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25 April 2021, Volume 42 Issue 8
Food Chemistry
Stability and in Vitro Digestion of Pectin-Walnut Proteins Stabilized Emulsions Encapsulating Curcumin
LÜ Siyi, LU Qi, PAN Siyi
2021, 42(8):  1-9.  doi:10.7506/spkx1002-6630-20200406-069
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This research aimed to develop novel complexes between walnut proteins and pectin to be used as a delivery system for curcumin. Curcumin was successfully encapsulated in pectin-walnut proteins stabilized emulsions. After 14 days of storage, the particle size (D4,3) of the curcumin-encapsulating emulsions increased slightly, and no phase separation occurred. Under ambient pressure, the pectin-walnut albumin stabilized emulsion encapsulating curcumin showed good stability against salt (up to 300 mmol/L) and heat (up to 90 ℃). The pectin-walnut globulin stabilized and pectin-walnut glutelin stabilized emulsions encapsulating curcumin showed instability against salt (100 mmol/L). During in vitro digestion, the pectin-walnut albumin stabilized emulsion could quickly release the encapsulated bioactive substances. The pectin-walnut globulin stabilized emulsion was a suitable encapsulating material, which could maintain the stability of the encapsulated curcumin in acid medium, and delay its release during in vitro digestion, improving its bioaccessibility and making it easier to be absorbed by the small intestine. However, the pectin-walnut glutelin stabilized emulsion could delay the release of the encapsulated curcumin during in vitro digestion, resulting in low bioaccessibility. The current work indicated that the pectin-walnut proteins stabilized emulsions could be applied as an effective delivery system for bioactive compounds in a wide range of functional foods.
Effect of Pyrophosphate on the Dissociation of Natural and Oxidized Actomyosin
LÜ Bowen, GENG Xiaoqian, ZHANG Yueshu, YU Ze, LI Chunqiang, SHAO Junhua
2021, 42(8):  10-16.  doi:10.7506/spkx1002-6630-20200319-287
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The effect of sodium pyrophosphate (TSPP) on the oxidation and dissociation of actomyosin was investigated together with the influence on the effect of adenosine triphosphate (ATP) in dissociating differently oxidized actomyosin. Results showed that TSPP could inhibit the oxidation of actomyosin by 1 mmol/L H2O2, but have no obvious inhibitory effect on actomyosin oxidation by 10 and 20 mmol/L H2O2. After being treated separately with TSPP and ATP, the intrinsic viscosity of non-oxidized actomyosin decreased by 30.4% and 41.9%, and the average particle size by 21.2% and 48.4%, respectively. After being treated sequentially with TSPP and ATP, the intrinsic viscosity and average particle size of non-oxidized actomyosin increased to some extent. ATP induced a slightly greater change in the relative viscosity of the oxidized samples than that of the nonoxidized samples. However, the increase in the relative viscosity was weakened by TSPP treatment. Undissociated fibrous actomyosin existed in TSPP treated samples with and without ATP treatment, forming large aggregates upon oxidation. In general, TSPP was far less than effective in dissociating actomyosin than ATP, and even it had an inhibitory effect. Oxidation could promote the dissociation of ATP to a certain extent, but this effect was inhibited by TSPP. The results provide a theoretical basis for applying TSPP for improving the quality of meat products.
Effects of Adding Magnesium Chloride and Glutamine Transaminase after Cooling to Different Temperatures on the Rheological Properties of Heat-induced Gels from Whole Soybean Flour
LI Jun, KANG Xin, PU Xueli, CUI Huaitian, WANG Shengnan, SONG Hong, ZHU Danshi, LIU He
2021, 42(8):  17-21.  doi:10.7506/spkx1002-6630-20191029-315
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Heat-induced gels were prepared by boiling ultrafine whole soybean flour in water and then adding magnesium chloride and glutamine transaminase (TGase) after cooking. This study aimed to investigate the effect of cooling temperature, magnesium chloride concentration and TGase concentration on texture, rheological and microstructural properties of heat-induced gels. The results showed that the strength and rupture distance of gels with both added magnesium chloride and TGase increased with decreasing the cooling temperature from 25 to 15 to 5 ℃, and were significantly affected by the different cooling temperatures. The gel strength reached the maximum value of 56.23 g when the temperature was cooled to 5 ℃ and 0.8% magnesium chloride and 1.2 U/g TGase were added. The rheological results showed that the storage modulus (G’) of gels with added TGase showed a rapid upward trend. And the G’ of gels with TGase addition after cooling to 5 and 15 ℃ was significantly larger than that of their counterparts without TGase. The lower the cooling temperature, the larger the G’ of the gel. The scanning electron microscopic (SEM) images showed that the network structure of the gels formed by adding magnesium chloride and TGase after cooling to 5 ℃ was more orderly and denser.
Structural Analysis and Characterization of Cross-linked Complex between Plasma Amine Oxidase and Gelatin
LIU Xinzhu, CHENG Shan, LI Yu, WANG Wenhang
2021, 42(8):  22-28.  doi:10.7506/spkx1002-6630-20200318-278
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In this study, the effect of cross-linking with different amounts of plasma amine oxidase (PAO) in the presence and absence of Cu2+ as a co-factor on gelatin structure and rheological properties was investigated. The results of sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that the contents of large molecular mass bands in gelatin increased with increasing PAO level. Meantime, scanning electron microscopic (SEM) results revealed that the structure of gelatin became more compact with smaller empty space. The results of differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy (FTIR) and fluorescence spectroscopy demonstrated that the network structure of gelatin was greatly affected by treatment with PAO and the best cross-linking effect was achieved with a PAO concentration of 50 U/g, as indicated by the largest number of large molecular mass components, the highest thermal stability and fluorescence intensity, and the denser and more orderly network structure. We found that 100 mmol/mL Cu2+ did not play a positive role in the cross-linking of gelatin. In view of the positive results above, this study provides a potential approach for crosslinking gelatin and other proteins.
Spectroscopic Analysis of Interaction between Gliadin and Quercetin under Different NaCl Concentrations
WANG Qiming, TANG Yuwan, LI Chunyi, ZHAO Jichun, ZHANG Yuhao, MING Jian
2021, 42(8):  29-39.  doi:10.7506/spkx1002-6630-20200724-329
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The interaction between gliadin (G) and quercetin (Q) under different NaCl concentrations was studied by fluorescence spectroscopy, ultraviolet-visible (UV-Vis) spectroscopy and Fourier transform infrared (FTIR) spectroscopy. The results showed that at all NaCl concentrations, Q led to fluorescence quenching of G accompanied by a distinct blue shift (about 10 nm) of the fluorescence intensity with increasing NaCl concentration. When the concentration of Q was 50 μmol/L, 96%–98% of the protein fluorescence was quenched, indicating that strong interactions existed between G and Q. Q quenched the fluorescence intensity of G through static and static-dynamic modes. At 50 mmol/L NaCl concentration, the association constant (Ka) and the number of binding sites (n) were the largest, 4.87 × 107 L/mol and 1.477 1, respectively, indicating stronger interactions between G and Q at proper salt concentration. Thermodynamic data indicated that hydrophobic interaction was responsible for the formation of complexes at 50 mmol/L NaCl concentration, while hydrogen bonding was the main driving force at other NaCl concentrations. Synchronous fluorescence and UV-Vis spectroscopy suggested that Q changed the microenvironment surrounding the aromatic amino acid residues and the protein conformation. Tryptophan residues greatly contributed to quenching the protein fluorescence. FTIR spectroscopy and Raman spectroscopy revealed that specific interaction modes of G with Q existed at specific NaCl concentrations, and hydrogen bonding and hydrophobic interaction played important roles in the formation of the complexes, changing the secondary and tertiary structures and the microenvironment around the amino acid side chain of G. The above results proved that NaCl concentration influenced protein-polyphenol interaction; adding proper NaCl concentrations could facilitate the binding between gliadin and quercetin and changes in protein conformation. Our study provides a theoretical basis for the development of quercetin as a natural food additive in wheat products.
Effects of Salt Concentration on Texture, Microstructure and Quality of Wet-Cured Grass Carp Muscle
JIANG Jingdan, YANG Mingyuan, XU Changhua, SHI Wenzheng, LU Ying
2021, 42(8):  40-45.  doi:10.7506/spkx1002-6630-20200226-298
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In order to investigate the effect of salt concentration on the texture, microstructure and quality of wet-cured grass carp muscle, we studied changes in the centrifugal loss, thiobarbituric acid reactive substances (TBARS), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) pattern, scanning electron microscopic (SEM) image, transmission electron microscopic (TEM) image, Fourier transform infrared (FT-IR) spectrum and low-field nuclear magnetic resonance (LF-NMR) spectrum of cured fish samples as a function of salt concentration. Also, we monitored the protein, moisture and fat contents, microstructure and texture. Our results showed that the crude protein content decreased by 2%, the centrifugal loss rate decreased by 8%, and the TBARS value increased about twice when salt concentration increased from 30 to 120 mg/mL. NMR data revealed that bound water and free water were converted into non-flowable water with increasing salt concentration. The second-derivative infrared spectrum showed that the intensity of the 1 747 cm-1 peak was significantly reduced after curing, and the intensity of the peaks in the ranges of 1 612–1 698 cm-1 and 1 523–1 553 cm-1 was significantly increased at a salt concentration of 30 mg/mL. The peak intensity gradually decreased with increasing salt concentration. SEM and TEM results showed that fish muscle fibers became blurred at 30 mg/mL salt concentration, while the sarcoplasmic tissue remained intact. The morphology of sarcoplasma became loose at 120 mg/mL salt concentration, and the spatial structure of fish protein was destroyed, thus making fish muscle tissue loose, increasing water loss, and reducing the eating quality of fish meat. However, 30 mg/mL salt concentration had the smallest effects on fish muscle texture. Our study can provide a scientific basis for the development of high-quality ready-to-eat aquatic products in the future.
Effect of Auricularia cornea Polysaccharide on Gelatinization, Rheological and Retrogradation Characteristics of Coarse Cereal Porridge
SUN Cong, ZHENG Mingzhu, XU Xiuying, CAI Dan, LIU Meihong, CAO Yong, LIU Jingsheng
2021, 42(8):  46-51.  doi:10.7506/spkx1002-6630-20191024-253
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A canned coarse cereal porridge was made from waxy corn, oats, millet, red beans, mung beans, and cowpea with added Auricularia cornea polysaccharide (ACP) as a stabilizer. The effects of A. cornea polysaccharide on the gelatinization, rheological, and retrogradation characteristics of porridge were measured by rapid visco analyzer (RVA), rheometer, and differential scanning calorimeter (DSC) combined with Avrami model. The results showed that the addition of ACP was positively correlated with the peak viscosity and final viscosity of porridge, but negatively correlated with the setback and breakdown values. The shear stress was significantly (P < 0.05) increased with increasing ACP addition, while ACP did not change fluid properties of the porridge. The porridge with ACP was still a pseudoplastic fluid. The increase in recrystallization temperature (T0 and TC) of the porridge samples was slowed down with the addition of ACP, and the aging ΔH was decreased by 46%. The Avrami model of aging kinetics showed that the amount of A. cornea polysaccharide added to the porridge was negatively correlated with the crystallization rate k. The porridge with 9% ACP had the best processing and storage quality.
Mechanism and Thermodynamic Characterization of Electrostatic Complexation between Gelatin and Sodium Alginate
CHEN Haihua, YU Rui, WANG Yusheng
2021, 42(8):  52-59.  doi:10.7506/spkx1002-6630-20200322-321
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This work aimed to investigate the formation mechanism of gelatin-sodium alginate complex (GEL-AG) at room temperature. The effects of GEL-to-AG ratio (m/m) and pH on the electrostatic interaction between the two compounds were studied by turbidity titration, zeta potential measurement, and methylene blue spectrophotometry. Isothermal titration calorimetry (ITC) was employed to elucidate the thermodynamic behavior of GEL-AG interaction. The results showed that the complex formation process was a spontaneous exothermic process driven by enthalpy through strong electrostatic interaction. The environmental pH influences the electrostatic interaction between GEL and AG through adjusting the net charge carried by them; insoluble complexes were formed at low pH and soluble complexes at high pH. Low concentration of NaCl (< 0.02 mol/L) could promote the combination between GEL and AG through electrostatic shielding, while high concentration of NaCl could inhibit the interaction. The electrostatic interaction between GEL and AG could be strengthened with increasing GEL-to-AG ratio or their respective total concentrations. At a GEL-to-AG ratio of 7:3, a stable complex suspension was obtained at a high pH, while pH in the range of 2.5–3.5 showed little influence on the properties of GEL-AG suspension. This study will be useful for the development of GEL-AG-based biomaterials and their application in the food and pharmaceutical fields.
Preparation and Physicochemical Characterization of Cinnamic Acid Based Oleogels and Oleogel Emulsions
LUAN Huilin, WU Yuqing, ZHENG Hongxia, KE Xiangyu, MAO Like
2021, 42(8):  60-66.  doi:10.7506/spkx1002-6630-20191024-269
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In this study, corn oil and cinnamic acid were respectively used as a solvent and a gelling agent to prepare oleogels. Its rheology, texture, oil-holding capacity, thermodynamic properties and gel crystal morphology were investigated. Further, oil-in-water (O/W) oleogel emulsions were prepared and its physical properties such as particle size, rheology and stability were analyzed. The critical gelling concentration of cinnamic acid was 4%, and the oil-holding capacity of oleogels stored at 4 ℃ was higher than that at room temperature. The hardness and thermal stability of oleogels were higher with increasing concentration of cinnamic acid. Cinnamic acid was randomly distributed in oleogels as long fibrous aggregates. Oleogel emulsions with lower oil phase content in the range from 10% to 20% had smaller particle size, lower viscosity and better physical stability. At room temperature, the oleogels had poorer stability, while the oleogel emulsions could maintain good stability, which will be useful for its application in the food industry.
Improvement Effects of Structural Changes of Pork Myofibrillar Protein Induced by Rutin on Its Gel Properties
JIA Na, LIN Shiwen, LIU Dan, LIU Dengyong
2021, 42(8):  67-73.  doi:10.7506/spkx1002-6630-20191018-183
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Different concentrations of rutin (0, 10, 50, 100 and 150 μmol/g protein) was added to Fenton oxidation system containing pork myofibrillar protein (40 mg/mL protein, 10 μmol/L FeCl3, 100 μmol/L VC and 1 mmol/L H2O2). To study the effects of rutin on the structure and gel properties of myofibrillar protein, we determined the sulfhydryl content, surface hydrophobicity, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) pattern, solubility, gel strength, water-holding capacity and rheological properties of the protein. The results showed that the addition of rutin significantly decreased the sulfhydryl content of myofibrillar protein (P < 0.05), and first decreased and then increased the surface hydrophobicity. Rutin addition reduced the intensity of myosin heavy chain (MHC). The intensity of actin band was decreased at higher concentrations of rutin (100 and 150 μmol/g). Most of the MHC and actin bands were restored after adding β-mercaptoethanol. The solubility was decreased with increasing rutin concentration, and the gel strength and water-holding capacity were increased significantly (P < 0.05). Similarly, the storage modulus (G’) in the final stage of gel formation was increased. Therefore, rutin can improve the gel properties of myofibrillar protein by covalent crosslinking with it or moderately increasing its surface hydrophobicity.
Comparison of Reactive Sites of Different Fatty Acid Molecules by Quantum Chemistry Calculation
WANG Chunchun, QIN Xiaoli, KAN Jianquan, LIU Xiong, SUO Huayi, ZHONG Jinfeng
2021, 42(8):  74-80.  doi:10.7506/spkx1002-6630-20200316-253
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In order to explore the similarities and differences in their reactive sites, the structures of six fatty acids with different chain lengths and unsaturation degrees were optimized by software Gaussian 09 at the theoretical level of m06-2x/def2tvzp, and the single point energy was calculated using the ma-def2tzvp basis in this study. Afterward, the surface electrostatic potential, average local ionization energy, frontier molecular orbital and condensed Fukui function were analyzed and visualization results were obtained. The results suggested that the nucleophilic reaction site of saturated fatty acids was hydroxyl hydrogens, and the electrophilic reaction site of caprylic acid was near the oxygen atoms. The carboxyl groups of palmitic acid and stearic acid showed no electrophilic activity with the increase in chain length. The unsaturated fatty acid oleic acid exhibited higher nucleophilic electrophilic activity near the oxygen atoms, and higher electrophilic activity at the double bonds. As the number of double bonds increased, the reactive sites of linoleic acid and linolenic acid were both double bonds. The reactivity of fatty acids with the same chain length enhanced with increasing unsaturation degree. The results of this study can provide a theoretical basis for revealing the relationship between the structure and properties of fatty acids, and provide a reference for the selection of fatty acids in the edible oil industry.
Construction and Properties of Polyglycerol Monolaurate-Oleic Acid Microemulsion System
ZHAO Jingsong, WANG Yusheng, LI Tianhao, ZHAO Wenqing, CHEN Haihua
2021, 42(8):  81-89.  doi:10.7506/spkx1002-6630-20191017-167
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In order to expand the application of curcumin in foods, a food-grade dilution-stable microemulsion with high curcumin in it was prepared with oleic acid as oil phase, polyglycerol monolaurates with different degrees of polymerization as surfactants and polyols with different carbon chain lengths as cosurfactants. A suitable surfactant and cosurfactant were selected by analyzing the pseudo ternary phase diagrams. Furthermore, the effects of the mass ratio of surfactant to cosurfactant (Km), aqueous phase pH, and NaCl concentration on the phase behavior of microemulsions were investigated. The structure of microemulsions was analyzed by the conductivity method. The particle size and morphology of microemulsion droplets were determined by laser particle size analyzer and transmission electron microscope (TEM). The results showed that the best emulsification efficiency together with the largest proportion of microemulsion area was obtained under the following conditions: a 2:1 mixture of hexaglycerol and decaglycerol monolaurates as the surfactant, ethanol the cosurfactant and a Km value of 1/2. The emulsion droplets were uniformly spherical in shape with particle size of 10–100 nm. The addition of NaCl and an aqueous phase at non-neutral pH could led to a decrease in the proportion of microemulsion area and an increase in the minimum dilute ratio (DR). An infinitely dilutable microemulsion could be formed at a mass ratio of surfactant to oil phase (SOR) of 9:1. A W/O microemulsion could be obtained with water content less than 45%. The microemulsion could be transformed into double continuous type with increasing water content up to 60% and then into O/W-type at higher water content. Microemulsification could increase the solubility of curcumin to 4.87 mg/mL without influencing the biological activity of curcumin. This indicated that the microemulsion might be an excellent carrier for curcumin that could efficiently improve its poor solubility and dilutability.
Effect of Soybean Oil and Pre-emulsified Soybean Oil on the Quality of Nemipterus virgatus Surimi Gel
SONG Chunyong, HONG Pengzhi, ZHOU Chunxia, CHEN Ailin, FENG Rui
2021, 42(8):  90-97.  doi:10.7506/spkx1002-6630-20200820-275
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In order to improve the quality of surimi products, we investigated the effect of 2 mL/100 g soybean oil and 10 mL/100 g pre-emulsified soybean oil (i.e. soybean oil emulsion pre-emulsified by whey protein isolate or sodium caseinate, containing 2 mL/100 g soybean oil) on the gel properties, microstructure and sensory characteristics of Nemipterus virgatus surimi gel, and we compared the effect on surimi gel quality. The results showed that the whiteness of Nemipterus virgatus surimi gel was improved by adding 2 mL/100 g soybean oil, while the texture properties were deteriorated, accompanied by decreased water-holding capacity and increased cooking loss rate (P < 0.05). An uneven distribution of oil droplets with a diameter in the range of 0.10–0.175 μm was observed in the gel matrix accompanied by obvious aggregation of oil droplets. And the gel with soybean oil exhibited a three-dimensional network with large cavities, and the oxidizability of lipids in it was significantly increased compared with the control without soybean oil (P < 0.05). However, after adding 10 mL/100 g pre-emulsified soybean oil, the whiteness, water-holding capacity and bound water content were significantly increased together with a decrease in the cooking loss rate (P < 0.05), whereas the texture and the oxidizability of lipids were not significantly changed (P > 0.05). Furthermore, the sensory properties of surimi gel were significantly improved. The pre-emulsified oil droplets with a diameter of less than 0.10 μm were evenly distributed in the gel matrix, and the gel exhibited a three-dimensional network with small cavities and smooth surface. Therefore, pre-emulsified soybean oil with milk protein can effectively improve the gel properties and flavor of Nemipterus virgatus surimi, which will be useful for the development of new surimi products.
Bioengineering
High-level Expression, Characterization, and Application of a Novel β-Mannanase from Penicillium chrysogenum
ZHEN Hongmin, HUA Xiaohan, MA Junwen, WEN Yongping, LI Yanxiao, YAN Qiaojuan, JIANG Zhengqiang
2021, 42(8):  98-105.  doi:10.7506/spkx1002-6630-20200819-248
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A cloned β-mannanase (PcMan26A) gene from Penicillium chrysogenum was successfully expressed in Penicillium chrysogenum. At the end of high cell density fermentation, the enzymatic activity of the fermentation supernatant reached 25 200 U/mL. The enzyme belonged to the glycoside hydrolase family 26 and shared the highest amino acid sequence identity (67.8%) with β-mannanase from Aspergillus niger CBS 513.88. The optimal reaction conditions for PcMan26A were pH 6.0 and 50 ℃, and it was stable at 45 ℃ and within a broad pH range of 4.0–8.0. The enzyme showed the highest specific activity (3 581.0 U/mg) towards konjac powder. Furthermore, PcMan26A was used for konjac powder hydrolysis, yielding konjac manno-oligosaccharide with a yield of 86.2%. The main composition of the konjac manno-oligosaccharide was manno-oligosaccharides with degree of polymerization > 4. The recombinant β-mannanase provides a new option for the enzymatic production of konjac manno-oligosaccharide.
High-throughput Sequencing Reveals the Microbial Community Structure in Taorong-type Daqu
CHEN Meng’en, ZHAO Cong, HAN Suna, ZHOU Danfeng, LI Jianmin, ZHANG Jin, HUANG Runna, WANG Xiaoyi, DENG Jie, HU Xiaolong
2021, 42(8):  106-113.  doi:10.7506/spkx1002-6630-20191229-339
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The microbial community structure in mature Taorong-type Daqu was studied by high-throughput sequencing, and its correlation with the physicochemical properties of Daqu was analyzed. The results showed that a total of 83 bacterial operational taxonomic units (OTUs) were detected in Daqu. The main bacterial phyla included Firmicutes (79.15%), Proteobacteria (15.30%), Actinobacteria (4.05%), Cyanobacteria (1.42%), and others (0.08%). The dominant genera were Lactobacillus, Weissella, Pantoea, Leuconostoc, Kroppenstedtia, Saccharopolyspora, Pediococcus, Bacillus, Enterobacteriaceae_unclassified, Mitochondria_norank, Burkholderia and Cyanobacteria_norank. A total of 34 fungal OTUs were detected, among them, Ascomycota was absolutely dominant (99.19%). The dominant fungi were unidentified, Thermomyces, Thermoascus, Aspergillus, Rhizomucor, Wickerhamomyces and Kazachstania. The bacterial community in Taorong-type Daqu was much more diverse than the fungal community. The major microorganisms were predominant, and all identified microbial genera were significantly correlated with the physicochemical properties of Daqu. This study will be of great significance to guide the optimization of the production of Taorong-type Daqu and liquor.
Screening of Five Non-Saccharomyces Strains with Low Ethanol Yield and Evaluation of Their Oenological Properties
ZHANG Boqin, FANG Zizhuang, CHENG Chifang, ZHANG Ruyi, DUAN Changqing, YAN Guoliang
2021, 42(8):  114-121.  doi:10.7506/spkx1002-6630-20200315-229
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This study aimed to obtain low ethanol-producing non-Saccharomyces strains for use in wine production. We evaluated the ethanol-producing ability of 38 indigenous non-Saccharomyces strains isolated in Hunan, Liaoning and Xinjiang provinces by pure culture fermentation. Five strains with lower ethanol-producing ability in terms of ethanol volume, ethanol yield and potential ethanol content than S. cerevisiae EC1118 were selected, including Rhodotorula mucilaginosa CVE-RM3, Hanseniaspora uvarum CVE-HU42, Pichia kudriavzevii CVE-PK1, Metschnikowia pulcherrima CVE-MP33 and H. vineae CVE-HV6. Their oenological characteristics were further investigated in mixed culture fermentation when inoculated with S. cerevisiae EC1118. The results showed that mixed culture fermentation with CVE-HV6 and EC1118 was the most effective in reducing ethanol, with a reduction of 0.72% (V/V) as compared to pure culture fermentation with EC1118. Furthermore, the mixed culture fermentation increased the contents of the major aroma compounds ethyl acetate, ethyl acetate, 2-phenylethanol and linalool by 281.22%, 3 503.87%, 36.53% and 49.03%, respectively, and reduced the contents of hexanoic acid and octanoic acid by 86.60% and 92.01%, respectively, significantly improving the aroma profile and enhancing the fruity and flower intensity of wine. In conclusion, H. vineae CVE-HV6, with low ethanol yield and good oenological characteristics, has the potential to be used for producing low-alcohol wine.
Screening of Low-temperature Protease-producing Planococcus and Its Application in Low-salt Fish Sauce Fermentation
ZHOU Jing, YUAN Li, GAO Ruichang
2021, 42(8):  122-128.  doi:10.7506/spkx1002-6630-20200314-220
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We investigated the effects of four strains of Planococcus with protease activity at low temperature (15 ℃) isolated from traditional fermented shrimp paste on the quality of low-salt fish sauce. The strains were screened for their enzymatic activity at 40 ℃. Among these, strain XJ11 showed the highest protease activity of (10.07 ± 0.11) U/mL. Based on morphological observation and 16S rRNA sequence alignment, all four strains were identified as Planococcus and named P. maritimus XJ2, P. plakortidis XJ10, P. dechangensis XJ11 and P. rifietoensis XJ12, respectively. The strains grew well in a low-temperature, low-salt, and alkaline environment. The protease produced by Planococcus exhibited the highest enzymatic activity at 40 ℃ at pH 9.0 and showed good salt tolerance. A 1:1:1:1 mixture of the four strains was inoculated at 3% as a starter culture in low-salt fish sauce fermentation. The results showed that the amino acid nitrogen content of low-salt fish sauce inoculated with the starter culture was (1.33 ± 0.08) g/100 mL, which was significantly higher than that of the control prepared without the starter culture ((0.89 ± 0.02) g/100 mL) (P < 0.01). The amounts of the key desirable volatiles such as alcohols, ketones, acids, esters and pyrazine were higher than in the control. In summary, the starter culture increased the amino acid nitrogen content of low-salt fish sauce and improved the flavor effectively.
Isolation of Antagonistic Bacteria from Smoked Bacon and Analysis of Antifungal Properties of Its Lipopeptides
GAO Zhaojian, QIN Chenkai, HUANG Lianghao, ZHU Shuang, ZHU Jinhu, ZHAO Yifeng
2021, 42(8):  129-136.  doi:10.7506/spkx1002-6630-20200321-309
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This study aimed to isolate and identify antagonistic bacteria from traditional smoked bacon, to determine the bacteriostatic activity of one selected isolate, and to purify and identify lipopeptide from its fermentation broth for the purpose of providing a theoretical basis for the application of the antimicrobial lipopeptide in food preservation. The inhibition zone method was adopted for screening the antagonistic strains. The antimicrobial lipopeptide was isolated and purified using a simple three-step procedure involving acid precipitation, silica gel column chromatography and reversed-phase high performance liquid chromatography (RP-HPLC), and it was identified by Fourier transform infrared spectroscopy (FT-IR) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). A total of 12 bacterial isolates were obtained, including XLP27, which had obvious antifungal and antibacterial effects on both Gram-positive and Gram-negative strains as well as one yeast strain and four mould strains. The antagonistic strain XLP27 was identified as Bacillus pumilu. The antibacterial substances produced by strain XLP27 were synthesized mainly in the stationary growth phase and the maximum yield was reached after 60 h of culture. One antimicrobial compound was purified from the fermentation broth and identified as lipopeptide iturin A, which had five isoforms with protonated masses of m/z 1 046.182, 1 060.213, 1 074.198, 1 088.201 and 1 102.175, representing fatty acid chain length of C14–C18. The lipopeptide was extremely heat stable (30 min at 40–100 ℃), remained active over a wide pH range (4–10) and was found to be resistant to all proteolytic enzymes tested. Iturin A had broad-spectrum antibacterial activity and good stability, and thus could have the potential to be an alternative to traditional antibacterial agents in the fields of food preservation and agricultural biological control.
Monascus Fermentation Enhanced the Umami Taste of Low-Salt Semi-Dried Grass Carp (Ctenopharyngodon idellus)
WU Kang, XIE Jinghui, WANG Qianqian, QI Ming, WU Jianzhong
2021, 42(8):  137-142.  doi:10.7506/spkx1002-6630-20200228-316
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To evaluate the effect of fermentation by Monascus purpureus on enhancing the umami taste of low-salt semi-dry grass carp, conventional low-salt semi-dry fish and low-salt semi-dried fish with inactivated Monascus purpureus were used as controls. The extent of proteolysis was evaluated by water soluble nitrogen (WSN) and tricholoracetate acid soluble nitrogen (TCA-SN) contents. Umami taste was evaluated by taste activity value (TAV) and equivalent umami concentration (EUC), which were calculated by free amino acids (FAA) and 5’-nucleotides contents, respectively. The fermented fish was reddish in color and richer in umami taste. Large amounts of amino acids and peptides were released during fermentation. The EDU of the fermented fish was 7.34 g/100 g, which was 4.38 times as high as that of the unfermented fish with Monascus purpureus. WSN and TCA-SN contents were significantly increased in the fermented sample. Likewise, the content of total free amino acids especially the umami amino acids glutamic acid (Glu) and aspartic acid (Asp), as well as the contents of umami nucleotides guanosine 5’-monophosphate (5’-GMP) and inosine 5’-monophosphate (5’-IMP), were significantly increased. The TAVs of 5’-GMP and Glu were more than 1, indicating that they are the major contributors to the umami taste. Monascus fermentation proved effective in enhancing the umami taste of low-salt semi-dried grass carp.
Analysis of Microbial Diversity in Sufu Using High-throughput Sequencing
TAO Kang, WU Lingwei, JIN Xiaofang, REN Kai, YU Zhengxian, LIU Tong, LIU Mingwei, WANG Shuixing
2021, 42(8):  143-149.  doi:10.7506/spkx1002-6630-20191230-362
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In order to study the influence of microorganisms present in the production environment on the quality and flavor of sufu, we analyzed the microbial diversity and abundance in natural fermented and pure-cultured fermented sufu by high-throughput sequencing based on 16S rDNA sequence for bacterial and internal transcribed spacer-1 (ITS1) sequence for fungi. The results showed that Proteobacteria, Firmicutes and Bacteroidetes were the main bacterial phyla in sufu. Ascomycetes, unclassified_k__Fungi, Basidiomycetes and Mucoromycota were the main fungal phyla. At the species level, the abundance of Pseudomonas fragi was very high in samples A (2.30%), B (20.95%), C (12.51%) and D (48.75%), being higher in natural fermented sufu than in pure-cultured fermented sufu. The main fungus was yeast, and sample A mainly contained unclassified_k__Fungi (63.20%) and Millerozyma farinosa (25.80%). In samples B, C and D, Debaryomyces prosopidis was dominant, with relative abundance of 37.60%, 51.70% and 85.90%, respectively.
Structure and Diversity Analysis of Mold Community in Main Maotai-flavor Baijiu Brewing Areas of Maotai Town Using High-throughput Sequencing
ZHU Zhiyu, HUANG Yongguang
2021, 42(8):  150-156.  doi:10.7506/spkx1002-6630-20191227-332
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The present work aimed to systematically study the structure and diversity of the mold communities in Daqu and environmental samples from seven main Maotai-flavor Baijiu brewing areas of Maotai town by high-throughput sequencing on an MiSeq platform. A total of 95 genera were detected, including 68 genera from Daqu samples and 89 genera from environmental samples. Aspergillus, Byssochlamys, Monascus, Thermoascus, Thermomyces, Rhizomorcor were identified as the dominant molds in Daqu, while Alternaria, Aspergillus, Byssochlamys, Cladosporium, Fusicolla, Lecythophora, Monascus, Penicillium, Phoma, Pyrenochaeta, Thermoascus, Thermomyces and Toxicocladosporium as the dominant molds in the environmental samples. We analyzed the differences in mold community in terms of diversity index, enrichment rate, decay rate and mobility between the main brewing areas, and between Daqu and environmental samples from each brewing area, and we dissected the interaction between molds in Daqu and those in the brewing environment, which will provide a theoretical basis for deciding where to site a new winery from a microbial perspective.
Preparation and Characterization of Peptide-Fe2+ Complexes from Tilapia Skin Protein
LIN Shanting, HU Xiao, LI Laihao, YANG Xianqing, CHEN Shengjun, WU Yanyan, HUANG Hui, RONG Hui
2021, 42(8):  157-164.  doi:10.7506/spkx1002-6630-20200307-117
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In this research, we optimized the preparation conditions of Fe2+-chelating peptides from tilapia skin protein, and we structurally characterize the complex. Five commercial proteases were screened for the highest Fe2+-chelating rate of peptides. The effects of chelating time, pH and temperature on the Fe2+-chelating rate were evaluated, and the changes in the fluorescence, ultraviolet and infrared spectra of the peptides were examined before and after chelating with Fe2+. In addition, the molecular mass distribution, amino acid composition and antioxidant activity of the peptides were measured. The results showed that the peptides obtained after 2 h of hydrolysis with trypsin had the highest Fe2+-chelating rate. The optimal chelating conditions were pH 5, 37 ℃, and 90 min. Chelating with Fe2+ resulted in folding of the peptide chain, decreased intrinsic fluorescence intensity, and red shift of the characteristic ultraviolet absorption band. Fe2+ was likely bound to the amino, carbonyl and carboxyl groups of the peptides. The molecular mass of the Fe2+-chelating peptides was about 500–3 000 Da, and the peptide chains were rich in Asp, Glu, Gly, Arg and Pro. All hydrolysates had antioxidant capacity. The antioxidant capacity was not significantly changed after chelating with Fe2+. Although the antioxidant capacity of the trypsinic hydrolysate was significantly lower than that of glutathione, the Fe2+-chelating activity was significantly higher than that of glutathione. The results showed that the peptides in the hydrolysate had excellent Fe2+ -chelating activity suggesting the potential to be developed into a new dietary iron supplement.
Component Analysis
Change of Volatile Flavor Compounds in Tan Sheep Muscle during Cooking
SHAN Qimei, ZHAO Xiaoce, LUO Ruiming, YOU Liqin
2021, 42(8):  165-171.  doi:10.7506/spkx1002-6630-20191024-252
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In this study, the changes of volatile flavor compounds in the Longissimus dorsi muscle of Yanchi Tan sheep during cooking were studied by purge-and-trap (P&T)-gas chromatography-mass spectrometry (GC-MS) and gas chromatography-olfactometry (GC-O). A total of 42 volatile compounds were detected, including aldehydes (16), ketones (6), alcohols (5), acids (3) and heterocyclic compounds (12). The results showed that the percentages of aldehydes, ketones and alcohols were high at cooking temperatures of 50–80 ℃, but decreased when the temperature increased to 90 ℃. After boiling, the percentages of pyrrole, pyridine, thiazole and other heterocyclic compounds increased with the prolongation of cooking time. 2-Undecanolaldehyde, 4-ethyl octanoic acid, 2-pentyl furan, 2,3,5,6-tetramethylpyrazine, dimethyl disulfide and benzyl methyl sulfide were identified as the characteristics flavor compounds of Tan sheep muscle by GC-O. The findings obtained in this study can provide a theoretical reference for the industrial processing and flavor control of Ningxia featured Tan sheep meat products such as hand-held mutton and stewed mutton.
Effect of Astaxanthin Microspheres on the Quality and Flavor of Yogurt
WANG Fuyun, WANG Chengcai, WANG Zheming, JIANG Hongrui, MIN Douyong, LIU Xiaoling, LI Shubo
2021, 42(8):  172-178.  doi:10.7506/spkx1002-6630-20200406-065
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Bovine milk added with astaxanthin microspheres was used to prepare yogurt containing astaxanthin microspheres in order to evaluate the effects of astaxanthin microspheres on the number of viable lactic acid bacteria, texture and sensory evaluation of yogurt. Furthermore, we tested the stability of astaxanthin incorporated in yogurt during storage. The volatile organic compounds of the yogurt were analyzed by headspace-gas chromatography-ion mobility spectrometry (HS-GC-IMS). The results showed that astaxanthin microspheres had a positive effect on the number of viable lactic acid bacteria, but had no significant influence on the texture of the yogurt (P > 0.05). The yogurt containing astaxanthin microspheres at 6 mg of astaxanthin/100 g of sample exhibited an orange-red color and the highest sensory evaluation score. The intensity of signal response to hexanal, contributing to the fishy smell, was decreased when astaxanthin was encapsulated by ethyl cellulose, and astaxanthin microspheres had no effect on the volatile flavor substances of yogurt. Compared with free astaxanthin, the retention rate of encapsulated astaxanthin incorporated in yogurt stored for 14 days at 4 ℃ under light was improved by 1.62-fold. Therefore, astaxanthin microspheres could enhance the application potential of astaxanthin in yogurt.
Effect of Different Pretreatments on the Free Amino Acids and Volatile Components of Bighead Carp Bone Protein Hydrolysates
DONG Ye, ZHANG Yiqi,, YAO Hongzheng, HE Guangxi, DAI Zhiyuan,
2021, 42(8):  179-185.  doi:10.7506/spkx1002-6630-20200406-068
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Bighead carp bone protein was first pretreated by steam explosion or thermally, and then hydrolyzed with trypsin. The resultant hydrolysates were designated as SFBPH and HFBPH, respectively, and their free amino acids, volatile components and relative molecular mass distribution were analyzed and compared with each other. The results showed that the hydrolysis degree of SFBPH was higher than that of HFBPH. In SFBPH and HFBPH, small molecular components with molecular mass less than 1 000 Da accounted for 73.78% and 66.56% of the total amount, respectively. The total amount of free amino acids in SFBPH was 28.36 mg/mL, which was 1.3 times higher than that of HFBPH. In SFBPH and HFBPH, the amount of bitter amino acids accounted for 64.88% and 64.23% of the total amino acids, respectively. The overall taste of the two hydrolysates was dominated by bitterness. A total of 77 volatile components were detected by gas chromatography-mass spectrometry analysis, of which, 61 were detected in SFBPH, and 47 in HFBPH, with hydrocarbons and aldehydes being the most abundant ones.
Flavor Characteristics of Porphyra haitanensis and Porphyra yezoensis: A Comparison Study Using Electronic Tongue and Gas Chromatography-Ion Mobility Spectrometry
CAO Rong, HU Mengyue, TAN Zhijun, WANG Lianzhu, LIU Qi
2021, 42(8):  186-191.  doi:10.7506/spkx1002-6630-20191110-101
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In order to evaluate the flavor characteristics of Porphyra haitanensis and P. yezoensis, we tested their taste profile and volatile components by electronic tongue (ET), gas chromatography-ion mobility spectrometry (GC-IMS), and investigated differences in the compositions of free amino acids and flavor nucleotides. The results showed that the umami, umami aftertaste, bitterness and astringency of P. haitanensis were pronouncedly higher than those of P. yezoensis, consistent with the results of free amino acids, flavor nucleotides and equivalent umami concentration. A total of 36 volatile compounds were isolated and identified from P. haitanensis and P. yezoensis by GC-IMS. The total amount of odorants in P. yezoensis was higher than that in P. haitanensis, with linalool oxide, 1-hexanal, butyrolactone, phenylacetaldehyde and nonanal being dominant. Generally, saturated alcohols and esters have high threshold values, contributing little to the odor of laver, while aldehydes such as malondialdehyde, phenylacetaldehyde and nonanal have low threshold values, greatly contributing to the overall pleasant odor of laver. It can be concluded that there are great differences in taste and aroma between P. haitanensis and P. yezoensis. The results of this study can provide reference for the high-valued and efficient utilization of lavers.
Quantification of Sweetness and Correlation with the Main Taste Compounds of Huangjincha Congou Black Tea
YU Penghui, HUANG Hao, ZHAO Xi, ZHONG Ni, GONG Yushun, ZHENG Hongfa
2021, 42(8):  192-197.  doi:10.7506/spkx1002-6630-20191103-024
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The sweetness intensity of Huangjincha Congou black tea infusion was quantitatively evaluated and the contents of the main taste substances in it were determined. Moreover, the correlation between them was investigated. The results showed obvious differences in the sweetness intensity of tea samples from different factories. Maltose was the most abundant sugar, and serine the most dominant sweet amino acid. The contents of sweet compounds including monosaccharides, disaccharides and sweet amino acids and theanine content were positively correlated with the sweetness intensity. The total sugar content was significantly (P < 0.05) positively (r = 0.781) correlated with the sweetness intensity. The contents of catechin, caffeine, gallic acid, theaflavins, and thearubigins were negatively correlated with the sweetness intensity. The contents of total catechins and theaflavins were significantly (P < 0.05) negatively (r = ?0.811 and ?0.781, respectively) correlated with the sweetness intensity. Epigallocatechin gallate (EGCG) could reduce the sweetness intensity of sugars in the tea. In view of the fact that sweetness is an important taste element of Congou black tea, this study provides a foundation for evaluating the quality of Congou black tea.
Analysis of Aroma Characteristics and Volatile Components of Juhong Tea, Manufactured from Black Tea with Added Citrus Peel
LIU Panpan, ZHENG Pengcheng, GONG Ziming, CHANG Zerui, SU Fangjun, HUANG Bo, FENG Lin, GAO Shiwei, ZHENG Lin
2021, 42(8):  198-205.  doi:10.7506/spkx1002-6630-20191102-013
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This study aimed to analyze the main volatile compounds and aroma profiles of Juhong tea and its raw materials. The volatiles were analyzed by combined use of quantitative descriptive analysis (QDA), headspace solid phase micro-extraction (HP-SPME) coupled with gas chromatography-mass spectrometry (GC-MS), gas chromatography-mass spectrometry-olfactometry (GC-MS-O), and multivariate statistical analysis. Sensory evaluation showed that fruity and sweet notes were the main odors of Juhong tea. A total of 92 volatile components, including 23 aldehydes, 23 alcohols, 20 alkenes, 9 ketones, 9 esters and 8 other compounds, were identified from Juhong tea and its raw materials. Alkenes and alcohols were the principal aroma types. The results of GC-O analysis combined with odor activity value indicated that limonene, β-ionone, linalool, decanal, β-damascenone, (E,E)-2,4-decadienal and octanal were the main aroma-active components. The results of correlation analysis showed that 11 volatile components, including myrcene, phenylacetaldehyde and linalool, contributed significantly to the fruity, floral, sweet, green and woody flavors. Orthogonal partial least squares discriminant analysis was performed, revealing the typical volatile components that differed between Juhong tea and its raw materials. These results provide an experimental basis for future studies on the aroma of Juhong tea and other herbal teas made from citrus fruits.
Analysis of Metabolites in Rice Produced in Different Regions by GC-MS-based Metabonomics
ZHANG Shu, WANG Changyuan, FENG Yuchao, SHENG Yanan, FU Tianxin, ZHANG Yiwei, JIANG Yingjun, YU Miao, ZHANG Liyuan
2021, 42(8):  206-213.  doi:10.7506/spkx1002-6630-20200409-128
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Using gas chromatograph-mass spectrometry (GC-MS)-based metabolomics, we investigated 60 rice samples collected from two geographical indication rice-producing regions in Heilongjiang province. The SIMCA software and the R software were used to analyze the obtained data by multivariate statistical analysis and metabolic pathway analysis such as principal component analysis (PCA), orthogonal partial least square discriminant analysis (OPLS-DA). We explored the effect of the geographical origin on rice metabolism and metabolites. A total of 167 metabolites were identified, and our analysis revealed that the geographical origin affected the number, type, and content of rice metabolites, having the most significant effect on the content. A total of 32 differential metabolites were identified, and hierarchical cluster analysis showed that they could be used as a reference index for identification of the geographical origin of rice. The OPLS-DA results showed a clear discrimination between rice samples from the two producing regions, and the samples showed a significantly clustered distribution, indicating that the geographical origin had a great influence on rice metabolism. Metabolic pathway analysis found that nine key pathways were highly correlated with the differential metabolites. We determined that the geographical origin had the greatest impact on the metabolism of lipids and amino acids in rice. The five metabolites of ureapropionic acid, squalene, cholesterol, lauric acid and fumaric acid could be of great reference significance for understanding the metabolic differences of rice from different producing regions. This research provides a theoretical basis for the identification of rice from different producing region and a reference for tracing the geographical origin of agricultural products.
Ultrasonic Assisted Cloud Point Extraction Combined with Liquid Chromatography-Tandem Mass Spectrometry for the Determination of Natural Vitamin K Homologues in Vegetables
LI Kailong, CHEN Tongqiang, XU Wenyang, LI Tao, WANG Liangliang, WANG Fang, SUN Guifang
2021, 42(8):  214-220.  doi:10.7506/spkx1002-6630-20200331-455
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In this study, an ultrasonic-assisted cloud point extraction combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS) with a triple quadrupole (QqQ) mass detector method was developed and optimized for determining the natural vitamin K homologues phylloquinone (PK) and menaquinone-4 (MK) in vegetables. The analytes were isolated from vegetable samples by ultrasound-assisted extraction using acetonitrile as the extraction solvent followed by microwave-assisted cloud point extraction with Triton X-45, chromatographed by gradient elution using ammonium formate solution-methanol as the mobile phase, detected using an electrospray ionization source in the positive ion mode (ESI+) with multiple reaction monitoring (MRM), and quantified by the external standard method. The results showed that the optimized conditions for ultrasonic-assisted cloud point extraction were as follows: 1.5 g of the sample was ultrasonically extracted with 2 mL of acetonitrile, and the extract was re-extracted with 50 μL of Triton X-45 after adding 0.15 g of sodium chloride to it. Good linearity was achieved for the two vitamin K homologues in the range of 1.0–500.0 ng/mL with correlation coefficients of greater than 0.997 0. The limits of detection (LODs) were 1.0 ng/g and 0.8 ng/g for PK and MK, respectively. The recoveries for samples spiked at three concentration levels of 40, 80 and 120 ng/g ranged from 94.5% to 106.2%, and the relative standard deviations (RSDs) of inter-day and intra-day precision were less than 10%. No natural vitamin K was detected in the root and stem vegetables. Only PK was detected in the leafy vegetable samples analyzed at levels in the range of 96.2–1 704.5 ng/g, with the highest value being observed in spinach.
Characterization of Microbial Community and Volatile Flavor Compounds of Deep Spoiled Panxian Dry-Cured Ham
MU Yu, SU Wei, MU Yingchun
2021, 42(8):  221-228.  doi:10.7506/spkx1002-6630-20191111-143
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This study aimed to establish an effective method for the prediction of the microbial spoilage of dry-cured ham. We compared the physicochemical characteristics of normal and spoiled Panxian hams. Meanwhile, we employed high-throughput sequencing and solid phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) to reveal the bacterial community structure and volatile flavor compounds of normal and spoiled hams, respectively. The results showed that low concentration of NaCl and high levels of water activity (aw), pH, and moisture were found in spoiled hams. Three members of the family Enterobacteriaceae, namely norank_Enterobacteriaceae, unclassified_Enterobacteriaceae, and Serratia, were determined as the predominant microorganisms in spoiled hams. Redundancy analysis (RDA) demonstrated that moisture and pH might be important factors influencing the two unknown strains of Enterobacteriaceae, and Serratia was closely related to NaCl content. The levels of ketones, hydrocarbons, benzene- and sulfur-containing volatile compounds in spoiled hams were higher than in normal hams. Linear discriminant analysis effect size (LEFSe) analysis indicated that phenol, phenylacetaldehyde, 1-pentadecene, dimethyl disulfide, and 3-methylbutyric acid were identified as characteristic volatile flavor compounds of spoiled hams. Network analysis between microorganisms and volatile flavor compounds illustrated that norank_Enterobacteriaceae and unclassified_Enterobacteriaceae were the main contributors to the volatile flavor compounds of spoiled hams. These findings provide a theoretical foundation for quality control of Panxian ham.
Analysis of Composition and Structures of Pigments in Roast Duck Skin by HPLC-MS/MS
HAO Min, XIA Qiang, HE Jun, SUN Yangying, DANG Yali, CAO Jinxuan, PAN Daodong
2021, 42(8):  229-234.  doi:10.7506/spkx1002-6630-20200405-052
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In order to explore the mechanism of color formation of roast duck, pigment was extracted ultrasonically and separated by high performance liquid chromatography-mass spectrometry (HPLC-MS/MS). In addition, the structure of the pigment was identified by ultraviolet visible (UV-VIS) spectroscopy and Fourier transform infrared spectroscopy (FT-IR). The results showed that the brightness value (L*) of roast duck skin decreased, and both the redness value (a*) and yellowness value (b*) increased with the increase of roasting time. The UV-VIS spectrum showed a maximum absorption peak at 210 nm. The FT-IR spectrum showed that the pigment contained at least six compounds with functional groups of –OH, C=O, –NH2, –CHO and high conjugated C=C , whose molecular masses were 365.11, 301.14, 277.22, 303.23, 353.27, and 279.23 respectively. These constituents included long-chain fatty acid esters, oligomers, redox reaction products, derivatives produced by lipid oxidation, amides and aldehydes.
Separation, Structural Identification and Anti-tumor Effects of New Compounds from Cordyceps militaris
XIANG Ting, XIA Chen, LIU Jianhua, WANG Chaoran, SHEN Jianfu
2021, 42(8):  235-242.  doi:10.7506/spkx1002-6630-20191017-168
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Preparative high performance liquid chromatography with C18 and C18-HCE columns was used to prepare nucleoside compounds from Cordyceps militaris including cordycepin, adenosine and two monomeric compounds, and their structures were identified by ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF MS) and nuclear magnetic resonance (NMR) spectroscopy. The inhibitory effects of cordycepin, compound I and compound II on the proliferation of HepG2 liver cancer cells were comparatively evaluated by methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay. The results showed that the C18-HCE column could effectively improve the retention and selectivity of polar compounds, facilitating the separation and purification of nucleosides. The purities of adenosine, cordycepin, compound I and compound II were 90.49%, 98.11%, 96.34%, and 98.33% and their yields were 0.053%, 0.253%, 0.368%, and 0.231%, respectively. Compound II was identified as a new substance, which was similar to compound I in structure, both being nucleoside-type compounds substituted with amino acids or amino acid derivatives. Anti-tumor activity experiments showed that all compounds could significantly inhibit the proliferation of HepG2 hepatoma cells, and their effects decreased in the order: compound I > cordycepin > compound II.
Safety Detection
Determination of Major and Trace Metal Elements in Beer by Microwave Plasma-Atomic Emission Spectroscopy
ZHANG Ping, LIU Hongwei, HUANG Jianhua, CHEN Lin
2021, 42(8):  243-247.  doi:10.7506/spkx1002-6630-20200509-106
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An analytical method was established for the determination of four major metal elements (Na, Mg, K, and Ca) and ten trace metal elements (Al, Mn, Fe, Ni, Cu, Zn, Sr, Cd, Ba, and Pb) in beer by microwave plasma-atomic emission spectroscopy (MP-AES). The samples were microwave digested using HNO3 in closed digestion vessels. The spectral interferences were corrected by selecting a suitable wavelength as well as using fast linear interference correction (FLIC). The matrix effect was corrected by using internal standard elements. Inductively coupled plasma-mass spectrometry (ICP-MS) was used to analyze the same sample solution to demonstrate the accuracy of the method. Under the optimized conditions, the limits of detection (LODs) for the analytes were in the range of 0.13–230 μg/L. The spiked recoveries ranged from 95.6% to 104% with relative standard deviations (RSDs) of 1.8%–4.0%. All the major and trace metal elements were successfully determined in beer samples by the MP-AES method and the results obtained were identical to those obtained by ICP-MS. The method has the characteristics of simple and fast operation, good stability, low running cost and good accuracy, and is suitable for high throughput analysis of multiple metal elements in beer.
Species Identification of Common Wild Edible Bolete in Yunnan by Fourier Transform Mid-infrared Spectroscopy Coupled with Support Vector Machine
HU Yiran, LI Jieqing, LIU Honggao, FAN Maopan, WANG Yuanzhong
2021, 42(8):  248-256.  doi:10.7506/spkx1002-6630-20191016-151
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In an effort to provide reference for the identification and quality control of edible fungi in Yunnan, Fourier transform mid-infrared spectroscopy combined with support vector machine (SVM) was used to identify different wild bolete species in Yunnan. The influence of different data mining methods on model classification performance was determined. Infrared spectra of 8 species with 827 samples of eight common wild bolete species were acquired and analyzed for spectral characteristics, and a discriminant model was established using SVM. Spectral information mining was performed by preprocessing, feature variable extraction or their combination, and the classification performance of the models developed was compared with each other to find the optimal method for species identification of wild bolete. The results showed that there was a large amount of noise and interference information in the original data, which reduced the classification performance of the model. All tested data mining methods could remove non-effective information but in different extents, improving model classification performance. Preprocessing combined with characteristic variables extraction had the highest ability to mine spectral information, providing the best model classification performance. The SVM model developed had excellent goodness of fit with high classification accuracy and wide applicability, allowing the accurate and quick identification of the eight wild bolete species.
Discrimination between Raw and Restructured Beef Steak Using Hyperspectral and Ultrasound Imaging
SUN Zongbao, Wang Tianzhen, Zou Xiaobo, LIU Yuan, Liang Liming, LI Junkui, Liu Xiaoyu
2021, 42(8):  257-263.  doi:10.7506/spkx1002-6630-20200414-187
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A novel method to discriminate between raw and restructured beef steal was developed by using hyperspectral and ultrasonic imaging. Hyperspectral and ultrasonic images of samples were collected from which texture feature values were extracted by gray level co-occurrence matrix (GLCM). Four discriminant models were established separately using linear discriminant analysis, K-nearest neighbor (KNN), back propagation artificial neural network, and extreme learning machine (ELM) based on the hyperspectral image data, the ultrasonic image data or their fusion. We comparatively evaluated three variable selection techniques: successive projections algorithm (SPA), competitive adaptive reweighted sampling (CARS), and variables combination population analysis (VCPA). The results suggested that the texture of restructured beef steak was uniform, and the ultrasonic image signal was weak and uniform, which was distinct from that of raw beef steak. The best models for hyperspectral and ultrasound imaging were KNN and ELM, with prediction set identification rates of 95.00% and 90.00%, respectively. After data fusion, the prediction set identification rate of the best model ELM was 97.50%. The prediction set identification rates of the models established based on the texture variables selected by CARS and VCPA were 100.00%. The results showed that data fusion between hyperspectral and ultrasonic imaging combined with a suitable variable selection method can discriminate between raw and restructured beef steak quickly and accurately.
Antimicrobial Resistance, Virulence Gene and Genetic Characteristics of Vibrio parahaemolyticus Isolates from Aquatic Products Imported to Shanghai
SHEN Jinling, ZHAO Lina, HAN Wei, XU Zhenjian, JIANG Yuan, YANG Jielin, GUO Dehua, XUE Feng
2021, 42(8):  264-269.  doi:10.7506/spkx1002-6630-20200318-276
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This work aimed to study the antimicrobial resistance, key virulence genes and genetic evolution of Vibrio parahaemolyticus isolated from aquatic products imported to Shanghai for the purpose of providing supportive data for the early warning of V. parahaemolyticus contamination risks and tracking its sources. A total of 68 isolates were obtained from aquatic products imported to Shanghai in the years between 2017 and 2019 and analyzed for antimicrobial susceptibility, key virulence genes and multi-locus sequence typing. These isolates showed the highest resistance rate to ampicillin (98.53%), whereas their resistance rates to other antimicrobials tested were relatively lower, and all the isolates were sensitive to levofloxacin and chloramphenicol. Intermediate resistance was observed for nine antimicrobials, among which, the intermediate resistance rates to aminoglycosides, piperacillin, ciprofloxacin and cefoxitin were relatively higher. In total, 20.59% of the isolates were resistant to two or more antimicrobials. One isolate was found to harbor trh, and no isolate harbored tdh. A total of 65 sequence types (STs) were identified, of which, 52 were novel STs. No apparent correlation was found between geographical origin, food type and ST type; however, isolates of the same specific STs were identified in the same type of food samples imported from the same country at different times. In conclusion, the V. parahaemolyticus isolates were highly genetically diverse and showed intermediate resistance and multiple resistance to drugs, and some of them were found to have high pathogenic potential. Thus, much attention should be paid to this issue.
Simultaneous Detection of Three Major Milk Allergens in Food by UPLC-MS/MS
NING Yawei, LIU Zhuo, YANG Zheng, MA Junmei, FAN Sufang, LI Qiang, ZHANG Yan
2021, 42(8):  270-276.  doi:10.7506/spkx1002-6630-20200419-248
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An ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed to simultaneously detect the three major milk allergens β-lactoglobulin, αs1-casein and αs2-casein in food, which could avoid false negatives caused by factors such as the degradation of single allergens during processing and low sensitivity. The standard protein hydrolysates were analyzed by nanoflow liquid chromatography-tandem orbitrap high resolution mass spectrometry, obtaining eight signature peptides. Wheat flour was selected as the blank matrix for methodological validation in the multi-reaction monitoring (MRM) mode by triple quadrupole mass spectrometry. This method showed a good linear relationship in the concentration range of 1.6–30 000 ng/mL with correlation coefficient (R2) > 0.999. The limits of quantitation (LODs) of β-lactoglobulin, αs1-casein, and αs2-casein were 50, 3.2, and 40 μg/g, respectively. The average recoveries were between 86.41% and 98.60% with relative standard deviations (RSDs) ≤ 8.95%, and the matrix effects were 86.04%–97.48%. This method could be applied to actual products with and without milk in order to identify whether there is any inconsistence between the allergen label and the actual allergens in food, and more broadly to reduce the potential health risks for milk-sensitive consumers.
Liquid Phase Microextraction with Deep Eutectic Solvent Combined with High Performance Liquid Chromatography for Determination of New Neonicotinoid Insecticide Residues in Edible Oil
WANG Suli, GUO Zhenfu, GENG Lili
2021, 42(8):  277-282.  doi:10.7506/spkx1002-6630-20200423-294
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In this study, a method for the determination of residues of four new neonicotinoid insecticides (thiamethoxam, imidacloprid, acetamiprid, and thiacloprid) in edible oil was developed using ultrasound-assisted liquid-liquid microextraction with deep eutectic solvent (UA-DES-LLME) followed by high performance liquid chromatography (HPLC). Samples were diluted with n-hexane and added with deep eutectic solvent (DES) before being subjected to ultrasonic-assisted dispersive liquid-liquid microextraction (UALLME). Then, the extract was centrifuged. The lower DES-rich phase was collected and injected into the HPLC system for analysis. Several important parameters influencing the extraction efficiency, such as the type and volume of DES, ultrasonication time, and centrifugation time, were investigated. Under the optimized conditions, the recoveries of the analytes were between 81.9% and 98.0%, with relative standard deviations (RSD, n = 5) of 5.5%–8.3%. The limits of detection (LODs) and limits of quantitation (LOQs) were 3.2?5.3 μg/L and 10.8?17.7 μg/L respectively. The method was successfully applied to real samples of soybean oil, sunflower seed oil and linseed oil. This method combined extraction and concentration in one step without the use of poisonous organic solvents. This method proved to be simple, rapid and efficient.
Determination and Quantitation of Ethyl Carbamate in Chinese Baijiu by Ultra-high Performance Liquid Chromatography-High Resolution Mass Spectrometry
XIONG Xiaotong, HU Feng, YOU Xiaolong, YIN Yanyan, CHEN Mingxue, CHENG Pingyan, ZHONG Fangda
2021, 42(8):  283-287.  doi:10.7506/spkx1002-6630-20191030-333
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A new method for the quantitation of ethyl carbamate (EC) in Maotai-flavor Baijiu was developed using ultra-high performance liquid chromatography-high resolution mass spectrometry with primary daughter ions. Based on the fact that it has the special property of in-source fragmentation, EC was detected under the full scan model. The primary parent ion with m/z 90.05 was used as qualitative ion and its daughter ion with m/z 62.02 as both qualitative and quantitative ions considering that a great noise and low response values were achieved respectively when using the primary parent ion and the secondary daughter ion as quantitative ions. The calibration curve showed good linearity in the range of 12.2–244 μg/L with a correlation coefficient greater than 0.999. The limit of detection (LOD) was 0.95 μg/L. The recoveries of EC from spiked samples were 83.46%–106.79%, with relative standard deviations (RSD) of 0.67%–3.54%. Overall, the method was suitable for the quantitation of EC in Maotai-flavor Baijiu with the advantages of rapidity, low detection limit and high accuracy.
Quantitative Detection of Cow Milk Adulteration in Goat Milk Based on a Non-Protein Biomarker Present in Cow Milk
SUN Jinzhi, FU Lujing, YUE Tianli, LI Jingyan, GUO Chunfeng
2021, 42(8):  288-293.  doi:10.7506/spkx1002-6630-20200402-028
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Enzyme-linked immunosorbent assay cannot detect the adulteration of heat-treated cow milk in goat milk. In this study, N-acetylglucosamine was revealed by gas chromatography-mass spectrometry to be a non-protein biomarker present in cow milk that discriminates it from goat milk, and a high performance liquid chromatographic method was developed for quantitative detection of cow milk adulteration in goat milk. Samples were derivatized with 1-phenyl-3-methyl-5-pyrazolone at 70 ℃ for 60 min, followed by chromatographic separation of the marker on a reversed-phase C18 column and detection at 245 nm. The amount of cow milk added to goat milk was calculated according to the N-acetylglucosamine content in the samples. The limit of detection and limit of quantification of the method were 0.3% and 1.0%, respectively. The method showed good linearity in the range of 1%–100% cow milk addition with correlation coefficient of 0.999 4. The recoveries of spiked samples were 100.4%–105.1%, and the intra-day and inter-day precisions (relative standard deviations) at spiked concentrations of 10, 50 and 100 mg/L were 1.8% and 2.0%, 1.7% and 3.7%, 2.6% and 2.7%, respectively. This method proved to be highly sensitive, accurate and precise, and therefore could be suitable for quantitative analysis of cow milk adulteration in goat milk.
Determination of Advanced Glycation End Products in Salmon and Exploration of Characteristic Indicators to Identify Its Geographical Origin
XU Zhenghua, WU Jiali, ZHU Kewei, LIANG Yushen, YANG Xuejiao, ZHAO Yong, LING Jing, ZHENG Siheng, ZHOU Henggang, ZHANG Kun, ZHU Yuzhen, ZENG Maomao
2021, 42(8):  294-298.  doi:10.7506/spkx1002-6630-20200418-235
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A method for simultaneous determination of carboxymethyllysine (N-carboxymethyllysine, CML) and carboxyethyllysine (N-carboxyethyllysine, CEL) in salmon by ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was developed in this paper. The chromatographic separation was performed on an ACQUITY UPLC HSS T3 column using a mobile phase composed of 100% acetonitrile and 0.1% formic acid, and the separated analytes were detected by electrospray ionization in the positive ion (ESI+) mode under multiple reaction monitoring (MRM) and quantitated by the isotope internal standard dilution method. Samples were degreased, reduced, and hydrolyzed and the hydrolysate was purified by solid phase extraction on an Oasis MCX column before being injected into the instrument. The average recoveries of the method were 90%–95%, the precision expressed as relative standard deviation (RSD) was less than 5%, the limit of detection (LOD) was 0.032 7 ng/g, and the limit of quantitation (LOQ) was 0.142 2 ng/g. The method was used to detect 21 salmon samples from Norway, Chile and the Faroe Islands, and the obtained data were analyzed by principal component analysis (PCA). The results showed that there were significant differences in CML and CEL contents between different countries and individuals of the same species. This study revealed CEL to be a potential characteristic indicator to distinguish Norway salmon from those from Chilean and the Faroe Islands.
Geographical Origin Traceability of Green Tea from Fanjing Mountain Area, Tongren, Guizhou
ZHANG Minglu, HUANG Congwei, LI Like, XIAO Yixuan, YU Haiyou, ZHAO Tairan, YIN Jie
2021, 42(8):  299-304.  doi:10.7506/spkx1002-6630-20200401-003
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The contents of 31 trace elements in 42 green tea samples collected from four tea-producing counties in Tongren city, Guizhou province were determined by inductively coupled plasma mass spectrometry (ICP-MS). The correlation between these elements was analyzed. Principal components analysis (PCA), stepwise linear discriminant analysis (S-LDA) and orthogonal partial least squares discriminant analysis (OPLS-DA) were used to develop classification models for the tea samples from different geographic origins. The results indicated that the contents of Co, Mo, Ag, Cd, Zn, Al, Y, Nd, Sc, Pr, Gd, Sm, Dy, Er, Yb, Ho, Tb, Tm and Lu in tea samples were significantly different among the different producing areas (P < 0.05). There was a synergistic correlation between some of the elements, while others showed an antagonistic correlation between each other. The OPLS-DA model displayed the best classification performance with a correct discrimination rate of 92.86%. This study revealed that the multivariate statistical classification model based on trace elements contents could effectively distinguish tea samples from neighboring producing areas, which will be useful for tea quality and safety traceability in the tea industry.
Aptamer Combined with Quantum Dot for Simultaneous Detection of Staphylococcus aureus and Escherichia coli O157:H7
WANG Yue, SHAO Lin, LI Qianxue, YI Le, QU Han, WANG Hongli, SHEN Minghao
2021, 42(8):  305-312.  doi:10.7506/spkx1002-6630-20190927-336
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Objective: To establish a rapid, efficient and sensitive method for the simultaneous detection of the foodborne pathogens Staphylococcus aureus and Escherichia coli by the combined use of aptamer and quantum dot. Methods: Surface-enhanced Raman spectroscopy was used to select two aptamers with good binding specificity and affinity to the target bacteria. The aptamers were coupled to immunomagnetic beads to specifically capture the target bacteria. Finally, two kinds of quantum dots giving rise to luminescence in different manners were selected and used in combination with quantum dot fluorescent labeling technology to construct a “sandwich structure” of “quantum dot + aptamer + target bacteria”. The structure was optimized and the limits of detection (LODs) for the two target bacteria were determined. Results: The selected aptamers allowed the simultaneous detection of the target bacteria. The optimal capture conditions were as follows: immunocapture time 45 min and magnetic separation time 2 min. The optimum concentration of aptamer was 400 nmol/L. The LOD for S. aureus was 101 CFU/mL, and the linear relationship between fluorescence intensity (Y) and logarithmic bacterial count (X) was fitted as Y = 28.51X + 126.67, with a correlation coefficient, R2, of 0.973. The LOD for E. coli was 102 CFU/mL, and the linear relationship between was Y and X was fitted as follows: Y = 92.86X ? 64.67, with R2 of 0.987. The recoveries of S. aureus and E. coli in spiked milk samples were 94.6%–102.8% and 93.4%–100.4%, respectively. Conclusion: The method established in this study is a rapid, efficient, sensitive and easy to operate method for the simultaneous detection of the two foodborne pathogens. It is promising for application in food processing and safety detection.
Fluorescent Microspheres Competitive Immunochromatography Combined with Naked Eye Reading for Rapid and Semi-quantitative Detection of Sibutramine in Food Supplements
TAN Guiliang, PAN Ziqiang, ZHANG Shiwei, HU Min, FENG Ronghu, YANG Maoxun
2021, 42(8):  313-318.  doi:10.7506/spkx1002-6630-20200409-129
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In this study, a novel semi-quantitative method to determine sibutramine in food supplements was developed using fluorescent microspheres competitive immunochromatography coupled with naked eye reading. In the method, four antigen-coated test lines were set on the test strip, allowing decreasing concentration gradients of the labelled antibody left after reaction with the tested antigen to react with the coating antigen, so that immune reaction could be achieved with multi-scale sensitivity. The excitation light source was electric torch at 470 nm. The concentrations of Tween-20, labeled antibody and coating antigen, nitrocellulose (NC) membrane type and reaction time were optimized. Semi-quantitative analysis was performed by counting the number of colored test lines. Under the optimized reaction conditions, the sensitivity of the method was 0.05 μg/mL and the detection ranged from 0 to 9.00 μg/mL. This method took 15 minutes without the need for any special instrument, giving results consistent with those of high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). The newly developed method was a simple, reliable and rapid method, and could be used in the rapid screening of sibutramine in food supplements.