The bacterial community and the metabolic activities involved at the flavor-forming stage during the fermentation of shrimp sauce were investigated using metatranscriptome and 16S rRNA gene sequencings. Results showed that the abundance of Tetragenococcus was 95.1%. Tetragenococcus halophilus was identified in 520 of 588 transcripts annotated in the Nr database. Activation of the citrate cycle and oxidative phosphorylation, along with the absence of lactate dehydrogenase gene expression, in T. halophilus suggests that T. halophilus probably underwent aerobic metabolism during shrimp sauce fermentation. The metabolism of amino acids, production of peptidase, and degradation of limonene and pinene were very active in T. halophilus. Carnobacterium, Pseudomonas, Escherichia, Staphylococcus, Bacillus, and Clostridium were also metabolically active, although present in very small populations. Enterococcus, Abiotrophia, Streptococcus, and Lactobacillus were detected in metatranscriptome sequencing, but not in 16S rRNA gene sequencing. Many minor taxa showed no gene expression, suggesting that they were in dormant status.
The chitinase‐producing bacteria Paenibacillus sp. was isolated from soil samples. The chitinase was purified successively by ammonia sulfate fractional precipitation followed by chromatography on DEAE 52‐cellulose column and then on Sephadex G‐75 column. The chitinase has a molecular weight of ca. 30 kDa as measured by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS‐PAGE) electrophoresis. Its optimum pH is 4.5, and its optimum temperature is 50 °C with colloidal chitin as a substrate. The enzyme is stable below 45 °C and in pH ranges between 4.5 and 5.5. It is activated by glucosamine, glucose, N‐acetylglucosamine, and metal ions including Ca2+, Fe2+, Fe3+, and Ni2+. It is inhibited by SDS, H2O2, ascorbic acid, Cu2+, Mg2+, Ba2+, Sn2+, Cr3+, and K+. With colloidal chitin as substrate, the Km and the Vmax of the chitinase are 4.28 mg/mL and 14.29 μg/(Min·mL), respectively, whereas the end products of the enzymatic hydrolysis are 14.33% monomer and 85.67% dimer of N‐acetylglucosamine. The viscosity of carboxymethyl chitin decreased rapidly at the initial stages when subjected to chitinase hydrolysis, which indicates that the chitinase acts in an endosplitting pattern.
This research intends to reduce the crystallinity of chitin with physical methods so as to improve the enzymatic hydrolysis efficiency of chitin. Scanning electron microscopy showed that both steam explosion and high‐pressure homogenization significantly enlarged the pores in shrimp shells, which are made up of chitin, but γ radiation did not. X‐ray diffraction showed that the crystallinity index of chitin decreased by 23.2% in the (110) plane and 24.7% in the (020) plane after three rounds of steam explosion and decreased by 12.4% in the (110) plane and 28.7% in the (020) plane after three rounds of high‐pressure homogenization. After γ radiation, however, there was no significant change. The corresponding deacetylation degrees of the chitin treated with steam explosion and high‐pressure homogenization increased by 13.2 and 5.7%, respectively. The natural, steam‐exploded and high‐pressure homogenized chitins respectively produced 0.251, 0.145, and 0.407 mg/ml reducing sugars with the hydrolysis of Streptomyces griseus chitinase. However, when hydrolyzed by Paenibacillus sp. A1 chitinase the steam‐exploded and the high‐pressure homogenized chitin respectively released 40 and 300% more reducing sugars than the natural chitin. The results found that steam explosion and high‐pressure homogenization can reduce the crystallinity of chitin, and make it prone to enzymatic hydrolysis.
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