| Pumpkin pentose is a kind of precious monosaccharide in Pumpkin, which has not been developed and utilized widely for its low content. However, there are a great number of six-carbon sugars and polysaccharides in Pumpkin, which could be transformed to pentoses by biotechnology. Pumpkin pentose can be metabolized in human body without insulin and it can be used to explore functional food and medicines for diabetes and obese patients, which will greatly increase the additional post-harvest value of pumpkin. Strain J503and B106were screened from1635Bacillus which can transform glucose to pentose. In this study, hexose was used for biotransformation by Bacillus subtilis B106and J503, which was deteced by HPLC-MS, E-Nose and GC-MS for content and type of pentose. By proteomics method, key protein analysis by LC-MS, protein homologous analysis, key enzyme proteins of pentose and metabolic were deduced, which provided a scientific basis for the functional development of the functional pumpkin products.The main results and conclusions were made as follows:(1Quantitative of pentose (xylose and arabinose) was detected by HPLC, glucose and mannose were detected could be converted to pentose (xylose and arabinose). Strain J503was chosen to convert glucose to generate pentose. Response surface method was used to optimise the fermentation condition: Fermentation temperature for37℃， fermentation time for36h, the substrate glucose concentration for3.4%, pH value for7. Under this condition,2.26±0.03mg/mL pentose had been fermentated by strain J503, which was very close to the predicted value equation2.28mg/mL.(2) By solid phase micro extraction (SPME) enrichment, gas chromatography-olfactometry resolution-mass spectrometry (GC-O-MS) analysis, characterized volatile products of Maillard reaction of xylose and four amino acids (cysteine, glycine, leucine, and days aspartic acid) were furfural, naphthalene, butylated hydroxytoluene,(2-methyl-furan-),(1-Propanone,1-(2-furanyl)-),(Ethanone,1-(2-furanyl)-),(phenyl methoxy oxime) and (2-Isoamyl-6-methylpyrazine). The main nine volatile components from the reaction for leucine with xylose and fermentation products by Strain J503and B106were acetone,2-methyl furan,3-methyl n-butyl aldehyde,3-methyl butyric acid,5-methyl hexanone,6-methyl-2-4-methyl heptyl methadone, pentanoic acid,2-isoamyl-6-methyl pyrazine and butylated hydroxytoluene.The main eight volatile components from the reaction for glycine with arabinose and fermentation products by Strain J503and B106wereacetone aldehyde, furfural, benzaldehyde,1,5-dinitrogen miscellaneous-5-ketone, nonyl aldehyde and2,4-tertiary butyl phenol, phthalic acid different butyl octyl ester and butyl phthalate. According to the main content of aroma components, the pentose contents of fermentation by glucose were higher than those fermentation by mannose using J503and B106. The generated pentose order of all samples is (J503+glucose)>(J503+mannose)>(B106+glucose)>(B106+mannose)>(J503+B106+glucose)>(B106J503++mannose)(3) Pentoses were more likely to have Maillard reaction than hexose, xylose were more likely to have Maillard reaction than arabinose, glucose were more likely to have Maillard reaction than mannose. The order of response intensity of color reaction of amino acid was Leu> Gly> Cys> Asp. The color of treated-group was lighter than control, the order of color difference was glucose and mannose fermentation by J503+B106> glucose and mannose fermentation by J503> glucose and mannose fermentation by B106.(4) Electronic nose could distinguish volatile components of Maillard reaction by different pentose and four amino acids (cysteine, glycine, leucine, and aspartic acid) and glucose, mannose, and pumpkin juice fermentation by Bacillus subtilis (J5O3and B106).The cumulative contribution of all the samples between the two main components was78%-85%, which could effectively responded effectively induction value of the sample. The results determined by dimensionality reduction of PCA were similar with headspace solid phase micro-extraction analysis of volatile components.(5) Glucose germentation by J503had the most protein species and content, the molecular weight of majority of the protein was below43KD, protein of26KD was;at the most content. Liquid-mass spectrometry (LC-MS) method was used to identify490kinds of pentose proteins which could participated in glucose conversion by J503. The database (NCBI/BLAST) matched seven proteins which may be involved in the transformation:①1,2-dihydroxy-3-keto-5-methylthiopentenedioxygenase [Bacillus amyloliquefaciens subsp. plantarum CAU B946],②XlyB [Bacillus amyloliquefaciens FZB42],③endo-xylanase [Bacillus subtilis subsp. inaquosorum KCTC13429],④sugar-phosphate epimerase/isomerase [Bacillus subtilis subsp. subtilis str.168],⑤arabinogalactan endo-1,4-beta-galactosidase [Bacillus amyloliquefaciens subsp. plantarum CAU B946],⑥glucuronoxylanase xynC [Bacillus amyloliquefaciens] and⑦PenP [Bacillus amyloliquefaciens FZB42]. The mechanism was deduced which converted six-carbon sugar pentose metabolism by strain J503. |
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