F1.Compared with the other four initial strains,the initial strain F1 had the best growth performance,and the acetaldehyde content in the fermentation broth was 14.31 mg/L,which met the requirements of beer fermentation.Then,17 mutant strains were screened by ARTP mutagenesis.The content of acetaldehyde in the fermentation broth of mutant strain F1-2 was 2.56 mg/L±0.01mg/L,which was 82.40% lower than that of the original strain F1.The aldehyde dehydrogenase Ⅱactivity of F1-2 mutant strain decreased to 71.22%.To further investigate the fermentation performance of the mutagenic strain,a 200 L fermentation pilot experiment was carried out on the initial strain F1 and the mutagenic strain F1-2.Through the experiments on the changes of p H,diacetyl and acetaldehyde of the two strains,it was found that the overall fermentation performance of F1-2 was higher than that of the initial strain F1,the beer flavor was more coordinated,and the storage time was prolonged.(3)In the subsequent molecular experiments on yeast,the cleavage of yeast cell wall is an important step.Through four methods to break the wall of yeast,the results showed that the genome extracted by the ultrasonic method had the highest content,but the purity was not high and the quality was poor,which had an impact on the subsequent molecular experiments.The concentration and purity of genome extraction by glass beads method are moderate,and the price of glass beads is cheap and economical.(4)In the acetaldehyde metabolic pathway,aldehyde dehydrogenase II is closely related to the level of acetaldehyde.In this study,by comparing the original strain F1 with the mutant strain F1-2,it was found that after ARTP mutagenesis,the mutant strain F1-2 had nine amino acid mutations in alcohol dehydrogenase II compared with the initial strain,namely D14 E,A75G,I76 M,D78E,C148 E,V152I,S168 I,V174A,D14 E and P179 A.These results demonstrated that the mutant strains with strong genetic stability could be screened by the combination of ARTP and ALE,which could be applied to practical production.At the same time,the determination of acetaldehyde in beer by headspace injection is also more accurate.The glass bead method is the best way to extract the yeast genome.Compared with other wall-breaking methods,the quality of the extracted genome is more suitable for later molecular experiments. |
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