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Food Chemistry
Interaction Mechanism of Moderately Oxidized Myofibrillar Proteins with Selected Aldehyde Compounds
DAI Xinxin, LÜ Yichao, YIN Xiaoyu, DOU Qingzhe, KONG Baohua, CHEN Qian, QIN Ligang
2023, 44(18):  1-8.  doi:10.7506/spkx1002-6630-20221114-162
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A hydroxyl radical generating system consisting of 0.1 mmol/L FeCl3, 0.1 mmol/L ascorbic acid, and various H2O2 concentrations (0, 0.25, 0.5, 1.0, 2.5, 5, 10 and 25 mmol/L) was established and used to investigate the interaction mechanism of differently oxidized myofibrillar proteins (MPs) with 3-methyl butanal, pentanal, hexanal and heptanal. First, the binding capacity of oxidized MPs to the four aldehydes were determined by gas chromatography-mass spectrometry (GC-MS). Then, ultraviolet (UV) absorption spectroscopy, fluorescence spectroscopy (quenching mechanism and thermodynamic analysis) and circular dichroism (CD) spectroscopy were used to reveal the interaction mechanism between oxidized MPs and aldehydes. The results showed that heptanal had the strongest binding capacity with MPs, and the binding capacity of oxidized MPs with a H2O2 concentration of 1.0 mmol/L to all four aldehydes was the highest (P < 0.05). UV absorption spectra and fluorescence spectra demonstrated that oxidized MPs interacted with the four aldehydes via static and dynamic quenching. The maximum absorption peaks of both tyrosine and tryptophan residues were red-shifted with an increase in heptanal concentration, indicating that the tyrosine and tryptophan residues of MPs were exposed to a more hydrophilic environment. Thermodynamic analysis showed that the interaction between MPs and heptanal was mainly driven by hydrophobic interaction. The CD spectra showed that the α-helix content decreased from 19.24% to 16.88% (P < 0.05) and the β-sheet content increased from 24.59% to 26.47% (P < 0.05) with increasing heptanal concentration, and the structure of MPs changed from the ordered to the disordered state. This study provides a theoretical basis for flavor regulation of meat products.
Formation and Digestion Characteristics of Self-Assembled Complex of Starch and Pomegranate Peel Polyphenols
GUO Jinbiao, QU Yinghui, LI Xinyu, ZHANG Lei, HE Xihong, ZHANG Liming, HAO Limin
2023, 44(18):  9-17.  doi:10.7506/spkx1002-6630-20221114-159
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The purpose of this investigation was to explore the fabrication of a self-assembled complex of potato starch (PS) and pomegranate peel polyphenols (PP) by ball milling and to investigate its digestion characteristics. The impact of ball milling time, ball milling speed, and PS/PP mass ratio on the total polyphenol content and digestion characteristics of the self-assembled complex was investigated. Then, the formed product was characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD) spectroscopy, Fourier transform infrared (FTIR) spectroscopy, and thermogravimetric analysis (TGA). The results showed that the content of total polyphenols was (19.91 ± 0.32) mg/g in the product obtained under the following conditions: milling time 10 h, rotational speed 500 r/min, and mass ratio of PS to PP 1:0.15. The SEM results showed that many cracks and gullies appeared on the surface of the product, which was accompanied by an agglomeration phenomenon. The XRD results showed that the crystalline domain of starch was destroyed by ball milling, and the intensity of the diffraction peak became weakened or even disappeared, indicating that the granules were changed into an amorphous structure. The FTIR results showed that the absorption peak at 3 600–3 400 cm-1 was enhanced, which illustrated that PP and PS were self-assembled through hydrogen bonds. Hence, ball milling can be used to fabricate PS/PP self-assembled complex. The TGA results demonstrated that the thermal stability of the self-assembled complex was improved. Furthermore, in vitro simulated digestion and in vitro simulated gastrointestinal release tests showed that the proportion of resistant starch in the complex was significantly increased, which was beneficial to maintain the stability of the guest molecules during the digestion process, and allowed the targeted release of polyphenols in the large intestine and colon, enabling the guest molecules to exert their functions better, thereby improving their bioavailability.
Preparation and Evaluation of Hydrophobic Chitosan Aerogel for Loading and Sustained Release of Curcumin
XU Dingyu, JIAO Siyu, YAO Xianchao, LIU Xin, CHEN Lifen, LIN Rihui
2023, 44(18):  18-25.  doi:10.7506/spkx1002-6630-20221009-075
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Hydrophobic chitosan aerogels were prepared using chitosan as a matrix and 2,6-dimethyl-5-heptenal as a cross-linking modifier to graft n-octadecanethiol and were characterized by scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy, and X-ray photoelectron spectroscopy (XPS). The results showed that the surface of the hydrophobic chitosan aerogel had a three-dimensional porous structure. 2,6-Dimethyl-5-heptenal was successfully grafted via the Schiff base reaction. The proportion of elemental sulfur was 1.17% in the aerogel. In the surface contact angle test, the oil droplet contact angle was 0°, and the water droplet contact angle was 126°. The chitosan aerogel exhibited good hydrophobic properties. The loading of fat-soluble curcumin in the aerogel reached the maximum value of 70.5 mg/g after 40 min, and the optimal loading temperature was 27 ℃. The hydrophobic chitosan aerogel had a good sustained-release performance, and the sustained-release rate of curcumin in simulated intestinal fluid was up to 89.6%.
Effect of Extraction pH on Gelling Properties and Characteristic Peptide Identification of Pigskin Gelatin
SHA Xiaomei, YAN Nongyang, CHEN Wenmei, XIE Zuohua, LU Ling
2023, 44(18):  26-33.  doi:10.7506/spkx1002-6630-20220903-027
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In order to study the effect of extraction pH on gelling properties and characteristic peptide identification of pigskin gelatin, the molecular mass distribution and gelling properties of pigskin gelatin extracted under different pH conditions were studied by dodecyl sodium sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), a texture analyzer and a rheometer. Moreover, the characteristic peptides were identified by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The results showed that with a decrease in pH, the relative molecular mass of pigskin gelatin initially increased and then decreased, the gel intensity gradually declined, and the gel melting temperature and gelling temperature initial increased and then decreased. Gelatin extracted at pH 1 had the worst gelling properties. HPLC-MS/MS analysis showed that pH significantly affected the traceability of gelatin. In this study, 62, 71, 79 and 76 characteristic peptides were detected from pigskin gelatins extracted at pH 1, 3, 5 and 7, respectively. Among them, 37 characteristic peptides were common to all pigskin gelatins. Compared with our previous research, 17 characteristic peptides were found to be common to pigskin gelatins extracted under different conditions. These stable common characteristic peptides could be used as an important basis for pigskin gelatin traceability with high accuracy.
Inhibitory Effect of Resveratrol on Thermally Induced Trans Fatty Acids in Peanut Oil
LI Tian, GUO Qin, QU Yang, LIANG Manzhu, ZHANG Qiaozhen, QIN Jingjing, HUANG Xuegang, ZHANG Yu, WANG Qiang
2023, 44(18):  34-39.  doi:10.7506/spkx1002-6630-20220826-318
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The inhibition of resveratrol on the formation of thermally induced trans fatty acids in peanut oil and its structure-activity relationship were studied by gas chromatography (GC), liquid chromatography (LC), and theoretical calculation. The results showed that the content of resveratrol decreased with an increase in temperature, and too high temperatures caused a cut-off effect. Resveratrol significantly inhibited the formation of trans fatty acids in peanut oil, and the inhibition rate of total trans fatty acids decreased with increasing temperature. The highest inhibition rate of cis-trans isomerization of fatty acids was 30.30%. The inhibitory effect could be attributed to the energy barrier formed by reducing the rate of isomerization reaction and increasing the amount of trans fatty acids. Its quantitative parameters (intrinsic thermodynamic energy and total entropy) determined the anti-isomerization effect. Moreover, the structure-activity relationship for the anti-isomerization effect was established, which will lay a theoretical foundation for the precise regulation of trans fatty acids.
Preparation and Properties of Zein Loaded Phytosterol Nanoparticles
YANG Tingting, REN Lishun, CHEN Guangwei, HUANG Aixiang
2023, 44(18):  40-48.  doi:10.7506/spkx1002-6630-20220818-222
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This study was undertaken in order to improve the stability, water solubility and gastrointestinal release of phytosterols (PS). Zein-PS nanoparticles were prepared by the anti-solvent method. Single factor experiments and orthogonal array design methods were adopted to optimize the preparation conditions based on the stability and encapsulation efficiency of zein-PS nanoparticles. The structure and functional properties of the prepared samples were characterized by using scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) and simulated gastrointestinal digestion. The results showed that the optimal preparation conditions were determined as follows: zein/PS ratio of 15:1 (m/m), hydration time of 2 h, hydration temperature of 55 ℃, and ultrasonic treatment time of 20 min. The encapsulation efficiency of the nanoparticles prepared using the optimal conditions was 84.97%, particle size (479.76 ± 0.38) nm, and zeta potential (−22.79 ± 0.015) mV, suggesting small particle size and high stability. The nanoparticles with a zein/PS ratio of 15:1 had better re-solubility and water solubility. SEM revealed the formation of a compact network structure on the surface of the nanoparticles and FTIR spectra showed that it changed at 3 313.16 and 1 523.51 cm-1, indicating the presence of hydrogen bonding and electrostatic interaction between zein and PS. Compared with PS, the release rates of the nanoparticles in simulated gastrointestinal fluids were reduced by 49.03% and 28.11%, respectively, showing a good sustained-release effect. In addition, after storage for 30 days, the change in the particle size of the nanoparticles was smaller at 4 ℃ than at 25 ℃, and the encapsulation efficiency remained above 70%. Therefore, zein-PS composite nanoparticles have good stability and sustained release property, which has potential application prospects in the food industry.
Stability and Separation of Peptides from Seabuckthorn Seed Protein
XIANG Huan, CUI Chun
2023, 44(18):  49-57.  doi:10.7506/spkx1002-6630-20221201-003
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In this study, peptides with inhibitory activity against porcine pancreatic lipase (PPL) from the alcalase hydrolysate of seabuckthorn seed meal protein were purified by ultrafiltration and macroporous resin separation, and their structures were investigated by ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and molecular docking. The peptides had good thermal and pH stability. The PPL inhibitory activity was significantly improved by adding appropriate amounts of Na+ and Mg2+ but not influenced by short-term exposure to the air. A total of 31 peptides were identified by UPLC/MS-MS, and six peptides were selected by molecular docking, whose contents in the ultrafiltration fraction with molecular mass less than 3 kDa were as follows: VR (2.90%), FR (7.40%), RDR (1.10%), APYR (1.50%), NLLHR (1.40%) and EEAASLR (1.10%), respectively. The half-maximal inhibitory concentration (IC50) values of VR, FR, RDR, APYR, NLLHR and EEAASLR prepared by solid phase synthesis were 371.07, 243.07, 250.50, 350.41, 220.70, and 510.55 μg/mL, respectively. The results of molecular docking showed that each of these peptides could combine with PPL by hydrogen bonding and π-π stacking interactions. Pearson correlation analysis showed that there was a positive correlation between molecular binding energy and the PPL inhibitory activity of the peptides from seabuckthorn seed protein (R2 = 0.865, P < 0.05).
Bacillus subtilis B-2 Improved the Quality of Low-Salt Fermented Fish Sauce through Regulating the Microbial Community
LI Yan, LI Wenjing, LI Chunsheng, YANG Daqiao, WANG Yueqi, WANG Di, CHEN Shengjun, WU Yanyan, LI Laihao
2023, 44(18):  58-67.  doi:10.7506/spkx1002-6630-20221226-246
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In this study, the effect of fermentation with Bacillus subtilis B-2, a salt-tolerant strain capable of producing high yield of protease, on the microbial community and the contents of biogenic amines and amino acid nitrogen (AAN) during the fermentation of low-salt fish sauce. The results of high-throughput 16S rRNA gene sequencing showed that fermentation with B-2 reduced the richness and evenness of the microbial community, Bacillus was dominant throughout the fermentation process and the abundance of spoilage microorganisms significantly decreased. B-2 significantly inhibited the formation of histamine, putrescine, cadaverine, and tyramine, and their contents at the end of fermentation decreased by 25.9%, 35.6%, 23.6% and 9.3%, respectively, compared with those in naturally fermented fish sauce. The content of AAN in low-salt fish sauce was significantly improved by inoculated fermentation, reaching 1.23 g/100 mL after 15 days of fermentation, which was significantly higher than that of naturally fermented fish sauce (0.79 g/100 mL). The correlation network map showed that the decrease in the abundance of Brevibacterium, Dietzia, Paracoccus, Aequorivita, and Brachybacterium was the major reason for the decrease in microbial diversity at the late stage of fermentation of naturally fermented fish sauce. The abundance of Stenotrophomonas showed a significantly positive correlation with the contents of many biogenic amines in naturally fermented and B-2 fermented fish sauce, suggesting its important role in the formation of biogenic amines in low-salt fish sauce. Comparative analysis of the microbial community and quality attributes at the end of fermentation showed that the metabolism of B-2 was the main reason for the decrease in the species and abundance of spoilage microorganisms, the increase in AAN content, and the decrease in the contents of key biogenic amines. B. subtilis B-2 is expected to be developed as a special fermentation starter for fish sauce to improve the quality and safety of rapidly fermented low-salt fish sauce.
Identification and Genome Analysis of Laceyella sacchari FBKL4.014 Isolated from Moutai-Flavor Daqu
TIAN Haojie, LI Dounan, QIU Shuyi, ZHOU Jianli, LONG Zehe, WANG Kejia, LIU Maoqiang, CHEN Jie, CHENG Du, PAN Fengshuang
2023, 44(18):  68-76.  doi:10.7506/spkx1002-6630-20220928-319
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A thermophilic strain of Laceyella sacchari designated as FBKL4.014 was isolated from high-temperature Moutai-flavor Daqu, and its whole genome was sequenced. The results showed that the full genomic length of L. sacchari FBKL4.014 was 3.35 Mb, with a GC content of 48.67%. The sequencing depth was 433 ×, and a total of 3 328 protein-coding genes were predicted. Strain FBKL4.014 had the potential for complete carbohydrate and amino acid metabolism, and it was further inferred that the strain may have the potential to biosynthesize tetramethylpyrazine. This study provides genetic data for understanding the mechanism for the production of flavor substances in Maotai-flavor baijiu by Thermoactinomycetaceae metabolism, and also provides a reference for further research on the discovery of unknown functional genes of this strain and its genetic modification.
Cloning, Expression and Functional Analysis of Lycopene Cyclase Gene from Maize
WANG Fanyu, HE Weiwei, LI Dajing, GUO Qingqi, LUO Hao, LU Yizhu, BAO Yihong, ZHANG Zhongyuan
2023, 44(18):  77-84.  doi:10.7506/spkx1002-6630-20221111-119
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In this study, the lycopene β-cyclase (LCYb) and lycopene ε-cyclase (LCYe) genes were cloned from maize, and the encoded products were analyzed by bioinformatics methods. After expression in Escherichia coli, the catalytic properties of LCYb and LCYe from maize were explored by color complementation and product analysis experiments. The results of sequence analysis showed that the full-length cDNA of maize LCYb and LCYe were 1 470 and 1 611 bp, respectively, which were more than 90% homologous to those of sorghum and millet. LCYe and LCYb proteins were successfully purified by fusion expression with glutathione thiotransferase tags. The results of color complementation test and high performance liquid chromatography (HPLC) analysis showed that maize LCYb had catalytic activity on β-ring, could cyclize both ends of lycopene to form β-carotene, and had very weak ε-ring catalytic activity, which could form α-carotene through the intermediate γ-carotene. Maize LCYe was also found to able to catalyze both ends of lycopene to form ε-carotene. This study can lay a foundation for exploring the molecular mechanism of the regulation of maize carotenoid.
Efficient Expression and Enzymatic Properties of L-Threonine Dehydrogenase from Escherichia coli
LIU Xinxin, WANG Yao, SHI Hongling, YAO Lunguang, WANG Xian, TANG Cunduo
2023, 44(18):  85-92.  doi:10.7506/spkx1002-6630-20221206-055
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Our aim was to improve the catalytic efficiency of L-threonine dehydrogenase (L-TDH) on L-threonine dehydrogenation to synthesize ethyl L-2-aminoacetate. An L-TDH gene from Escherichia coli was mined and solubly expressed in E. coli BL21(DE3) through pACYCDuet-1 expression system. The expressed enzyme was purified and characterized. The results showed that high-level soluble expression of L-TDH was achieved in E. coli BL21(DE3), and the enzyme activity in the lysate was 19.13 IU/mL, which was about 79 times higher than that the level of E. coli BL21 (DE3) background expression. The specific activity of the purified L-TDH was 12.77 IU/mg; its optimal reaction temperature was 45 ℃, and its optimal reaction pH was 9.0. The residual enzyme activity was still more than 90% after being held at 35 or 40 ℃ for 120 min. In addition, the kinetic parameters of EcTDH were better than those of other reported L-TDHs, and EcTDH was superior in the synthesis of 2,5-dimethylpyrazine (2,5-DMP) by converting L-threonine. Our findings could lay a theoretical foundation for the industrial production of 2,5-DMP.
Inhibition Activity and Mechanism of 1-Octen-3-ol on the Growth and Deoxynivalenol Biosynthesis of Fusarium graminearum
QIAN Shen’an, HU Zheng, YU Yinan, MENG Jiajia, ZHANG Zhiqi, HUANG Qingwen, ZHAO Zhihui, NIE Dongxia, HAN Zheng, FAN Kai
2023, 44(18):  93-101.  doi:10.7506/spkx1002-6630-20221223-227
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The inhibitory effect of fumigation with 1-octen-3-ol against the growth and deoxynivalenol (DON) biosynthesis of Fusarium graminearum PH-1 was determined, and the possible mechanism involved was investigated by evaluating cell membrane integrity, oxidative stress level and the expression of key genes related to DON biosynthesis. The results showed that 1-octen-3-ol significantly inhibited the mycelial growth, spore germination and mycotoxin production of F. graminearum in a concentration-dependent manner (P < 0.05). After 7 days of fumigation with 100 μL/L 1-octen-3-ol, the inhibition rates of mycelial growth, spore germination and deoxynivalenol production were 60.70%, 100.00% and 97.50%, respectively. Further studies showed that 1-octen-3-ol treatment effectively damaged the cell membrane integrity and increased the membrane permeability of F. graminearum, leading to a significant reduction of ergosterol levels and serious leakage of cell contents. Moreover, 1-octen-3-ol interfered with the oxidative stress balance of F. graminearum and downregulated the expression of DON biosynthesis-related genes such as TRI4, TRI5, TRI8, TRI10, TRI12, and TRI101. In conclusion, 1-octen-3-ol can effectively inhibit the growth and DON biosynthesis of F. graminearum by damaging cell membrane integrity, interfering with oxidative stress balance and regulating key gene expression.
Safety and Fermentation Characteristics of Meat-Derived Staphylococcus spp.
LIU Silu, CHEN Shanshan, SHAO Liangting, SHAO Xuefei, XU Xinglian, WANG Huhu
2023, 44(18):  102-109.  doi:10.7506/spkx1002-6630-20221111-118
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In this study, the physiology, biochemistry, safety and functional properties of Staphylococcus used as a starter culture were analyzed. Among the 33 strains of Staphylococcus, 11 tested positive for catalase and negative for hydrogen sulfide and their safety was evaluated by hemolysis, plasma coagulase and heat-resistant nuclease tests. In the functional study, 8 of the 11 strains were able to produce lipase and decompose fat and had high cholesterol degradation capacity. In the acid tolerance test, 4 strains were selected for strong acid resistance. The 4 strains could degrade casein, myofibrillar protein, sodium nitrite and some bioamines. They were identified as S. warneri 5F’-2, S. vitulinus 8A-1, S. warneri 5F-2 and S. succinus A31. These strains were positive for catalase, unable to produce hydrogen sulfide, and negative for hemolysis, plasma coagulase negative and heat-resistant nuclease. They had the functional properties of lipase production, cholesterol degradation, acid tolerance, protease production, myofibrillar protein degradation, nitrite degradation, and inability to degrade amino acids to produce bioamines, and biogenic amine degradation. Therefore, they had excellent fermentation potential, and could be used as strains for meat product fermentation.
Characterization of Enzymatic Properties of Single Mutants of Homoserine Dehydrogenase from Corynebacterium glutamicum
JIANG Zeyuan, LIU Yuzhe, GAO Xin, ZENG Qi, MIN Weihong
2023, 44(18):  110-116.  doi:10.7506/spkx1002-6630-20220830-353
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Site-directed mutagenesis was used to improve the catalytic activity of homoserine dehydrogenase (HSD) to reduce its feedback inhibition and repression by metabolites in the pathway. HSD was docked with the substrate homoserine, and its spatial structure was analyzed. Two key sites, Gly25 and Asp61, were selected for site-directed saturation mutation. The activity screening showed that the mutants A61L and G25G had significantly increased enzyme activity when compared to the wild type (WT). The kinetics and enzymatic properties of these two mutants were studied. It was found that compared to the WT enzyme, the Km values of G25G and A61L decreased, the substrate affinity increased, and the enzyme activity increased by 1.21 and 1.35 times, respectively; the n value decreased, and the positive synergy increased. The optimum temperature for A61L and G25G was 40 ℃, the same as that for WT; the optimum pH for A61L and WT was 8.0, which was lower than that (8.5) for G25G. The half-lives of A61L and G25G were 1 h longer and 0.5 h shorter than that of WT, respectively. Low concentrations of K+, Mg2+, and Ca2+ could activate the mutants and WT, while different concentrations of methanol, ethanol, acetonitrile and dimethyl sulfoxide had significantly inhibitory effects on the mutants and WT. At inhibitor concentrations of 1–25 mmol/L, the inhibitory effect was significantly weaker on the mutants than on WT. The mutants G25G and A61L showed improved enzyme activity and weakened allosteric inhibition. This study provides a reference for optimizing the biosynthetic pathway of HSD and constructing strains capable of producing high yield of methionine, threonine and isoleucine.
Application Potential of Native Meyerozyma guilliermondii in Pilot Production of Dry Red Wine
ZANG Wei, LIU Ye, LIU Yu, PENG Shuai, CHEN Xuelian, WANG Jing
2023, 44(18):  117-125.  doi:10.7506/spkx1002-6630-20220907-069
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The purpose of this study was to explore the application potential of Meyerozyma guilliermondii NM218 in dry red wine pilot production. Cabernet Sauvignon wines were made by sequential inoculation of NM218 and commercial Saccharomyces cerevisiae (FX10) at an interval of 48 h (treatment group) or single inoculation of FX10 (control group). The alcoholic fermentation process and yeast growth status were monitored, and the physicochemical indicators, color and aroma components of wine were measured at the end of alcoholic fermentation and after five months of aging. Moreover, sensory evaluation was carried out to the effect of mixed culture fermentation on the quality of ‘Cabernet Sauvignon’ dry red wine. The results showed that the viable count of M. guilliermondii remained higher than 105 CFU/mL before the middle stage of alcoholic fermentation, and completely disappeared at the end of fermentation. The process of alcoholic fermentation in the treatment group was prolonged by 2 to 3 days compared with that in the control group. The basic physicochemical indicators of the wine samples met the requirements of the national standard GB/T 15037-2006 Wines. Compared with the control group, the total anthocyanin content and the percentage of individual anthocyanins in the treatment group at the end of alcoholic fermentation and after five months of aging were significantly decreased (P < 0.05), and the CIELab color parameters and ionization index were significantly increased (P < 0.05). At the end of alcoholic fermentation, the contents of aroma substances such as ethyl butyrate (97%), isoamyl acetate (123%), ethyl caproate (59%), methyl caprylate (43%), phenylethyl acetate (61%), and citronellol (55%) in the treatment group increased compared with the control group. After five months of aging, the contents of aroma substances such as ethyl heptanoate (151%), diethyl succinate (63%) and ethyl myristate (83%) increased compared with the control group. Two aroma compounds were exclusively found in the treatment group, which could impart a rich fruity aroma to the wine. Sensory evaluation showed that the mixed culture fermented wine had a purple-red color, harmonious body, and rich aroma and scored high overall after five months of aging. In conclusion, sequential inoculation of NM218 and FX10 can enhance the aroma quality and sensory pleasure of dry red wine, and has the potential for industrial application.
Effect of Cyperus esculentus Dietary Fiber on the in Vitro Production of Short-Chain Fatty Acids by Human Intestinal Flora and Its Distribution Characteristics
WANG Lingyu, YAN Chunmei, JIA Meng, HUANG Jing, WANG Anqi, WANG Xuanyu, WANG Xintao, LI Xiuhe, WANG Huifang, DAI Fuhong, LI Ning, ZHOU Zhongkai, GAO Tiecheng
2023, 44(18):  126-132.  doi:10.7506/spkx1002-6630-20221007-047
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In this study, in vitro colonic fermentation and 16S rRNA high-throughput sequencing were used to explore the effects of the Cyperus esculentus dietary fiber on the production of short-chain fat acids (SCFAs) by the human gut microbiota and its structure. Soybean dietary fiber served as the control. The results showed that C. esculentus dietary fiber significantly promoted the production of SCFAs, but its effect was less pronounced than that of soybean dietary fiber. This study also found that high concentrations of dietary fiber significantly changed the diversity and richness of the gut microbiota, and the effects of the two dietary fibers were significantly different. C. esculentus dietary fiber significantly promoted the relative abundance of Firmicutes, Proteobacteria and Actinobacteria, but inhibited the growth of Bacteroidota. Its effect was slightly different from that of soybean dietary fiber. Moreover, compared with soybean dietary fiber, C. esculentus dietary fiber significantly had a stronger capacity to promote the growth and reproduction of Subdoligranulum, Bifidobacterium, Lactobacillus and Faecalibacterium, and significantly suppressed the growth of Prevotella and Bacteroides. This study highlights the potential of C. esculentus dietary fiber as a novel prebiotic.
Metagenomic Analysis of Microbial Diversity and Biogenic Amine Metabolism in Naturally Fermented Mutton Sausage
WU Shuanghui, NIU Yin, HE Jikun, DAO Xiaofang, TANG Junni, CHEN Juan
2023, 44(18):  133-140.  doi:10.7506/spkx1002-6630-20221024-252
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The changes in biogenic amine contents of naturally fermented mutton sausage samples during different fermentation stages were detected by high performance liquid chromatography (HPLC), and the microbial community succession was explored using metagenomics. Meanwhile, the correlation between biogenic amines and microbial diversity was established, and the abundance and functional annotation of microorganisms and enzymes were investigated by means of the Non-Redundant (NR) Protein Sequence database, evolutionary genealogy of genes: Non-supervised Orthologous Groups (eggNOG) database and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. The results showed that biogenic amine contents increased first and then decreased with fermentation time, and the content of spermine was the highest among eight tested biogenic amines. A total of 43 phyla, 60 classes, 112 orders, 201 families, 465 genera and 2 156 species of microorganisms were identified in fermented mutton sausage samples. Staphylococcus, Vibrio, Oenococcus and Haemophilus were the dominant bacteria. Spearman correlation analysis showed that the contents of tryptamine, cadaverine, tyramine, spermidine, and spermine were significantly (P < 0.05) related to some of the top 30 most abundant bacterial genera, while the contents of histamine, putrescine, and phenylethylamine were not. After analyzing the metagenomic data using the KEGG database, biogenic amine metabolic pathways and related enzymes and bacteria were found, including those that could metabolize cadaverine, putrescine, spermidine and spermine, those that could catabolize tryptamine, phenylethylamine and tyramine, but not those that could metabolize histamine. Apart from the correlation between biogenic amines and microbial flora, this study also clarified the relationship of biogenic amine metabolism with enzymes and microorganisms.
Recombinant Expression, Renaturation and Biological Activity of Temperature-Responsive Nanobodies against Aflatoxin B1
ZHANG Leping, TU Zhui, LI Yanping, LI Xiaojiang, SHUAI Wenyuan, ZHANG Hang, HE Qinghua
2023, 44(18):  141-148.  doi:10.7506/spkx1002-6630-20221028-289
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In this study, temperature-responsive anti-aflatoxin B1 (AFB1) nanoantibodies were obtained by fusion expression of elastin-like polypeptides (ELPs) of different lengths with nanoantibodies. The recombinant expression vectors pET30a-G8-ELP20, pET30a-G8-ELP40, pET30a-G8-ELP60 and pET30a-G8-ELP80 were synthesized by recursive directional ligation (RDL) and transformed into Escherichia coli BL21 (DE3) cells, separately. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed that the four fusion proteins were expressed in the form of inclusion bodies in the bacterial precipitate. The inclusion bodies were dissolved in denaturing buffer and refolded by dilution, inverse transition cycling (ITC) purification, dialysis or column refolding. SDS-PAGE analysis showed that the purity of the four proteins obtained achieved using the four refolding methods was similar. After refolding by ITC, the highest yield of 83%, 90.7%, 89.5%, and 88.3% was obtained for the four fusion proteins, respectively. The half maximal inhibitory concentration (IC50) of G8-ELP80 obtained by dilution refolding was the lowest (4.35 ng/mL), as determined by indirect competitive enzyme-linked immunosorbent assay (ELISA). The phase transition temperatures (Tt) of nanoantibodies tagged with the different ELPs, as determined by the turbidity method, were 45, 38, 32 and 28 ℃, respectively. Circular dichroism (CD) spectroscopy showed that the main secondary structures of the four fusion proteins were β-sheet and β-turn. This study provides a basis for subsequent AFB1 detection and analysis.
Using Illumina MiSeq High-throughput Sequencing to Explore the Succession of Microbial Community in Takifugu flavidus during Cold Storage
ZENG Lu, LIU Shuji, CHEN Xiaoting, LIN Hetong, LIU Zhiyu
2023, 44(18):  149-156.  doi:10.7506/spkx1002-6630-20221108-082
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In order to explore the microfloral succession in Takifugu flavidus during storage at 0 ℃, the spoilage bacteria were detected by 16S rDNA sequencing, the dominant spoilage bacteria in the late stage of storage were identified, and the changes in the microbial flora and the spoilage capacity of the dominant spoilage bacteria were analyzed. The results showed that the refrigerated shelf life of T. flavidus was about 5 days. In the early stage of storage (0–3 days), T. flavidus had the highest number of species of bacteria, including Pseudomonas, Streptophyta, and Shewanella, while in the late stage (6–15 days), the bacterial species were concentrated mainly in Pseudomonas and Shewanella. Ten dominant spoilage bacteria were isolated and identified, Shewanella and Pseudomonas accounting for a high proportion. Moreover, the spoilage capacity of several common dominant spoilage bacteria was analyzed, revealing that the decreasing order of the spoilage capacity was Pseudomonas fluorescent > P. azotoformans > Shewanella hafniensis > S. baltica. The results of this study provide a reference for understanding the spoilage mechanism of T. flavidus during cold storage and extending its shelf life.
Effect of Soymilk Fermented by Lactobacillus amylolyticus L6 on Intestinal Microorganisms and Short-chain Fatty Acids in Vitro
FEI Yongtao, HUANG Yihe, HUANG Li, LIU Gongliang, BAI Weidong
2023, 44(18):  157-164.  doi:10.7506/spkx1002-6630-20220729-333
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This study was conducted to investigate the effect of soymilk fermented by Lactobacillus amylolyticus L6 on intestinal microorganisms and short-chain fatty acids in vitro. After simulated gastrointestinal digestion of the fermented soymilk, the survival rate of L. amylolyticus L6 was determined. Then, the digested product was inoculated with human fecal microorganisms and cultured at 37 ℃ under anaerobic conditions for 48 h, and the changes of short chain fatty acids were determined during the fermentation process. The relative abundance of intestinal flora was analyzed by 16S rRNA sequencing. The results showed that L. amylolyticus L6 in the fermented soymilk maintained a high survival rate after simulated gastrointestinal transport. After 48 h in vitro fermentation with the fermented soymilk, the relative abundance of many harmful intestinal bacteria decreased, and the content of short-chain fatty acids (SCFAs) increased significantly (P < 0.05). These findings suggest that L. amylolyticus L6 fermented soymilk can increase the content of SCFAs after 48 h in vitro fermentation. This study provides a theoretical guidance for the development of fermented soybean products.
Effect of High-density Culture on the Fate of Porcine Muscle Stem Cells
LIN Ling, ZHU Haozhe, JIANG Yichen, ZHENG Yanyan, LIU Zheng, WU Zhongyuan, DING Shijie, ZHOU Guanghong
2023, 44(18):  165-174.  doi:10.7506/spkx1002-6630-20220928-315
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In this study, four different initial densities (2.7 × 103 as a control, 5.4 × 103, 1.1 × 104, and 2.2 × 104 cells/cm2) of porcine muscle stem cells were cultured for three days to explore the effect of high-density culture conditions on the fate of porcine muscle stem cells. The counting results showed that the suitable culture density of porcine muscle stem cells was 2 × 104–3 × 104 cells/cm2, and the proliferation-related protein phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2) was down-regulated when the initial cell density was outside this range, causing a proliferation inhibitory effect. The transcriptome sequencing results showed that 2 125 differentially expressed genes were identified between the highest density and control groups, mainly enriched in some signaling pathways such as those related to cell senescence and cell cycle and the PI3K-Akt signaling pathway. Furthermore, the expression of the stemness genes PAX7 and PAX3 and the quiescence-related gene SPRY1 was upregulated under high-density conditions. The expression of the differentiation genes MYOD, MYOG and MYH3 and the senescence genes P21, P53 and P15 was upregulated. The results of real-time PCR and Western blot assay showed that the expression of PAX7, SPRY1, MYOG, and P21 were upregulated as the cell density increased. The above results showed that porcine muscle stem cells had multiple fats such as returning to a quiescent state, differentiation and senescence under high-density conditions. The results of this study can help in understanding the mechanism of cell fate determination and provide a theoretical basis for studying the regulation of cell proliferation and differentiation under high-density condition.
Isolation and Identification of Lactic Acid Bacteria and Microbial Diversity in Fresh Cow Milk from Xilin Gol
SUN Jie, SU Qian, DU Ping, XIA Yanan, ZHAO Jie, YU Jie, CHEN Yongfu
2023, 44(18):  175-182.  doi:10.7506/spkx1002-6630-20221030-303
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This work was undertaken in order to study the microbial diversity in dairy products from ethnic minority habitats in Xilin Gol. Six fresh cow milk samples from Sunit Left Banner and East Ujimqin Banner (n = 3 for each region) were selected for metagenomic sequencing, and lactic acid bacteria (LAB) from these samples were isolated and identified by the traditional culture method. There results showed that a total of 165 microbial species were identified in the six milk samples, the dominant ones being Lactococcus lactis and Leuconostoc mesenteroides. Many microbial species were common to the two groups, and there was no significant difference in α- or β-diversity, but there were still significantly differential species identified between these groups. Fresh milk from Sunit Left Banner was dominated by Lactococcu phage BM13, while that from East Ujimqin Banner was dominated by Enterobacter sp. MGH8. A total of 1 408 048 genes and 412 metabolic pathways were annotated using the Uniprot database, whose functions were concentrated on amino acid metabolism and glycolic acid metabolism. A total of 47 strains were isolated from the six fresh milk samples, and L. lactis was the dominant strain, which was consistent with the metagenomic results. This study is expected to provide a theoretical basis for the safe production of fermented dairy products and the development of starter cultures.
Effect of Optimized Thioglycolate Medium on the Growth and Metabolites of Akkermansia muciniphila
WU Xiaoling, XU Jinzhao, XU Yuanzhi, NING Ke, XIE Qinggang, XU Xiaoxi
2023, 44(18):  183-192.  doi:10.7506/spkx1002-6630-20221109-098
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In this study, Akkermansia muciniphila (Akk) was cultured to a concentration of more than 109 CFU/mL in optimized liquid thioglycolate medium. Besides, the differential metabolites in the fermentation supernatants from the original and optimized medium were analyzed by non-targeted metabolomics, and the physiological functions of the characteristic products were evaluated. The results showed that when cultured in the optimized medium, Akk produced higher amounts of various functional metabolites such as propionic acid, bardoxifene, docetaxel and vinblastine by affecting protein digestion and absorption, aminoacyl tRNA biosynthesis, bile secretion, ATP-binding cassette (ABC) transporter, and the metabolic pathways of amino acid biosynthesis. These metabolites could be directly related to anti-cancer, anti-inflammatory, anti-obese and other functions. This study provides solid evidence that Akk can function as a probiotic agent and postbiotic to regulate many diseases, and lays a theoretical foundation for future industrial production.
Screening and Compositional Analysis of Lactic Acid Bacteria with Tyrosinase Inhibitory Activity
ZHANG Xue, LI Xuefeng, HU Ruifeng, GUO Wanchun, LIU Xuejun
2023, 44(18):  193-199.  doi:10.7506/spkx1002-6630-20221224-233
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In this study, the fermentation supernatant, cells and disrupted cell supernatant of five strains of lactic acid bacteria (LAB) were screened for tyrosinase inhibitory activity and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging capacity. The results showed that the tyrosinase inhibitory activity of the fermentation supernatant was much higher than that of the cells and the disrupted cell supernatant. The percentage inhibition of tyrosinase activity by the fermentation supernatant of strain MNJ-9 was 68.9% compared to only 50.9% with the cells and 34.8% with the disrupted cell supernatant. The fermentation supernatant of strain MNJ-9 scavenged 93.1% of DPPH radical. Meanwhile, MNJ-9 was identified by 16S rDNA gene sequencing. The major components of the fermentation supernatant of MNJ-9 were investigated using untargeted metabolomics based on liquid chromatography-mass spectrometry (LC-MS). The tyrosinase inhibitory activity and DPPH radical scavenging capacity of 10 of the major components were measured. The results showed that 5-aminovaleric acid had the highest tyrosinase inhibitory activity and DPPH radical scavenging capacity. The cytotoxicity of 5-aminovaleric acid on mouse melanoma cells (B16F10), and its impact on the melanin content and tyrosinase activity of B16F10 cells were determined to evaluate its whitening ability. The results showed that the cell survival rate was equal to or over 80% after culture for 72 h with 70 μg/mL of 5-aminovaleric acid, indicating no apparent cytotoxicity. At 5-aminovaleric acid concentrations of 10–70 μg/mL, the intracellular melanin content and tyrosinase activity were 85.4%–45.2% and 76.53%–40.5%, respectively. This indicates that 5-aminovaleric acid has some whitening ability.
Untargeted Metabolomics Based on Ultra-high Performance Liquid Chromatography-Mass Spectrometry for Analysis of Differential Metabolites in Lactobacillus-fermented Ultra-fine Whole Pulp of Chimonobambusa quadrangularis Shoot
Wang Lei, Jia Yulong, Luo Yanyu, ZOU Chunxia, ZHANG Ying
2023, 44(18):  200-213.  doi:10.7506/spkx1002-6630-20221019-177
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In this work, a non-targeted metabolomic study was performed on the ultrafine whole pulp of Lactobacillus-fermented Chimonobambusa quadrangularis shoot using ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS). Based on the variable importance in projection (VIP) > 1 and P < 0.05 in t-test, 361, 392 and 398 differential metabolites were selected in raw (control) versus Limosilactobacillus reuteri (LR) fermented samples, control versus Lactobacillus plantarum (LP) fermented samples, and control and LR + LP (LRP) fermented samples in the positive ion mode; and 268, 304 and 321 differential metabolites in the negative ion mode, respectively. The major differential metabolites were lipids and lipid-like compounds, organic acids and their derivatives, organic heterocyclic compounds, and benzenes including histamine, D-mannan, indole-3-lactic acid, DL-4-hydroxyphenyl lactate, ornithine, and proline. Four significant metabolic pathways were identified by Kyoto Encyclopedia of Genes and Genomes metabolic pathway enrichment analysis, namely purine metabolism, pyrimidine metabolism and phenylalanine metabolism and citric acid cycle. This study provides theoretical support for the development of fermented bamboo shoot products.
Isolation, Identification and Biological Characteristics Analysis of a New Proteus Phage
SU Yahang, HOU Zhongyu, KOU Xiaodi, LIU Shuang, ZHU Chenglin, CHEN Juan, DAO Xiaofang, TANG Junni
2023, 44(18):  214-222.  doi:10.7506/spkx1002-6630-20221003-018
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Objective: To isolate, identify and characterize a new Proteus phage and analyze its genome. Methods: The strains isolated from yak slaughtering in the laboratory were cultured, their DNA was extracted and subjected to 16S rRNA sequencing, and the host bacteria were identified and used for bacteriophage separation from the environmental sewage of a yak slaughterhouse in Chengdu, Sichuan. The bacteriophage was separated and purified by the double-layer agar plate method, and its optimal multiplicity of infection (MOI), one-step growth curve and tolerance were determined. Its morphology was observed by negative staining with phosphoric acid and transmission electron microscopy (TEM). The whole genomic DNA of the phage was extracted using a commercial kit, sequenced on the Illumina MiSeq sequencing platform, assembled, annotated, and genomically analyzed. Results: Totally 10 Proteus strains were identified from the 67 isolates, and one phage with strong lytic activity, named Proteus phage PV66, was isolated from the sewage. PV66 could lyse Proteus and Enterococcus faecalis. The optimal MOI was 0.1. The results of one-step growth curve showed that the incubation period of bacteria infected with PV66 was about 10 min, the lytic period was about 80 min, and the average lytic amount was 321 PFU/cell. The tolerance pH range was 3–12, and the tolerance temperature was 65 ℃. The bacteriophage had a C3 morphotype under electron microscopy, with a head length of (140 ± 1) nm, a width of (54 ± 2) nm, and a tail length of (36 ± 1) nm. The genome sequencing results showed that the whole sequence length of PV66 was 90 492 bp. The phylogenetic tree analysis showed that the phage had high homology with Proteus mirabilis phage and Cronobacter phage and was in the same branch as non-Kuravirus. Conclusion: The Proteus phage has good application potential. This study provides a basis for future development of new bacteriophages, and enriches the database of Proteus phages.
Differences in Bacterial Community Structure between Bottom and Wall Pit Mud for Nongxiangxing Baijiu
ZENG Bo, ZOU Yongfang, RAO Jiaquan, WANG Xiaoping, WEN Jing, HUANG Zhiguo
2023, 44(18):  223-230.  doi:10.7506/spkx1002-6630-20221013-125
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This study was performed in order to understand the differences in bacterial communities and physicochemical indicators between bottom and wall pit mud for nongxiangxing baijiu. Five physicochemical indicators of pit mud were analyzed. High-throughput sequencing technology was used to investigate the bacterial communities, and correlation analysis was carried out between bacterial communities and physicochemical indicators. The results showed that the pH and catalase activity in bottom pit mud were significantly higher than those in wall pit mud (P < 0.05), while the opposite was true for ammonium nitrogen content (P < 0.05). The dominant bacterial phyla in pit mud were Firmicutes, Bacteroidetes, and Synergistetes. The dominant genera in bottom pit mud were Hydrogenispora, Caproiciproduces, Aminobacterium, and Proteiniphilum, the relative abundance of Hydrogenispora being significantly higher than that in wall pit mud (P < 0.05). The dominant genera in wall pit mud were Capriocipiroducens, Lactobacillus, Syntrophaceticus, and Aminobacterium, the relative abundance of Capriocipiroducens, Lactobacillus and Syntrophaceticus being significantly higher than that in bottom pit mud (P < 0.05). The physicochemical indicators and bacterial communities in bottom and wall pit mud were different. Among the physicochemical indicators, pH had the greatest impact on the bacterial community in bottom pit mud, and Hydrogenispora was the iconic genus of bacteria in pit bottom mud. The relative abundance of enzymes involved in acid metabolism was higher in pit wall mud. In conclusion, there were spatial differences in the bacterial communities between bottom and wall pit mud, which could have an important impact on the fermentation of baijiu.
Correlation between Taste Quality and Bacterial Community of Dazhu Rice Wine
XIANG Fanshu, CAI Wenchao, GUO Zhuang, LIU Huijie, YU Haiyan, SHAN Chunhui
2023, 44(18):  231-238.  doi:10.7506/spkx1002-6630-20220829-347
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This study was undertaken to reveal the correlation between the taste quality and bacterial community of Dazhu rice wine. Taste attributes were measured using an electronic tongue, the types and contents of organic acids and amino acids were analyzed, and lactic acid bacteria were isolated and identified from the rice wine. Moreover, analysis and functional prediction of its bacterial community were carried out using MiSeq high-throughput sequencing technology. Finally, the correlation between various taste attributes and bacterial community was calculated. The results showed that Dazhu rice wine varied widely in terms of saltiness, sourness and bitterness. The rice wine contained seven organic acids and 15 amino acids, the predominant organic acids being lactic acid (8.82 mg/g) and acetic acid (6.27 mg/g), and the predominant amino acids being glutamic acid (8.06 mg/g), leucine (4.10 mg/g) and aspartic acid (3.94 mg/g). A total of 14 lactic acid bacteria were isolated by pure culture and identified as five species, namely Limosilactobacillus fermentum (four strains), Pediococcus pentosaceus (four strains), Lacticaseibacillus paracasei (two strains), Lactiplantibacillus plantarum (two strains) and Leuconostoc pseudomesenteroides (two strains). High-throughput sequencing results showed that the average relative contents of Firmicutes and Proteobacteria were greater than 1.0%, with a cumulative content of 96.93%; the relative contents of Lactobacillus, Pediococcus, Weissella, Lactococcus, Leuconostoc and Acinetobacter were greater than 1.0%, with a cumulative content of 89.43%. Correlation analysis showed a significant negative correlation (P < 0.05) between Weissella and taste indicators such as aftertaste A, astringency and bitterness. These findings showed that the major organic acids in Dazhu rice wine were lactic acid and acetic acid, that the major amino acids were glutamic acid, leucine and aspartic acid, and that bacterial community was dominated by lactic acid bacteria such as Lactobacillus, Pediococcus and Weissella. Weissella could have a positive significance on the formation of rice wine quality.
Component Analysis
Change of Aroma Components in Instant Baiyaqilan Tea Powder during Processing
LIN Shidi, JIANG Qingxiang, LIN Qi, XU Yongquan, LI Lijun,, WENG Shuyi, NI Hui, LI Qingbiao
2023, 44(18):  239-248.  doi:10.7506/spkx1002-6630-20221011-105
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In this study, the change of aroma components in instant Baiyaqilan tea powder during processing was investigated by sensory evaluation, gas chromatography-mass spectrometry (GC-MS) and odor activity value (OAV). The results showed that the over aroma profile and the types of aroma components in instant Baiyaqilan tea powder were similar at different stages of processing, while the aroma intensity and volatile components were significantly different. 3-Ethyl-2,5-dimethylpyrazine, hotrienol, methyl salicylate, safranal, geraniol, indole, β-damascenone, cis-jasmonone, geranyl acetone, trans-nerolidol and methyl jasmonate were found to make key contributions to aroma changes. The extraction process significantly increased the OAV of safranal, geraniol, indole, β-damascenone and cis-jasmonone. Disc separation, reverse osmosis and freeze drying significantly reduced the OAV of geraniol, indole, β-damascenone, cis-jasmonone, methyl jasmonate, trans-nerolidol, 3-ethyl-2,5-dimethylpyrazine and safranal. Moreover, ultrafiltration treatment removed tea polyphenol complexes, promoting the release of aroma volatile compounds combined with polyphenols and thus increasing the OAV of geraniol, indole, β-damascenone, cis-jasmonone and methyl jasmonate. This study clarified the aroma change during the processing of instant Baiyaqilan tea powder and showed that ultrafiltration treatment promoted the release of aromatic volatiles. This study can provide a theoretical basis for improving the processing of instant Baiyaqilan tea powder for improved aroma quality and the processing technology of instant tea.
Characterization of Volatile Aging Markers in Traditional Semi-Dry Huangjiu by Comprehensive Two-Dimensional Gas Chromatography-Time of Flight Mass Spectrometry and Chemometrics
LIU Shaopu, ZHOU Zhilei, YU Hongbo, MAO Laigen, PAN Xingxiang, JI Zhongwei, MAO Jian
2023, 44(18):  249-258.  doi:10.7506/spkx1002-6630-20221128-318
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In this investigation, headspace solid-phase microextraction (HS-SPME) coupled with comprehensive two-dimensional gas chromatography-time of flight mass spectrometry (GC × GC-TOFMS) was used to analyze the volatile compounds of traditional semi-dry Huangjiu (Chinese yellow rice wine) of different ages. Furthermore, chemometrics was used to identify the aging markers. The results showed that a total of 351 compounds were identified in Huangjiu of six ages, of which 40 compounds were found in Huangjiu for the first time. The total number of volatile compounds showed an increasing trend with aging time. One-way analysis of variance (ANOVA) and correlation analysis found a strong correlation between the contents of 107 compounds that significantly changed and aging time, among which 55 compounds such as 2-acetylthiophene were found to be significantly correlated with aging time for the first time. Principal component analysis (PCA) and cluster analysis revealed that the aging process of traditional semi-dry Huangjiu could be divided into three stages: short-term aging (1–3 years), medium-term aging (6–12 years) and long-term aging (more than 15 years). By partial least squares-discriminant analysis (PLS-DA), 37 aging markers were selected, the contents of 12 compounds of which decreased significantly while the other 25 increased significantly during the aging process. Ten most representative key aging markers were finally identified based on Pearson correlation coefficients, among which 5 compounds including isoamyl butanoate, butanoic acid, 2-furanmethanol, 2-acetyl-5-methylfuran and o-cresol were defined as key aging markers of Huangjiu for the first time. This study provides a scientific basis for understanding the aging process, evaluating the quality and identifying the age of Huangjiu.
Differences in Key Flavor Substances and Quality between Charcoal-Baked and Electrically Baked White Tea
WU Quanjin, ZHOU Zhe, QI Siyu, WU Ying, SUN Weijiang
2023, 44(18):  259-267.  doi:10.7506/spkx1002-6630-20221109-094
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In this study, the differences in the aroma substances, major flavor substances and sensory quality of different grades of charcoal-baked and electrically baked white tea were studied by gas chromatography-mass spectrometry (GC-MS) and high performance liquid chromatography (HPLC) combined with chemometrics methods. The results showed that a total of 262 volatile compounds were detected, with terpenes, heterocyclic compounds, alcohols, esters and ketones being the major components, and 2-phenyl ethanol, linalool, geraniol, methyl salicylate, laurene, and (E)-linalool oxide accounting for a higher proportion of the total amount of volatile compounds. Compared with electrically baked white tea, 77, 16 and 93 differential volatile substances were identified from charcoal-baked Yinzhen, Baimudan and Shoumei tea, respectively. Odor activity value (OAV) analysis showed that linallinol, geraniol, 2-phenyl ethanol and laurene were important aroma substances of charcoal-baked white tea, which contribute to natural refreshing, floral and fruity or sweet aromas. Analysis of non-volatile compounds showed that there were significant differences in the content of total and individual catechins, tea polyphenols, caffeine and water extract between charcoal-baked and electrically baked white tea. The results of sensory evaluation were consistent with those of volatile and non-volatile substances. This study is helpful to understand the aroma and flavor quality of charcoal-baked white tea and provides theoretical support for improving the quality of white tea.
Effect of the Addition of Scallion, Ginger and Perilla on the Changes in Volatile Compounds and ATP and Related Compounds in Pre-prepared Ovalipes punctatus
WANG Qingzheng, XU Yanru, LIU Min, GU Guizhang, XU Dalun, ZHANG Jinjie
2023, 44(18):  268-276.  doi:10.7506/spkx1002-6630-20221031-311
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The effects of scallion, ginger and perilla as garnishes on the eating quality of pre-prepared Ovalipes punctatus were investigated. The results of sensory evaluation showed that all three garnishes had a deodorizing effect, with scallion being most effective. Ginger could impart a strong pungent flavor to crab meat, and perilla could endow crab meat with a rich and strong odor. There were significant differences in the volatile odor components of crab meat without (control) and with added garnish, the latter containing a richer variety of volatile compounds. A total of 47 volatile compounds were identified in the control group, and 50 in crab meat with added scallion. Scallion addition decreased the relative contents of fishy odorants such as hexanal, heptaldehyde, octanal, nonanal, and 1-octen-3-ol. Fifty-six volatile compounds were identified in crab meat with added ginger, including 14 aldehydes such as decanal and 3-methylbutyraldehyde, which could impart good fruity, nutty and floral aromas to crab meat. Totally 70 volatile compounds were detected in crab meat with added perilla, including 15 key volatile compounds such as ketones, esters and pyrazines in rich amounts, making the flavor more unique. Adding garnishes did not change the degradation pathway of adenosine triphosphate (ATP) in crab meat at high temperature, but affected the degradation of related compounds. The content of adenine (Ad) in the ginger addition group was highest, 1.95 mg/100 g. The content of IMP in the perilla addition group was highest, 72.52 mg/100 g, contributing to the distinctive umami taste. In the scallion addition group, which had the weakest bitter taste, the contents of hypoxanthine riboside (HxR) and hypoxanthine (Hx) were lowest, 2.12 and 3.69 mg/100 g, respectively. The findings of this study may provide a technical and theoretical basis for the development of high-quality pre-prepared Ovalipes punctatus products.
Gas Chromatography and Electronic Nose Combined with Chemometrics to Investigate Differences in Key Aroma Components in Eight White-Fleshed Peach Cultivars
LIU Gege, BI Jinfeng, GOU Min, LÜ Jian, WU Xinye, CHEN Qinqin
2023, 44(18):  277-285.  doi:10.7506/spkx1002-6630-20221117-202
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In order to explore the differences in the aroma quality of different white-fleshed peach cultivars, an electronic nose and headspace solid-phase microextraction combined with gas chromatography-mass spectrometry (HS-SPME-GC-MS) were used to analyze the volatile components of eight cultivars of white-fleshed peach including Baiyintao (BYT), Longmi 5 (LM5H), Xinchuanzhongdao (XCZD), Qiutong (QT), Xindajiubao (XDJB), Yihongmi (YHM), Yihongshuimi (YHSM) and Okayama (GSB). Relative odor activity value (ROAV) and variable importance in projection (VIP) were used to analyze the contribution of each volatile component to the overall aroma of white-fleshed peaches. In the electronic nose radar fingerprint, the W1W, W5S, W1S and W2W sensors showed different responses to the eight cultivars. Principal component analysis (PCA) demonstrated differences in the overall aroma of white-fleshed peach cultivars. In total, 49, 46, 46, 47, 49, 47, 47, 53 volatile compounds were identified in BYT, LM5H, XCZD, QT, XDJB, YHM, YHSM, and GSB by HS-SPME-GC-MS, respectively, the major ones being aldehydes, alcohols, lactones and esters. 2-Hexenal, hexanal, trans-2-hexenal and trans-2-hexen-1-ol, contributing to grass aroma; linalool, contributing to floral aroma; and n-hexanol, responsible for fruity aroma, were identified as key aroma compounds due to their ROAV ≥ 1 and VIP > 1. Variations in their concentrations caused differences in the key aroma of the eight white-fleshed peach cultivars.
Metabolomics Analysis of the Effect of Reduced Glutathione on Metabolites of Chardonnay Wine
LIANG Hongmin, GAO Deyan, QUAN Qiaoling, WANG Cong, GUO Yayun, SHI Hongmei
2023, 44(18):  286-293.  doi:10.7506/spkx1002-6630-20221017-157
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We undertook this study in order to explore the effect of reduced glutathione (GSH) on the physicochemical indexes and non-volatile and volatile metabolites of Chardonnay wine during a five-month storage period after the end of alcoholic fermentation. Five different concentrations of GSH were added into the wine after the end of alcoholic fermentation. The changes of physicochemical indexes, total phenolic content and color during the storage period were measured. Widely targeted metabolomic analysis and volatile metabolomic analysis based on ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and gas chromatography-mass spectrometry (GC-MS) were performed on wine samples with and without the addition of 20 mg/L GSH. The results showed that during the five-month storage period, the addition of different concentrations of GSH had no significant effect on the volatile acid content, total acid content and pH of white wine (P > 0.05) but significantly influenced its total phenol content, total flavonoid content, color and free SO2 content. The total phenol content of the wine sample added with 30 mg/L GSH was highest after storage for five months, and the color index of the wine sample added with 20 mg/L GSH was lowest. The addition of GSH could delay the decrease of free SO2 content. A total of 693 non-volatile metabolites and 816 volatile metabolites were identified. Based on variable importance in projection (VIP) greater than 1.0 and fold change (FC) equal to or greater than 1.4, a total of 33 non-volatile differential metabolites and 22 volatile differential metabolites were selected. Among the 32 non-volatile metabolites, 17 were down-regulated and the rest 16 were up-regulated. Among the 22 volatile metabolites, three were down-regulated and the other 19 were up-regulated. Some phenolic acids, fatty acids and amino acids were associated with changes in volatile metabolites.
Effect of Scenting on Aroma of Different Types of Dark Tea
LIU Jiashun, AN Huimin, CHEN Yuan, LI Shi, HUANG Yiwen, CHEN Jinheng, ZHANG Zhanghan, HUANG Jian’an, LIU Zhonghua
2023, 44(18):  294-304.  doi:10.7506/spkx1002-6630-20221008-058
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In this study, solid-phase microextraction (SPME) and comprehensive two-dimensional gas chromatography-quadrupole time-of-flight mass spectrometry (GC × GC-Q-TOF-MS) were used for the qualitative and quantitative determination of volatile components in dark tea samples, and the characteristic volatile components of jasmine dark tea were identified by multivariate statistical analysis and their regulatory effects on tea aroma were investigated. The results showed that the scenting process could coordinate the aroma characteristics of dark tea (such as aged, woody and smoky pine aroma), and endow dark tea with the aroma of jasmine, contributing to the aroma quality of jasmine dark tea that is defined as “fresh flowery fragrance, and pure tea-like fragrance”. A total of 366 volatile components were identified in dark tea and jasmine dark tea, of which 38 volatile components such as linalool, α-farnesene, and benzyl acetate were characteristic volatile components to distinguish the aroma quality of dark tea before and after scenting, and most characteristic volatile components were positively correlated with jasmine and fungus fragrance, and negatively correlated with aged woody and smoky pine aroma. These findings led us to conclude that scenting is important for improving the quality of jasmine dark tea. This study provides a theoretical basis for improving the processing technology and quality and regulating the aroma of jasmine dark tea.
Determination of 46 Ginsenosides in Different Processed Ginseng Products by Dispersive Solid Phase Extraction Combined with Ultra-high Performance Liquid Chromatography-Tandem Mass Spectrometry
WANG Yanhong, WU Yutong, CAO Hongfang, XU Xuanwei, ZHAO Dan, LI Yueru
2023, 44(18):  305-315.  doi:10.7506/spkx1002-6630-20221016-147
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A method was developed for the simultaneous detection of 46 ginsenosides in different processed ginseng products by dispersive solid phase extraction combined with ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The samples were ultrasonically extracted with 80% methanol aqueous solution, and the extract was purified by dispersive solid phase extraction (DSPE) using primary secondary amine (PSA) as a sorbent, separated on a KINETEX XB-C18 column (2.1 mm × 100 mm, 2.6 µm) by gradient elution using a mobile phase consisting of acetonitrile and 0.1% formic acid aqueous solution, detected in the negative ion mode with multiple reaction monitoring (MRM), and quantified by an external standard method. The 46 ginsenosides were detected within 35 minutes. Under the optimized conditions, the calibration curves showed good linearities for 46 compounds at concentrations ranging from 0.02 to 50 µg/mL with correlation coefficients (R2) over 0.99. The limits of detection (LODs) and the limits of quantitation (LOQs) were in the range of 0.4–3.2 mg/kg and 1.4–10.5 mg/kg, respectively. The average recoveries at three spiked levels were in the range of 80.5%–111.6% with relative standard deviations (RSDs) of 1.3%–6.8%.  In addition, the RSDs for intra- and inter-day precision were 0.5%–4.5% and 1.1%–6.6% (n = 6), respectively. The RSDs for repeatability ranged between 1.5% and 7.3% (n = 6), and the RSDs for sample stability within 24 hours between 0.9% and 5.8% (n = 6) within 24 h. In this method, the type and dosage of DSPE sorbent were optimized, and the influence of DSPE purification on the matrix effect was investigated. The results showed the matrix interference was effectively reduced by DSPE with PSA, and the sample pretreatment was easy to operate, efficient and cost-saving. This method was successfully applied to the analysis of 46 ginsenosides in sun-dried ginseng, red ginseng and black ginseng. The results revealed that ginsenosides were transformed into rare ginsenosides in red ginseng and black ginseng due to steaming, and the contents of rare ginsenosides in black ginseng were much higher than those in red ginseng. The proposed method was accurate and reliable, fast, easy to operate, immune to impurity interference, and suitable for the simultaneous quantitative analysis of 46 ginsenosides in processed ginseng products.
Safety Detection
Amine-Responsive Anthocyanin-Loaded Hydrogel for Monitoring the Freshness of Fish and Shrimp
DONG Shengye, ZHANG Yaqin, FU Zhaodi, SHI Xingbo
2023, 44(18):  316-323.  doi:10.7506/spkx1002-6630-20221011-093
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A visual, non-destructive method for monitoring fish and shrimp freshness was established using an amine-responsive agarose hydrogel loaded with purple sweet potato anthocyanin (PSPA). Trimethylamine (TMA) was selected as an indicator of fish and shrimp spoilage. The redshift of the absorption peak of PSPA hydrogels after reaction with different concentrations of TMA (0–194.22 × 10-9 mol/L) was investigated. The red (R), green (G), and blue (B) channel values of the hydrogels were analyzed using smartphone imaging combined with the Image J software. The concentration of TMA was quantified using G/R as the output signal. The limit of detection (LOD) was 0.84 × 10-9 mol/L and the linear range was 8.78 × 10-9–19.386 × 10-9 mol/L (R2 = 0.992 1). When used to detect the freshness of fish and shrimp, the proposed method showed a good correlation with the national standard method for measuring total volatile basic nitrogen (TVB-N) content with a Pearson correlation coefficient of 0.998 4 and 0.996 6, respectively. Due to the application of functional hydrogel and the introduction of smartphones, the method had the advantages of simplicity, non-destructiveness, and portability. This study provides a new method for on-site monitoring the real-time freshness of aquatic products, which has a broad application prospect in the field of aquatic food safety detection.
Gold Nanoparticle-based Colorimetric Aptasensor for Detection of Malathion
TIAN Hengqi, WANG Yao, YANG Haitao, LI Zhaozhou, WU Jiabei, WANG Lin, LIU Haoyu, CHANG Yunhe, CHEN Xiujin
2023, 44(18):  324-330.  doi:10.7506/spkx1002-6630-20220915-142
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In this study, a novel colorimetric sensing method for the detection of malathion was established based on the principle that gold nanoparticles (AuNPs) can aggregate in high-salt solutions, leading to color changes in the solutions. The optimal concentration of NaCl was 500 mmol/L and the optimal concentration of aptamer was 1 μmol/L in the reaction system. This method showed a good linear relationship in the range of malathion concentration of 50–1 500 ng/mL, with a detection limit of 45.54 ng/mL. The recoveries of malathion in spiked lettuce and apple samples were 98.24%–100.91% and 99.48%–101.29%, respectively. The AuNPs-based colorimetric aptasensor could be used for the sensitive, specific, accurate and rapid detection of malathion. Furthermore, this study may provide a reference for developing colorimetric aptasensors for the detection of other pesticide residues.
Application of Surface Enhanced Raman Scattering Spectroscopy in the Rapid Detection of Ethylenediaminetetraacetic Acid Disodium Salt in Prepackaged Dried Rice Noodles
KONG Hongxing, CHEN Qiying, MENG Haisen, CHEN Ruijue, FENG Jun, CHENG Hao
2023, 44(18):  331-338.  doi:10.7506/spkx1002-6630-20221109-095
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Fe3O4@SiO2@silver nanoparticles (AgNPs) were synthesized by a facile seed-mediated method and used as a surface enhanced Raman scattering (SERS) substrate to detect the content of the trace additive ethylenediaminetetraacetic acid disodium salt (EDTA-2Na) in prepackaged dried rice noodles. Fe3O4@SiO2@AgNPs were characterized by transmission electron microscopy, X-ray energy spectroscopy, X-ray diffraction spectroscopy, infrared spectroscopy, and SERS spectroscopy. The results showed that Fe3O4@SiO2@AgNPs were a magnetic substrate with a core-shell structure, and a diameter of about 660 nm. The minimum detectable concentration of rhodamine 6G (R6G) solution was measured to be 1 × 10-9 mol/L and the enhancement factor was 4.27 × 108. The limit of detection (LOD) of EDTA-2Na solution using Fe3O4@SiO2@AgNPs was 1.75 × 10-6 mg/mL. Good linearity was observed between the SERS intensity of the characteristic peak at 1 625 cm-1 and the logarithmatic concentration of EDTA-2Na with a correlation coefficient (R2) of 0.994. The LOD was 0.1 mg/kg for EDTA-2Na in prepackaged dried rice noodles, and the recoveries for spiked samples were 96.50%–104.67% with relative standard deviation (RSD) of 1.2%–4.2%. Accordingly, this method can accurately, sensitively and rapidly detect whether EDTA-2Na is added in excess in prepackaged dried rice noodles.
Analysis of 500 Pesticide Residues in Milk and Infant and Young Children Formula Milk Powder by Ultra-high Performance Liquid Chromatography-Quadrupole-Time of Flight Mass Spectrometry
NING Xiao, ZHANG Jingran, JIN Shaoming, CAO Jin
2023, 44(18):  339-346.  doi:10.7506/spkx1002-6630-20220819-225
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Objective: An ultra-high liquid chromatography-quadrupole-time of flight mass spectrometry (UPLC-QTOF-MS) was established and applied for the screening and quantitation of pesticide residues in raw milk and infant and young children formula milk powder. Methods: A stable and reliable pretreatment procedure involving quick, easy, cheap, effective, rugged and safe (QuEChERS) extraction and Captiva EMR-lipid cleanup was developed and validated. The samples were separated on a Luna Omega Polar C18 (2.1 mm × 100 mm, 1.6 μm) by gradient elution using mobile phase A consisting of 2 mmol/L ammonium formate and 0.01% formic acid and mobile phase B consisting of 2 mmol/L ammonium formate and 0.01% formic acid. By analyzing commercial pesticide certified reference materials, a single injection method using two fragmentation techniques, collision-induced dissociation (CID) and electron activated dissociation (EAD) was developed for the simultaneous qualitative screening and quantitative analysis of 500 pesticide compounds, in which mass spectrometry data were collected using the Zeno sequential window acquisition of all theoretical fragment ion spectra (SWATH) data-independent acquisition (DIA) mode. Comprehensive scores obtained by the OS software from four factors, including mass deviation, retention time deviation, differential isotope ratio and spectrum library similarity, were used for qualitative screening. The matrix-matched external standard method was used for quantitative analysis based on the peak area of precursor ions in the extracted ion chromatogram (XIC). Results: The comprehensive scores of the 500 pesticides and their metabolites ranged from 90.4 to 99.4 points, higher than the minimum score of qualitative screening (90 points), and the limits of detection (LOD) were in the range of 0.025–0.5 μg/kg. The correlation coefficients (r2) of the matrix-matched calibration curves for 103 pesticides and their metabolites with maximum residue limits (MRL) in milk set by the national standard of China GB 2763-2021 were above 0.99. The average recoveries ranged from 72.7% to 114.5% at three spiked concentration levels with relative standard deviations (RSDs) of 2.1%–13.6% (n = 6), and the limits of quantitation (LOQ) were in the range of 0.15–3 μg/kg. Conclusion: This method was sensitive and accurate, and could be used for the screening identification of multiple pesticides and their metabolites and the quantitative analysis of restricted pesticides with high detection rate in raw milk and infant formula milk powder at low cost.
Establishment of a Dual Enzymatic Recombinase Amplification Method for Rapid Detection of Foodborne Pathogens in Infant Formula Powder
LIN Zhiwei, WANG Shuai, WANG Yingchun, WU Zhanwen, LI Tao, LI Hongna, YANG Yange, YUAN Fei
2023, 44(18):  347-354.  doi:10.7506/spkx1002-6630-20221013-127
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For quick and multiple detection of the contamination of foodborne pathogens in infant milk powder, a dual enzymatic recombinase amplification (ERA) method for the rapid detection of Cronobacter, Salmonella, Listeria monocytogenes and Staphylococcus aureus were established in this study. Firstly, specific and efficient primers and probes were selected for these foodborne pathogens. Then, two groups of dual-ERA systems were established and optimized. The minimum detection limit was 10-2 ng/μL for Salmonella, L. monocytogenes and S. aureus, and 1 ng/μL for Cronobacter. The results of simulated contamination test showed that the four foodborne pathogens could be detected simultaneously after 6 hours of enrichment culture at an initial contamination level of 1 CFU/mL. When this method and real-time polymerase chain reaction (PCR) were used to detect commercial infant milk powder, consistent results were obtained, which confirmed the accuracy and reliability of the dual-ERA method. This method could simultaneously detect the four foodborne pathogens in about 20 minutes. Compared with the traditional method and real-time PCR, the detection efficiency was significantly improved, which is of great significance for promoting the rapid screening of foodborne pathogens.
Rapid Detection of Norovirus in Shellfish by Dual Fluorescence Reverse Transcription-Enzymatic Recombinase Amplification
WU Zhanwen, WANG Shuai, KANG Jie, LI Tao, LI Hongna, CAI Jie, ZHANG Hao, YANG Yange, YUAN Fei
2023, 44(18):  355-363.  doi:10.7506/spkx1002-6630-20221126-300
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Objective: To establish a dual fluorescence reverse transcription-enzymatic recombinase amplification (RT-ERA) method for the rapid detection of GI and GII noroviruses (NoV) in shellfish using MS2 as the model process control virus. Methods: The target sequences of GI and GII NoV were individually cloned into vectors with a T7 promoter. Then, high-purity RNA was obtained by in vitro transcription. Dual fluorescence RT-ERA was performed using the primers and probes of MS2 phage designed in this study and the primers and probes of GI and GII NoV selected previously. The reaction procedure and system were optimized. The sensitivity of the method was analyzed. Finally, this method was used to detect real samples, and its accuracy and viability were determined by comparing the results with those obtained by the reference method specified in the national standard GB 4789.42-2016. Results: The optimal volumes of primers and probes for the dual fluorescence RT-ERA assay were 4.1 and 1.8 μL, respectively. The method took only about 10 min to perform, and could detect as low as 102 copies of NoV nucleic acid. When the established method was applied to detect 29 real samples, the results were consistent with those from GB 4789.42-2016. Conclusion: The dual fluorescence RT-ERA assay can simultaneously detect MS2, GI and GII NoV with high sensitivity and accuracy, which will lay a foundation for rapid on-site detection of NoV in the future.
Determination of Dioxins and Dioxin-like Polychlorinated Biphenyls in Pork by Gas Chromatography-Tandem Mass Spectrometry
LU Jing, JIAO Yanna, CHENG Jing, SHEN Tiantian, ZHU Shaohua, FU Shanliang
2023, 44(18):  364-371.  doi:10.7506/spkx1002-6630-20221108-080
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A gas chromatography-tandem mass spectrometry (GC-MS/MS) with stable isotope dilution (SID) method was established for the determination of dioxins (PCDD/Fs) and dioxin like polychlorinated biphenyls (DL-PCBs) in pork. The sample was extracted by Soxhlet extraction, defatted using acidified silica gel, and purified by sequential chromatography on a multi-layer silica gel column, an alkaline alumina column and an activated carbon column. PCDD/Fs- and DL-PCBs-rich fractions were collected separately, concentrated and redissolved for GC-MS/MS analysis. The influence of defatting method and fat content on the purification efficiency was investigated, and the blank contamination source of the system was discussed. The results showed that gel permeation chromatography (GPC) degreasing had a great influence on the recovery of PCDD/Fs and DL-PCBs, and acidified silica gel degreasing was more suitable for the analysis and detection of ultra-trace dioxins. The major blank pollution source of the dioxin detection system was the Soxhlet extractor. The recoveries for internal standard were 45.3%–122.4% with relative standard deviation (RSD) of 1.0%–14.7%. The recoveries for the analytes were 64.9%–118.8% with RSD of 1.1%–14.7%. Both the linearity and limit of detection (LOD) met the requirements of the national standard of China GB 5009.205-2013. The proposed method is suitable for the determination of PCDD/Fs and DL-PCBs in actual pork samples.
Synthesis of Bifunctionalized Mesoporous Silica and Its Application in the Detection of Aminoglycosides
TANG Minmin, SHAO Xuemei, GAO Wei, SONG Jinzhu, ZHU Xiaojun, FENG Yun, ZOU Jie, XU Danke
2023, 44(18):  372-380.  doi:10.7506/spkx1002-6630-20230130-178
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In this work, a novel reversed phase/cation exchange mixed-mode sorbent for solid phase extraction (SPE) was synthesized by modifying the mesoporous material MCM-41 with 3-aminopropyltriethoxysilane (APTES) and octadecyltrimethoxysilane (C18 TMS) followed by succinic acid glycoside (SAA) in two steps. Various SPE parameters were optimized, and an analytical method was developed for the determination of six aminoglycosides (AGs) in milk by ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) using an amide column for chromatographic separation. The results of structural characterization demonstrated that the synthesized material had a large specific surface area, uniform pore size, and modified functional groups, exhibiting strong adsorption capacity and selectivity for AGs. The bifunctionalized material significantly improved matrix interference of milk compared to the carboxylated material. The proposed method exhibited good linearity for the six AGs in the concentration range of 5–500 ng/mL with correlation coefficients of 0.992 7–0.999 5. The limits of detection (LOD) achieved using this method were 0.30–2.50 ng/mL. The average recoveries ranged from 74.3% to 107% with relative standard deviation (RSD) between 2.6% and 15.7%. The material is characterized by low cost, simple preparation and good operability as a SPE filler. It can be extended to the detection of AGs in other food or agricultural samples.
Preparation of Test Strip Based on Au@Pt Nanozymes for Rapid Detection of Hydrogen Peroxide Residues in Foods
LI Boran, CHENG Luyao, CAO Zhihong, CHENG Nan, LUO Yunbo
2023, 44(18):  381-388.  doi:10.7506/spkx1002-6630-20220909-089
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A gold@platinum (Au@Pt) nanozyme-based test strip for the convenient and sensitive detection of hydrogen peroxide residues in foods was developed using Whatman No.1 filter paper as the carrier, Au@Pt nanozyme as the catalyst, 3,3’,5,5’-tetramethylbenzidine as the color developer, and acetic acid-sodium acetate solution at pH 4.6 as the buffer system. The optimized test strip soaking time was 10 s. The test paper could detect hydrogen peroxide in samples within 3 min with a minimum detection limit of 0.34 mg/kg and a good linearity (R2 = 0.993 1). The test paper is easy to make, highly accurate, stable and simple to operate, and can be used in the real-time on-site detection of food products in various distribution channels.