Daqu is an essential starter for the brewing of Shanxi Aged Vinegar, and high contents of total acid and total ester are two important indices for highly qualified vinegar. The present study is amied to investigate the aromaenhancing role of a yeast strain screened from Daqu in fortified vinegar fermentation. A Daqu-originated yeast strain Y14 showed the highest esterproducing ability, up to 36.055 g/L, and was identified as Pichia manshurica by sequence analysis of the D1/D2 domain of the 26S rRNA gene in combination with morphological and physiological properties. When it was reinoculated into the Daqu-based fermentation, Gas Chromatography Mass Spectrometry (GC/MC) analysis for alcohol fermentation showed a significant increase of total alcohol and total ester. The levels of ethanol, ethyl acetate, and isoamyl acetate increased from 31.912%, 9.839%, and 1.025% to 39.861%, 22.707%, and 1.291%, respectively. Moreover, an additional seven volatile aroma components appeared in the Y14 fortified fermentation (FF). Correspondingly in the resulting vinegar, the total acid and total ester markedly increased from 37.4 and 15.3 g/L to 38.0 and 21.5 g/L, respectively. Sensory assay revealed no abnormal odour but a distinct fruity, flowery fragrance and a lingering ester taste in the vinegar of the FF. With this approach, Pichia manshurica isolated from Daqu, when reintroduced into the Daqu-based fermentation, can significantly increase the acid and the ester production, improve the vinegar quality by endowing it flowery and fruity notes, and a long-lingering buttery and nut-like taste.

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Both the calcium and collagen in bone powder are hard to be absorbed by the body. Although enzymatic hydrolysis by protease increased the bio-availability of bone powder, it was a meaningful try to further increase Ca2+ release, oligopeptide formation and antioxidant activity of the sheep bone hydrolysate (SBH) by lactic acid bacteria (LAB) fermentation. Lactobacillus helveticus was selected as the starter for its highest protease-producing ability among 5 tested LAB strains. The content of liberated Ca2+ was measured as the responsive value in the response surface methodology (RSM) for optimizing the fermenting parameters. When SBH (adjusted to pH 6.1) supplemented with 1.0% glucose was inoculated 3.0% L. helveticus and incubated for 29.4 h at 36℃, Ca2+ content in the fermented SBH significantly increased (p<0.01), and so did the degree of hydrolysis and the obtaining rate of oligopeptide. The viable counts of L. helveticus reached to 1.1×1010 CFU/mL. Results of Pearson correlation analysis demonstrated that LAB viable counts, Ca2+ levels, obtaining rates of oligopeptide and the yield of polypeptide were positively correlated with each other (p<0.01). The abilities of SBH to scavenge the free radicals of DPPH, OH and ABTS were also markedly enhanced after fermentation. In conclusion, L. helveticus fermentation can further boost the release of free Ca2+ and oligopeptide, enhance the antioxidant ability of SBH. The L. helveticus fermented SBH can be developed as a novel functional dietary supplement product.

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Cadmium (Cd) is a toxic metal leading to multiple forms of organ damage. Zinc (Zn) was reported as a potential antagonist against Cd toxicity. The present study investigates the antagonistic effect of Zn (20 μM) on Cd (20 or 50 μM) cytotoxicity in macrophages in vitro. The results shows that Cd exposure caused dose-dependent morphologic and ultrastructural alterations in RAW 264.7 macrophages. Zn supplement significantly inhibited Cd cytotoxicity in RAW 264.7 or HD-11 macrophages by mitigating cell apoptosis, excessive ROS output, and mitochondrial membrane depolarization. Notably, Zn supplement for 12 h remarkably prevented intracellular Cd2+ accumulation in 20 μM (95.99 ± 9.93 vs 29.64 ± 5.08 ng/106 cells; P = 0.0008) or 50 μM Cd (179.78 ± 28.66 vs 141.62 ± 22.15 ng/106 cells; P = 0.003) exposed RAW 264.7 cells. Further investigation found that Cd promoted metallothioneins (MTs) and metal regulatory transcription factor 1 (MTF-1) expression in RAW 264.7 macrophages. Twenty μM Zn supplement dramatically enhanced MTs and MTF-1 levels in Cd-exposed RAW 264.7 macrophages. Intracellular Zn2+ chelation or MTF-1 gene silencing inhibited MTs synthesis in Cd-exposed RAW 264.7 macrophages, which was accompanied by the declined expression of MTF-1, indicating that regulation of Zn on MTs was partially achieved by MTF-1 mobilization. In conclusion, this study demonstrates the antagonism of Zn against Cd cytotoxicity in macrophages and reveals its antagonistic mechanism by preventing Cd2+ bioaccumulation and promoting MTs expression.

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