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Table of Content

25 July 2017, Volume 38 Issue 14
Bioengineering
Construction of Genetically Engineered Strain for Nattokinase Production and Enzyme Activity Analysis
CUI Qing, QIAN Bingjun, YAO Xiaomin, JI Shunli, LU Feifeng, WU Jing, ZHANG Jianhua
2017, 38(14):  1-8.  doi:10.7506/spkx1002-6630-201714001
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Nattokinase (NK), encoded by the aprN gene of Bacillus subtilis natto, has strong fibrinolytic activity both in vitro and in vivo. In this research, the aprN gene from B. subtilis was cloned and the codons which encode the first 30 amino acids were optimized on the basis of the codon preference of B. subtilis. Recombinant vector pHT01-aprN was constructed. Through restriction enzyme digestion analysis, PCR amplification and sequencing, the subcloned gene was confirmed to be aprN. The pHT01-aprN was transformed into B. subtilis 168 by electroporation, and the engineered bacterium (B.s 168/pHT01-aprN) was isolated on LB plates containing chloramphenicol. NK expression was induced by IPTG, and the highest enzyme activity in shaking flask culture was up to (289.00 ± 3.42) U/mL with good stability, which was 3.9 times as high as that of wild-type B. subtilis natto.
Cloning and Bioinformatics Analysis of Acidophilic xynA Gene from Penicillium janthinellum
HOU Jie, LI Qin, Xiong Ke, XU Youqiang, LI Xiuting,
2017, 38(14):  9-16.  doi:10.7506/spkx1002-6630-201714002
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Acidic xylanases have extensive application in feed and wine industries. The whole sequence of the gene xynA encoding acidic xylanase was amplified from Penicillium janthinellum MA21601 by genome walking. A cDNA sequence was obtained through the elimination of introns by overlapping PCR and analyzed by bioinformatics. The whole sequence was about 720 bp in length with only one intron of 63 bp. The cDNA sequence was 657 bp long and putatively encoded a protein which contained a 28-amino acid (aa) signal peptide and a 190-aa mature peptide. The molecular weight of the protein was estimated to be about 20.61 kD, which had an isoelectric point of 7.0. Bioinformatics analysis showed that XynA was a hydrophilic protein without disulfide bond. The amino acid sequence comparison of XynA with other fungal GH11 acidophilic xylanases indicated that the XynA had an identified specific recognition site of Asp, which displayed a β-jelly-roll architecture as a conserved region which was the characteristic of the GH11 family xylanases. The recombinant xylanase was successfully expressed in Escherichia coli with a specific activity of up to 220.5 U/mg.
Metaproteomic Analysis of Traditional Fermented Soybean Paste in Northeast China
WU Rina, XUE Yating, ZHANG Ping, TANG Xiaoyang, TAO Dongbing, YUE Yuanyuan, ZHANG Pengfei, CHEN Wei
2017, 38(14):  17-23.  doi:10.7506/spkx1002-6630-201714003
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In order to under the function of the microbial community in fermented soybean paste, we established a metaproteomic expression profile by stepwise extraction and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The proteins extracted were identified by liquid chromatography tandem mass spectrometry-mass spectrometry (LC-MS/MS). Results obtained were as follows: the number of proteins identified in fermented soybean paste was 232, including 60 proteins participating in sugar metabolism, 57 proteins involved in nucleic acid metabolism, 48 proteins involved in metabolism, 15 proteins related to energy metabolism, 3 proteins related to lipid metabolism and 49 other functional proteins. The number of proteins derived from bacteria was almost three times as large as the number of proteins derived from fungi. Bacterial proteins were mainly derived from Bacillus subtilis, Leuconostoc, Fecal streptococcus, Leuconostoc mesenteroides and Deshi Lactobacillus, while fungal proteins mainly came from Saccharomyces cerevisiae, Fission yeast, Alternaria, white rot fungi and Ashbya gossypii.
Screening of Phenyllactic Acid-Resistant Escherichia coli and Its Application in the Synthesis of Phenyllactic Acid
XU Yan, GUO Qian, ZHU Yibo, WANG Limei, QI Bin
2017, 38(14):  24-29.  doi:10.7506/spkx1002-6630-201714004
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Screening phenyllactic acid (PLA)-resistant strains enables effective reduction of the inhibitory effect of PLA on the synthesis process and thus an increase in PLA production. In this report, Escherichia coli BL21 (DE3) was mutagenized by UV irradiation and screened for a mutant resistant to PLA, E. coli Z2016 (CCTCC Accession No. M2016332). E. coli Z2016 was used the host strain to construct recombinant strains E. coli Z2016 pET-28a-ldhY52V and E. coli Z2016 pET-28a-ldhL for the synthesis of D- and L-PLA. The results showed that E. coli Z2016 was able to grow normally in a medium containing 1 g/L D-PLA. The production of D- and L-PLA by the recombinant strains were 6.75 and 6.97 g/(L·h), which were increased by 14.02% and 8.95% as compared to those produced by the original strain, respectively. During 120 min of fed-batch fermentation, the production of D-PLA by E. coli Z2016 pET-28a-ldhY52V was 20.02 g/L with a conversion rate of 90.97%, which was increased by 22.17% as compared to that produced by the control. The L-PLA concentration produced by E. coli Z2016 pET-28a-ldhL was 20.87 g/L, which was increased by 16.85% as compared to produced by the control, with a conversion rate of 91.24%. Accordingly, the use of PLA-resistant strains provided an effective method to increase PLA production.
Diversity and Dynamic Changes of the Bacterial Community during Fermentation of Soybean Paste
ZHANG Ying, WU Rina, SUN Huijun, WU Junrui, TAO Dongbing, WANG Hongyu
2017, 38(14):  30-35.  doi:10.7506/spkx1002-6630-201714005
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In this study, the diversity and dynamic changes of?the bacterial community during the natural fermentation of soybean paste, a traditional fermented food popular in northeast China, were analyzedusing polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) technology and high-throughput sequencing. Results showed that bacterial operational taxonomy unit (OTU) values were fluctuant during the fermentation and reached a maximum on day 21, indicating the largest bacterial population. Analysis of DGGE profiles and the distribution of bacteria at the genus level suggested that Leuconostoc sp., Enterococcus sp., Tetragenococcus sp. and Lactobacillus sp. were predominant bacterial strains for different fermentation stages of soybean paste. Among these, Tetragenococcus sp. was more fluctuant and and vulnerable to both internal and external environments. Enterococcus faecium, Leuconostoc sp. and Weissella viridescens decreased with fermentation time indicating that these bacteria might be mainly involved in the production of catabolic enzymes in the early fermentation stage and played a role in the adjustment of the internal environment during the whole fermentation process.
Effect of Main Metabolites on Carotenoids Degradation during the Fermentation of Chinese Wolfberry Wine
LIU Ya, LIU Jianhua, ZHANG Huiling, QI Xiaoqin, LI Jinpeng, FU Lixia, WANG Xiaochang
2017, 38(14):  36-41.  doi:10.7506/spkx1002-6630-201714006
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In this paper, Chinese wolfberries were inoculated with yeast and fermented to produce a wine. High performance liquid chromatography (HPLC) and gas chromatography (GC) were used to analyze the contents of the main metabolites alcohols, esters, carboxylic acids, aldehydes and ketones as well as changes in carotenoid content before and after fermentation. On the basis of the obtained data, we established a model wine system to elucidate the effects of various metabolites on carotenoid content and further to determine the most important factor affecting carotenoid degradation. The results showed that carboxylic acids were the main factors affecting the degradation of carotenoids, followed by aldehydes and ketone. Esters and alcohols had a minimal impact.
Screening of an Aroma-Producing Fungal Strain and Its Synergistic Effect with Rhizopus oryzae on Rice Wine Fermentation
ZHAO Tingting, LU Qianwen, SONG Feifei, XU Gurong, CAI Ting, XIANG Wenliang, ZHU Jianqing
2017, 38(14):  42-48.  doi:10.7506/spkx1002-6630-201714007
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A fungal strain, ZT018, isolated from a traditional rice wine starter culture for its potential to produce aroma compounds was identified as Mucor indicus based on its 18S rRNA sequence and morphology. When cooperated with Rhizopus oryzae to ferment rice wine, the fungal isolate soon became the dominant genus, and its cellular morphology was also gradually transformed from a filamentous form to a yeast-like one which benefited the production of aroma compounds. In the rice wine fermented by a mixture of ZT018 and R. oryzae (MIX), the volatile aroma components were analyzed by headspace solid-phase micro-extraction coupled with gas chromatography-mass spectrometry (HS-SPME-GC-MS). The results showed that among the primary volatile aroma compounds of MIX, the amounts of 2-phenethyl acetate, ethyl hexadecanoate, isoamyl alcohol and 2-phenethyl alcohol were significantly improved when compared with the one fermented by R. oryzae alone (SIM). Moreover, other volatile aroma compounds that could coordinate well with each other to improve the aroma, such as isobutanol, propionic?acid and limomene, were also found in the MIX. The MIX fermented for 5–7 days exhibited the best sensory quality, and it had 2-phenethyl acetate, ethyl hexadecanoate, isoamyl alcohol and 2-phenethyl alcohol contents of 5.270–5.750, 2.270–4.877, 4.375–8.419 and 39.983–63.046 mg/L, respectively, which were increased by 30.03%–38.06%, 46.83%–15.68%, 28.45%–23.81% and 55.60%–22.91%, respectively, as compared with those in the SIM. Thus, the MIX had better quality than SIM, which will be of great significance to improve rice wine aroma.
Optimization of Preparation Process and Properties of Microcapsules Containing Lactobacillus helveticus MG9-2
SONG Jiaojiao, BAO Qiuhua, WANG Yali, SHANG Yina, HUO Qiwen, ZHANG Xiaoning, CHEN Jing, LI Minghui, WANG Junguo
2017, 38(14):  49-57.  doi:10.7506/spkx1002-6630-201714008
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Lactobacillus helveticus MG9-2 was microencapsulated with milk protein as wall material by means of emulsification/internal gelation. The preparation process was optimized, and the effect of different calcium salts (CaCO3 and CaCl2) as crosslinking?agents on microencapsulation efficiency was investigated. The results showed that the optimal process parameters were as follows: stirring speed, 800 r/min; water-to-oil ratio, 1:10; concentration of skim milk, 0.35 g/mL; and concentration of calcium salt, 1.5 mol/L. Under these conditions, the microencapsulation efficiency (ME) of viable cells with CaCO3 and CaCl2 were (78.4 ± 3.6)% and (75.7 ± 4.9)%, respectively, with no significant difference being observed between them. Microcapsules prepared with CaCO3 were smaller in size than those prepared with CaCO3, exhibiting a more compact surface, and a more regular spherical shape and a narrower size distribution and displaying a better resistance to intestinal release and gastric acid. The findings demonstrated that microcapsules with improved properties could be prepared more simply using CaCO3 as crosslinking?agent than CaCl2.
Thermal Inactivation Properties of Salmonella paratyphi A in Different Media
ZHANG Peipei, NIU Huimin, LI Miaoyun, ZHAO Gaiming, CUI Wenming, SUN Lingxia
2017, 38(14):  58-63.  doi:10.7506/spkx1002-6630-201714009
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This study aimed to study the thermal inactivation profile of Salmonella paratyphi A in different media. Salmonella paratyphi A was inoculated at a concentration of 108 CFU/g into different media, and then subjected to thermal treatments at different temperatures. The surviving cells were counted on selective media. DoseResp model was used to fit the thermal inactivation curve by Origin 8.0 software. The decimal reduction time (D) values were calculated, which represented the heat resistance of Salmonella paratyphi A. The results showed that thermal inactivation curves were well fitted to the DoseResp with a correlation coefficient (R2) > 0.97. The D values of Salmonella paratyphi A in different media were different. The D value in sausage high in fat and protein with a lean meat-to-fat ratio of 1:1 (m/m) was greater, indicating that that it was relatively heat resistant. Salmonella paratyphi A could be killed by thermal treated at 55 ℃ for 25.0 min, 63 ℃ for 2.03 min, or 72 ℃ for 0.31 min in common meat products.
Effect of Nitrate Pulse-Feeding Cultivation on Cell Growth and Cell Composition of Thermo-Tolerant Desmodesmus sp. F51
XIE Youping, ZHAO Xurui, YANG Xuqiu, LU Yinghua, ZHENG Xiangnan, CHEN Jianfeng
2017, 38(14):  64-70.  doi:10.7506/spkx1002-6630-201714010
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The aim of the present study was to investigate the effect of nitrate pulse feeding on the cell growth and cell composition of Desmodesmus sp. F51. Under nitrate pulse-feeding cultivation, the protein content of cells of Desmodesmus sp. F51 decreased from (560 ± 16) mg/g to (456 ± 17) mg/g, while the contents of carbohydrate and lipid increased from (209 ± 13) mg/g to (310 ± 12) mg/g and (98 ± 3) mg/g to (120 ± 4) mg/g, respectively. By investigating the changes in carotenoid, lipid and carbohydrate compositions, it appeared that nitrate pulse feeding could promote β-carotene biosynthesis and enhance the bioconversion of α-carotene to lutein. Lutein accumulation was positively associated with polyunsaturated fatty acid formation. Desmodesmus sp. F51 tended to accumulate carbohydrate as an energy storage product after 3 days of cultivation under nitrate pulse-feeding conditions, and the major accumulated monosaccharide was glucose (75.7 ± 1.4)%. Therefore, the nitrate-pulse feeding strategy is a highly promising approach to simultaneously produce lutein and carbohydrate in Desmodesmus sp. F51.
Surface Modification of Polypropylene Fiber Membranes with Dopamine to Enhance the Formation of Lactobacillus paracasei Biofilms and Its Fermentation Performance for Lactic Acid Production
ZHAO Zishu, LI Jinian, GUO Qian, YE Xiangyu, HU Mengxin,
2017, 38(14):  71-77.  doi:10.7506/spkx1002-6630-201714011
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Since dopamine can self-polymerize and adhere to the material surface in mild aqueous environments, dopamine was used for surface modification of polypropylene fiber membranes. In this paper, the effect of surface modification of polypropylene fiber membranes with dopamine on the formation of Lactobacillus paracasei biofilms was studied. The L. paracasei biofilms were characterized by scanning electron microscope, attenuated total reflectance Fourier transform infrared spectroscopy, and water contact angle measurement. The stress resistance and fermentation performance for lactic acid production were studied in detail. The results demonstrated that the dopamine layer on the surface of polypropylene fiber membranes remarkably enhanced the formation of L. paracasei biofilms. It is worth to note that as compared with the L. paracasei on polypropylene fiber membranes, L. paracasei biofilms on dopamine-modified polypropylene fiber membranes possessed higher resistance to severe environments, such as high temperature, strong acid and strong base. Besides, the purity of L-lactic acid produced by L. paracasei biofilms in a biofilm reactor was higher than 99%. The lag phase of fermentation scarcely existed, which means that high fermentation efficiency.
Effects of Lactobacillus plantarum Fermentation on the Structure of Soybean Meal Proteins
ZHANG Lili, CUI Xian, ZHANG Gongsheng, JIANG Fan, LIU Rongxu, HAN Jianchun,
2017, 38(14):  78-83.  doi:10.7506/spkx1002-6630-201714012
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In the present study, the structural changes of soybean proteins in aqueous suspension of skimmed soybean meal after being fermented by Lactobacillus plantarum were studied. With the extension of fermentation time, the degree of hydrolysis and in vitro digestibility of fermented samples were increased, and the particle sizes of soluble proteins became smaller. These results indicated that soybean proteins were degraded to a certain degree after fermentation. Simultaneously, the molecular structure of proteins in fermented samples was further unfolded, resulting in an increase in beta-sheet content and a decrease in alpha-helix content, as compared to the original soybean proteins and those sterilized at 0 h of fermentation. A red shift in the wavelength of maximum fluorescence emission was observed for the fermented soybean proteins. More sulfhydryl groups were exposed after fermentation. In addition, scanning electron microscopy (SEM) analysis showed that the surface of fermented soybean proteins was not as smooth as that of the original ones, with corrosion pores appearing.
Antimicrobial Activity of Lysozyme-Functionalized Gold Nanoparticles
YANG Longping, YAN Wenjing, HUANG Mingming, QIAO Weiwei, ZHANG Jianhao
2017, 38(14):  84-90.  doi:10.7506/spkx1002-6630-201714013
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In order to improve the stability of lysozyme and its antibacterial activity against Gram-negative bacteria, as a green and highly efficient antibacterial agent, lysozyme-functionalized gold nanoparticles were prepared for the first time by surface modification of gold nanoparticles with lysozyme. The influences of the mass ratio of lysozyme to gold nanoparticles and the concentration of functionalized nanoparticles on the antibacterial activity against Gram-negative and Gram-positive bacteria were examined. The antibacterial mechanism of the prepared materials against Escherichia coli and the cytotoxicity on SH-SY5Y cells were also investigated. The results showed that compared with pure gold nanoparticles and lysozyme, the antibacterial activity of lysozyme-functionalized gold nanoparticles against E. coli and Staphylococcus aureus were obviously enhanced, which indicated the synergistic antibacterial effect of lysozyme and gold nanoparticles. Under optimized conditions, 0.1 g/L lysozyme-functionalized gold nanoparticles were able to completely kill the two types of bacteria. Besides, lysozyme-functionalized gold nanoparticles exhibited durable antibacterial properties and good biocompatibility, which were not toxic to mammalian cells. Transmission electron microscopy (TEM) images and live/dead bacterial assay showed that lysozyme-functionalized gold nanoparticles could kill bacteria by disrupting the cell membrane and cell wall.
Optimization of Extraction and Structural Analysis of Polysaccharide from Paecilomyces hepiali Chen & Dai
ZHANG Lili, FAN Linlin, NIE Qixing, HUANG Yansheng, ZHANG Quancai, YIN Junyi, ZHANG Shuang, NIE Shaoping
2017, 38(14):  91-96.  doi:10.7506/spkx1002-6630-201714014
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Response surface methodology was applied to optimize polysaccharide extraction from Paecilomyces hepiali Chen & Dai. The extracted polysaccharide was structurally elucidated. The optimal extraction conditions were found to be 4.5 h extraction with hot water at 98.0 ℃ at water/material ratio of 25:1 (mL/g). Under these conditions, the maximum polysaccharide yield of 9.69% was obtained and the polysaccharide mainly composed of neutral polysaccharide (84.7%) and uronic acid (9.5%). High performance size exclusion chromatography (HPESC), high performance anion exchange chromatography and FT-IR spectroscopy were used to analyze the structural characteristics of the polysaccharide. Results showed that the molecular weight of the polysaccharide was 61.6 kD, displaying a single chromatographic peak, and its structure contained a pyran ring. In addition, the polysaccharide was composed of arabinose, galactose, glucose, xylose, mannose and galacturonic acid, with a molar ratio of 2.5:31:36:1:15:4. The findings may be useful for future studies on the structure-activity relationship of polysaccharides from Paecilomyces hepiali Chen & Dai and for the development of new polysaccharide-based products.
Optimization of Enrichment Medium and High Cell Density Cultivation of Lactobacillus reuteri IMAU10240
YAO Guoqiang, ZHANG Xuemei, GAO Zhimin, ZHAO Yanxia, SUN Tiansong, ZHANG Heping
2017, 38(14):  97-105.  doi:10.7506/spkx1002-6630-201714015
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Lactobacillus reuteri IMAU10240 is a potential probiotic lactic acid bacterium (LAB). This study aimed to achieve high viable counts of L. reuteri IMAU10240 by optimization of the composition of deMan Rogosa Sharpe (MRS) medium and culture conditions for industrialization application of this strain. The optimization was done using one-factor-at-a-time method, orthogonal array design and response surface methodology. The optimal medium composition was determined as follows (g/L): sucrose 100.00, soy peptone 13.25, yeast extract powder 8.84, yeast peptone 13.25, Na2HPO4 19.85, citric acid 2.58, MgSO4·7H2O 0.40, MnSO4·5H2O 0.12, glycerol 2.76, Tween-80 1.00, and L-cysteine hydrochloride 0.50, and the optimal culture conditions were 7–8 h culture at 37 ℃ and constant pH 5.5 under nitrogen atmosphere. In small-scale and pilot-scale experiments, the optimized conditions yielded a viable cell count of 7.57 × 109 CFU/mL, which increased to 2.59 × 1011 CFU/g after lyophilization. The enrichment medium and culture conditions can?be further verified in industrial production.
Effect of Fermentation on Rheology of Apple Pomace Polysaccharides
JIA Feng, LIU Dong, GUO Yurong, LI Jie, SUN Jiaojiao, SU Fan
2017, 38(14):  106-111.  doi:10.7506/spkx1002-6630-201714016
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The influences of concentration, temperature and storage time on the apparent viscosity and shear stress of apple pomace polysaccharide (APP), cider apple pomace polysaccharide (CAPP) and cider vinegar apple pomace polysaccharide (CVAPP) were studied in this research in order to elucidate the effect of fermentation on the rheology of APP. The results showed that APP, CAPP, and CVAPP were all pseudoplastic fluid, displaying a shear thinning behavior. Both CAPP and CVAPP exhibited significantly lower viscosity with decreased concentration dependence and enhanced temperature resistance as well as resistance within a period of time. The rheology of all three polysaccharides changed little during 3 days of storage. Both CAPP and CVAPP were superior to APP in term of processing characteristics to a certain extend.
Immobilization of Cross-Linked Amylase Aggregates on Polymer Containing Phenanthrene Skeleton
LI Kechun, LU Jianfang,, ZHOU Juying, XU Haitang, ZHAO Yanzhi,
2017, 38(14):  112-119.  doi:10.7506/spkx1002-6630-201714017
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Amylase was precipitated with 80% isopropanol followedcross-linking with glutaraldehyde to obtain cross-linked amylase aggregates (CLEAs-A). Then CLEAs-A was covalently immobilized on a macroporous polymer carrier containing a phenanthrene skeleton to obtain immobilized CLEAs-A. Herein, various process parameters were systematically evaluated. Moreover, the enzymatic properties and physical structure of immobilized CLEAs-A were investigated. The thermal and storage stability were improved remarkably as compared with those of the free amylase. After seventh repeated use, the activity recovery of immobilized CLEAs-A was still as high as 63.29%. Furthermore, scanning electron microscopy and porosity measurements indicated the surface and pore structure of immobilized CLEAs-A. The proposed immobilization strategy would provide a general approach for preparing immobilized enzymes with robust and high bio-catalytic properties for in industrial production.
Bacterial Diversity during Natural Fermentation of Soybean Paste under Variable Temperature Conditions
ZHOU Hongli, LI Sha, ZHANG Ling, JIANG Liwen, TAN Xinghe,
2017, 38(14):  120-126.  doi:10.7506/spkx1002-6630-201714018
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The diversity of the bacterial community in soybean paste fermented under variable temperatures was analyzed by DNA extraction, PCR amplification and high-throughput sequencing technology. The results showed a total of 2081 and 1 870 operational taxonomic units (OTU) were obtained from the samples fermented at low temperature first and then high temperature (mode 1) or at high temperature first and then low temperature (mode 2), respectively. Firmicutes was the most dominant phylum for both fermentation modes with an abundance value of 72.24% and 43.84%, respectively. Staphylococcus was the most dominant genus with an abundance value of 44.13% and 32.52% for modes 1 and 2, respectively. The bacterial diversity and abundance of mode 1 were generally higher than those of mode 2.
Effect of Natural Fermentation on the Gel Properties and Molecular Structure of Millet Starch
CAO Longkui, KOU Fang, KANG Lijun, SHEN Meng, NING Dongxue, XIA Tiantian, WANG Weihao
2017, 38(14):  127-131.  doi:10.7506/spkx1002-6630-201714019
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In this study, we investigated the effect of natural fermentation on the gel properties and molecular structure of millet starch, aiming to provide a scientific and theoretical basis for the study of the mechanism of the modification of millet starch after fermentation for mechanized and industrial production of fermented millet products. Changes in the molecular structure and gel properties of millet starch (the ‘Huangjinmiao’ variety from Inner Mongolia) were examined after 144 h of natural fermentation. The results were showed that the surface of fermented starch granules was eroded, whereas the crystalline?structure did not change and remained A-type. The position of X-ray diffraction peaks of starch granules did not change after fermentation, but the peak intensity was reduced. Moreover, the weight-average molecular?mass of both region Ⅰ (amylopectin) and Ⅱ (amylose) was decreased from 2.5 × 104–5.9 × 105 to 2.2 × 104–5.4 × 105 g/mol and the number-average molecular?mass was increased. The setback, peak viscosity of millet starch were decreased by 425 and 470 mPa·s as compared to those before fermentation, gelatinization temperature and enthalpy value were increased by 1.62 ℃ and 7.05 J/g, and gel hardness and elasticity were decreased by 60.735 g and 0.707, respectively. After fermentation for 96 h, the gel properties of starch tended to be stable. The findings revealed that both the molecular structure and gel properties of millet starch after natural fermentation were changed, being more suitable for the production of starch products with resistance to retrogradation.
Effect of Enzymatic Hydrolysis on Quality of Red Jujube Juice Fermented by Lactobacillu plantarum
ZHANG Lihua, ZHAO Guangyuan, WU Shunhong, DONG Yarui, LIU Nian, ZONG Wei,
2017, 38(14):  132-137.  doi:10.7506/spkx1002-6630-201714020
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In order to clarify the effect of enzymatic hydrolysis on the quality of fermented red jujube juice, red jujube juice was hydrolyzed by a combination of pectinase and hemicellulase, fermented by Lactobacillu plantarum and then stored at 4 ℃, and changes in the viable bacterial count, major nutrients and color were investigated at different storage times. The results showed that enzymatic hydrolysis significantly improved the contents of soluble solids and titratable acid in fermented jujube juice as compared with the control. Enzymatic treatment was beneficial to maintaining the contents of ascorbic acid, polyphenol and protein of fermented jujube juice during storage, which were 0.7, 0.28 and 0.58 mg/100 mL higher than those of the control after storage for 28 days, respectively. Additionally, the polysaccharide content was also increased slightly and the color was significantly improved. In addition, the viable number of lactic acid bacteria in fermented jujube juice was increased by enzymatic pretreatment, which was significantly higher than that of the control (0.17 (lg (CFU/mL))) at 28 days of storage. These results showed that the quality of fermented jujube juice could be improved by enzymatic pretreatment before fermentation.
Component Analysis
Screening and Identification of Flavor Characteristics of Three Pear Wines Based on Odor Activity Value Using Partial Least Squares Regression
ZHOU Wenjie, WANG Peng, ZHAN Ping, TIAN Honglei
2017, 38(14):  138-143.  doi:10.7506/spkx1002-6630-201714021
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The aroma compounds of three commercial pear wines were determined using solid phase microextraction (SPME) in combination with gas chromatography-mass spectrometry (GC-MS). A total of 43 aroma compounds were identified, including 16 alcohols, 15 esters, 4 aldehydes, 2 ketones, 1 phenol, 3 acids and 2 other compounds. Partial least squares regression (PLSR) were used to determine the characteristics of aroma compounds based on their odor activity values (OAV) and to deduce their contribution to the aroma of the wines. The characteristic aroma compounds identified mainly included isobutanol, 1-octano, 1-alcohol, phenylethyl alcohol, ethyl butyrate, ethyl 3-methyl butyrate, isoamyl acetate, ethyl caproate, ethyl caprylate, β-damascus ketone, and eugenol, and their possible correlation with six sensory attributes (fermented, acidic, fruity, floral, sweet, and delicate aromas) was fitted to a PLSR model. It turned out that the OAV of benzyl alcohol, n-butanol, and ethyl succinate were less than 1, which contributed to the aroma of pear wines. There was a good correlation between OAV of wine aroma compounds and fermented and sweet aromas while no striking correlations were observed with delicate, fruity and floral aromas.
Simultaneous Determination of Four Active Compounds in Sea Buckthorn Three Different Geographic RegionsHigh Performance Capillary Zone Electrophoresis
YANG Xia, FENG Feng, LIU Lizhen, CHEN Zezhong
2017, 38(14):  144-150.  doi:10.7506/spkx1002-6630-201714022
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A high performance capillary zone electrophoresis (HPCZE) method was developed for the simultaneous separation and determination of rutin, kaempferol, isorhamnetin and quercetin in sea buckthorn. The effects of different electrophoresis conditions on separation efficiency were investigated. The running buffer was 20 mmol/L Na2B4O7-H3BO3 with 1.5 mg/mL β-cyclodextrin at pH 9.55, the detection wavelength was set at 370 nm, applied voltage 25 kV and injection time 5 s. Under these optimal conditions, rutin, kaempferol, isorhamnetin and quercetin could be fully separated each other within eleven minutes with good linearity in the range of 0.01 to 0.51, 0.05 to 0.93, 0.02 to 0.65, and 0.03 to 0.81 mg/mL, respectively and correlation coefficients between 0.997 1 and 0.999 1. The lowest detection limits were 5.05 × 10-5, 2.10 × 10-5, 3.75 × 10-5, and 1.31 × 10-5 mg/mL, respectively (RSN 3). The average recoveries of spiked samples were in the range 95.51% to 104.66%, with a relative standard deviation (RSD) lower than 3.92% (n 3). Under the optimized conditions, a mixture of four standard compounds and four active compounds in extracts of sea buckthorn three different producing areas were determined. The results showed that the total content of active compounds was the highest in seabuckthorn ürümqi, Xinjiang among three different producing regions, and lowest in the sample Wuwei, Gansu. Furthermore, the sample Lüliang, Shanxi had the highest quercetin content while the contents of kaempferol and isorhamnetin were the highest in the sample ürümqi, Xinjiang. This method is rapid, simple, accurate and reliable for the simultaneous detection of active compounds in sea buckthorn.
Optimization of Headspace Solid Phase Microextraction (HS-SPME) for Analysis of Volatile Constituents of Flaxseed Oil from Xinjiang by Gas Chromatography-Mass Spectrometry (GC-MS)
WEI Changqing, ZHOU Qi, LIU Wenyu
2017, 38(14):  151-157.  doi:10.7506/spkx1002-6630-201714023
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This study optimized the headspace solid phase microextraction (HS-SPME) conditions for the analysis of the volatile constituents of flaxseed oil from Xinjiang by gas chromatography-mass spectrometry (GC-MS). The optimum HS-SPME conditions established were as follows: 40 min extraction using a DVB/CAR/PDMS fiber at 50 ℃ with magnetic stirring followed by 4 min desorption. Under these conditions, a total of 46 volatile compounds were identified consisting mainly of aldehydes, ketones, alcohols, acids, esters, alkanes and heterocyclics and other miscellaneous compounds with aldehydes, heterocyclics and alcohols being predominant among these, accounting for 30.14%, 12.73% and 9.25% of the total identified components, respectively. A good chromatographic separation was achieved in this study.
Effects of Drying Conditions on Aroma Compounds of Hami Jujube from Xinjiang
CHEN Kai, LI Qiong, ZHOU Tong, FU Bing, LI Jinyu, LI Huanrong
2017, 38(14):  158-163.  doi:10.7506/spkx1002-6630-201714024
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The objective of the current study was to explore the effect of different drying methods on the aroma compound composition of red jujubes. Fresh Hami jujubes from Xinjiang and dried jujubes obtained by different drying methods, namely, hot air drying at 40 ℃ and 45 ℃, sun drying, and shade drying were analyzed by head-space solid phase micro-extraction coupled with gas chromatography-mass spectrometry (GC-MS) for their aroma compound composition. The results showed that a 50/30 μm divinylbenzene/carboxen/polydimethylsiloxane (DVB/CAR/PDMS) fiber exhibited high sensitivity and selectivity in the extraction of aroma compounds. A total of 66 aroma compounds were identified from five samples, the predominant ones being aldehydes, esters, acids and ketone. Principal component analysis (PCA) was adopted to investigate the eight main aroma compounds for Chinese jujube. Jujubes dried by hot air at 45 ℃ had the best aroma quality, followed by those dried at 40 ℃ and the sun dried ones, and the aroma quality of the shade dried samples was the worst.
Comparative HPLC Analysis of Flavonoids from Different Raw Honeys
WANG Qin, LI Zhi, DANG Xiaoqun, ZHOU Zeyang, WANG Linling
2017, 38(14):  164-169.  doi:10.7506/spkx1002-6630-201714025
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In this study, by using high performance liquid chromatography (HPLC), we compared the flavonoid composition and contents of raw honey from different honeybee species, different geographic origins and different floral sources and commercial processed honey. The results showed that Chinese honeybee (Apis cerana cerana) and Italy honeybee (Apis mellifera ligustica) rape honey were similar in terms of total peak number and peak shape, but the total flavonoids content of Italy honeybee rape honey was higher than that of its Chinese counterpart. Italy honeybee rape honeys from different producing areas had a high similarity in the HPLC profiles, and they were almost identical with respect to the number and amount of flavonoids identified. The number and amount of flavonoids detected in processed honey were reduced as compared to the raw one. The HPLC profile of processed honey indicated that the chromatographic peaks of flavonoids occurred earlier, which were intensively distributed in the water-soluble phase. Considerable differences in total peak number and peak shape were found among raw honey from different floral sources; a total of 65, 58 and 70 flavonoid peaks were identified from rape honey, acacia honey and citrus honey, respectively, and citrus honey had the highest flavonoid content (161.62 μg/100 g). The HPLC peak shape of flavonoids in monofloral honey showed some specificity, providing a characteristic fingerprint for the identification of honey species.
Simultaneous Determination of Water-Soluble Sugars in Fresh Jujube by High Performance Liquid Chromatography with Evaporative Light-Scattering Detector (HPLC-ELSD)
PU Xiaoqiu, BAI Hongjin, MA Qianqian, WU Cuiyun
2017, 38(14):  170-174.  doi:10.7506/spkx1002-6630-201714026
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A method for the simultaneous determination of water-soluble sugars in fresh jujubes by high performance liquid chromatography with evaporative light-scattering detector (HPLC-ELSD) was established. The carbohydrates were separated on a chromatographic column of Waters XBridgeTM Amide (4.6 mm × 250 mm, 5 μm) with a mobile phase consisting of A (0.2% triethylamine aqueous solution) and B (0.2% triethylamine in acetonitrile) (A:B = 24:76, V/V). The column temperature was 30 ℃ and the atomization temperature was 60 ℃. The drift tube temperature of ELSD was 60 ℃. Nitrogen was used as the carried gas with a flow rate of 1.6 L/min and a gain value of 1.0. These experimental conditions allowed good separation of water-soluble sugars, such as rhamnose, arabinose, glucose, mannose, glucose, sucrose, maltose and lactose with a resolution of 1.5 or above. Good linearity was observed for the analytes in the concentration range of 0.099–1.040 ng/μL. The recoveries of spiked real samples ranged from 93.1% to 111.2%, with relative standard deviation (RSD) values below 5%. The limits of detection (RSN = 3) were in the range of 0.008–0.03 ng/μL. This method had a lot of advantages including simple operation, effective separation and time saving, and could be used to determine water-soluble sugars in fresh jujube. Compared with the traditional spectrophotometric method, HPLC-ELSD was more sensitive and accurate making it more suitable for quality identification of fresh jujube.
Change of Taste Components in Thermal Reaction Model System Containing Chicken Enzymatic Hydrolysate with Reaction Time
HOU Baihui, XIA Yangyi, ZHOU Tao, ZHANG Weiyue
2017, 38(14):  175-180.  doi:10.7506/spkx1002-6630-201714027
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In this study, a thermal reaction model system consisting of papain hydrolysate of chicken breast and glucose was built and used to investigate the change of taste components during the reaction process through analysis by?gas chromatography-mass spectrometry (GC-MS) and using an automatic amino acid analyzer. The?results?indicated?that with the increase in heating time, the contents of free amino acids and taste-active amino acids increased significantly (P < 0.05), with bitter amino acids being the dominant ones. A total of 66 volatile compounds including 25 aldehydes, 7 alkanes, 6 ketones, 8 alchols, 13 acids, 1 heterocyclic compound, 2 aromatics, 3 esters and 1 alkene compound were detected, 37 of which were detected in the control group (not heated), and 44, 51 and 52 of which were detected at different reaction times of 30, 60 and 90 min, respectively. Twenty-four volatile compounds were common to these four samples. The content of volatile compounds increased with increasing reaction time, reaching a maximum at 90 min. In conclusion, volatile compounds could be more easily produced chicken enzymatic hydrolysate under thermal reaction conditions.
Processing Technology
Removal of Cadmium from Lentinus edodes Using Citric Acid: Optimization of Processing Conditions Using Response Surface Methodology
HU Jing, SUN Junshe, TAN Xiaoyan, HU Shaofeng, QIN Zifang, NING Huijuan, ZHANG Xiuqing
2017, 38(14):  181-186.  doi:10.7506/spkx1002-6630-201714028
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The removal of the heavy metal cadmium in dried Lentinus edodes using citric acid content was optimized using a combination of one-factor-at-a-time method and response surface methodology. The removal efficiency of cadmium was investigated as a function of three independent variables: citric acid concentration, pH and rotation speed. It turned out that the maximum removal rate of cadmium of 57.0% was obtained under the following optimized conditions: citric acid concentration 0.32 mol/L, pH 5.0, and rotation speed 70 r/min. After the treatment, the maximum residual cadmium content was 0.41 mg/kg, lower than the national limit of 0.5 mg/kg. Meanwhile, low losses of polysacccharides and proteins were observed suggesting good retention of nutrients in L. edodes. The proposed method could have a high application potential for reducing residual cadmium in L. edodes.
Optimization of Chelation of Juncao Ganoderma lucidum Peptides with Selenium by Response Surface Methodology
ZHAO Lina, CHEN Zihong, CHEN Hao, HUA Pengpeng, LIU Bin,
2017, 38(14):  187-192.  doi:10.7506/spkx1002-6630-201714029
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The purpose of the present study was to optimize the preparation of Juncao Ganoderma lucidum peptide-selenium chelate using response surface methodology. The independent variables were pH, the ratio of sodium selenite solution to protein hydrolysate (V/V), reaction time, and temperature. The response was selenium content. A quadratic polynomial regression model was established and its reliability was validated. The chelate was characterized by infrared (IR) spectroscopy. The results showed that in decreasing order significance, pH, reaction temperature, the ratio of sodium selenite solution to protein hydrolysate and reaction time affected the selenium content of chelates, and the optimal values of these factors were 9, 75 ℃, 1:2 and 60 min, respectively. The selenium content of the obtained product was (2 985.89±10.59) μg/g. IR spectral analysis confirmed the chelation reaction between Juncao Ganoderma lucidum peptides and selenium ions and the generation of a new chelate. This study can provide a new approach for the development and utilization of Juncao Ganoderma lucidum.
Optimization of Neutroproteinase-Assistant Extraction of Polysaccharide from the Stroma of Shiraia bambusicola Henn by Response Surface Analysis and Characterization of the Extracted Polysaccharide
GE Xuejun, ZHOU Dejian, WANG Bin, CHEN Yin
2017, 38(14):  193-199.  doi:10.7506/spkx1002-6630-201714030
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Objective: This study aimed to optimize the neutroproteinase-assisted extraction of polysaccharides from the stroma of Shiraia bambusicola Henn using response surface methodology (RSM) and to investigate the separation, purification, characterization and antioxidant activity of the polysaccharides. Methods: The effects of extraction time, enzyme concentration and extraction temperature on the yield of polysaccharides were examined by using one-factor-at-a-time experiments. These extraction conditions were optimized through mathematical modeling. The crude extract was fractionated into two fractions: SBH-1 and SBH-2, with SBH-1 being among them. The monosaccharide composition, molecular mass and structure of SBH-1 were detected by high performance gel permeation chromatography (HPGPC) and infrared spectroscopy, respectively. The in vitro antioxidant activity was measured in comparison with VC by superoxide anion scavenging, 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging, and lipid peroxidation inhibition assays. Results: The maximum yield of crude polysaccharides of 7.63% was obtained at 115.95 min of extraction at 46.68 ℃ with 0.83% neutral proteinase. The molecular mass of SBH-1 was determined to be 18.3 kD, and its monosaccharide composition consisted of glucose, mannose, and galactose with a ratio of 1.3:2.3:1.2. The EC50 values for scavenging of DPPH and superoxide anion radials were 2.03 and 1.038 mg/mL respectively. Conclusions: The neutroproteinase-assistant extraction procedure was simple and easy to operate. SBH-1 was a neutral heteropolysaccharide with high activity. This study can provides a theoretical basis for further research and development of polysaccharides from S. bambusicola Henn.
Simultaneous Extraction Purification of Forsythoside A, Rutin and Phillyrin from Forsythia suspensa Leaves
SUN Jianrui, WANG Dahong, QIU Zhijun, YUAN Jiangfeng
2017, 38(14):  200-205.  doi:10.7506/spkx1002-6630-201714031
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In this study, we optimized the simultaneous separation and purification of forsythiaside A, phillyrin and rutin from an ethanolic extract of Forsythia suspensa leaves. These target compounds were quantitatively determined by high performance liquid chromatography (HPLC). Different types of macroporous adsorption resin were compared for their abilities to separate and purify three active components from Forsythia suspensa leaves. AB-8 resin was selected as the best one, and the optimum chromatographic conditions on this adsorbent were obtained as follows: sample-to-resin ratio, 1.5:1 (g/g); sample loading flow rate, 2 bed volumes (BV)/h; sequential elution with 8 BV of deionized water, 6 BV of 30% ethanol and 6 BV of 50% ethanol at a flow rate of 3 BV/h. Further purification of forsythiaside A was carried out using a C18-silica gel column by reversed-phase chromatography, while purified phillyrin and rutin were obtained by precipitation after standing. After purification, the purity of forsythiaside A and rutin was more than 97%, and that of phillyrin was over 95%. In conclusion, under the optimized conditions, three effective components of high purity were obtained from Forsythia suspensa leaves. This study may be helpful for the comprehensive utilization and industrialization exploitation of Forsythia suspensa leaves.
Formulation and Application of Non-Oil-Based Coffee Mate
HOU Chunyan, WU Tao, LIU Rui, ZHANG Min
2017, 38(14):  206-212.  doi:10.7506/spkx1002-6630-201714032
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This study aimed to develop a formulation for non-oil-based coffee mate using a protein-based fat simulant and to investigation its application. The non-oil-based coffee mate with good external appearance and stability was formulated with the protein-based fat simulant obtained by physical modification of whey protein concentrate and egg white protein without added oil. The optimal formulation found was protein-based fat simulant 88.68%, glucose syrup 6.03%, beta cyclodextrin 0.89%, sucrose 1.95%, calcium phosphate 0.05%, flaxseed gum 0.09%, guar gum 0.09%, sucrose fatty acid ester 0.44%, and hydroxypropyl starch phosphate 0.89%. The formulated product was demonstrated to have good sensory and functional properties. In addition, the microrheological and reconstitution properties were evaluated, and the sensory quality differences in comparison to a commercial product were determined?using a triangular?sensory?test. The experimental results showed that the sensory and physicochemical properties of the non-oil-based coffee mate were similar to those of the commercial sample, with no significant differences being observed between the two. The 24 h stability index under ambient temperature was less than 2 for both samples, suggesting their comparable stability. Moreover, they were comparable to each other in terms of microrheological and reconstitution properties.
Separation and Purification of α-Linolenic Acid from Phyllanthus emblica L. Seed Oil by Silver Iron Complexation
GE Shuangshuang, ZHANG Wenwen, LI Kun, FENG Ying, GAN Jin, ZHENG Hua, ZHANG Hong
2017, 38(14):  213-220.  doi:10.7506/spkx1002-6630-201714033
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The objective of this research was to purify α-linolenic acid from Phyllanthus emblica L. seed oil by using silver iron complexation in order to obtain high purity and yield of α-linolenic acid. Firstly, the effects of silver nitrate concentration, volume fraction of methanol, complexation time and complexation temperature on the purity and yield of α-linolenic acid were investigated by one-factor-at-a-time method. Afterwards, three independent variables were selected out of the four process parameters for the optimization experiments which were done using Box-Behnken design with the purity and yield of α-linolenic acid as response variables. The results indicated that the optimal process parameters were obtained as follows: complexation temperature, 0 ℃; silver nitrate concentration, 2.29 mol/L; methanol volume fraction, 38.00%; and complexation time, 1.93 h, under which the purity and yield of α-linolenic acid were 93.30% and 73.37%, respectively. The remaining silver nitrate was recovered for a second complexation. The recovery of silver nitrate was 93.83% and the recovered Ag+ still had good complexation effect. Particularly, exogenous Ag+ was not introduced in the whole complexation process. Therefore, this method allowed successful separation and purification of α-linolenic acid from P. emblica L. seed oil while reducing the operating cost. Meanwhile, the method of silver iron complexation was safe and helpful for industrial application of P. emblica L. seed oil.
Response Surface Optimization of GABA Accumulation in Germinated Quinoa under Citric Acid Stress and Its ACE Inhibitory Activity in Vitro
GUO Xiaomeng, ZHAO Fushi, YE Xiaohui, MA Tingjun,
2017, 38(14):  221-226.  doi:10.7506/spkx1002-6630-201714034
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This study aimed to optimize the culture conditions for γ-aminobutyric acid (GABA) production in germinated quinoa under citric acid stress. GABA from germinated quinoa was ultrasonically extracted with ethanol and analyzed by high performance liquid chromatography (HPLC). The optimization of citric acid concentration, culture time and temperature, the factors affecting GABA content in germinated quinoa, were optimized by the combined use of one-factor-at-a-time method and response surface methodology. Results indicated that the highest GABA content of 1.538 mg/g in germinated quinoa was obtained after 48 h of culture at 25 ℃ in the presence of 2.00 mmol/L citric acid, which was 3.8 times as high as that observed for the ungerminated sample. The percentage of ACE inhibition by germinated quinoa under citric acid stress was 63%, which was 1.3 and 1.9 times higher than that that by the germinated one without citric acid stress and the ungerminated one, respectively. This finding suggested that germination under citric acid stress could improve the antihypertensive activity of quinoa, which will provide a guideline for further study of quinoa.
Optimization of Ultrasonic-Assisted Extraction of Trypsin Inhibitor from Pea Seeds by
YU Hong’en, LIU Hongkai, KANG Yufan
2017, 38(14):  227-232.  doi:10.7506/spkx1002-6630-201714035
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The purpose of the current study was the optimization of the solvent extraction of trypsin inhibitor from pea seeds with ultrasonic assistance. A sample granularity of 80 mesh, an ultrasonic frequency of 40 kHz, a radiation time of 13 min, an extraction temperature of 40 ℃, a solid-to-solvent ratio of 1:80 (g/mL), and an ultrasonic power of 500 W were determined to be optimal using one-factor-at-a-time method. Ultrasonic treatment time, extraction temperature, solid-to-solvent and ultrasonic power were taken as independent variables for further optimization by establishing a quadratic polynomial regression model to predict trypsin inhibitor activity using a Box-Behnken design and analyzing the model by response surface methodology. The final optimized extraction conditions were found to be 80 mesh, 40 kHz, 16 min, 50 ℃, 1:100 (g/mL) and 500 W for sample granularity, and ultrasonic frequency, ultrasonic treatment time, extraction temperature, 50 ℃ and solid-to-solvent ratio, respectively. Under these conditions, the maximum trypsin inhibitor activity of 574.54 TIU/g was obtained experimentally, showing a relative error compared to the predicted value of 1.42%. The developed model exhibited an adjusted coefficient of determination (R2Adj) of 0.886 4 and a coefficient of determination (R2) of 0.943 2, indicating?an?excellent goodness of fit. The model is of great guidance significance.
Optimization of Ultrasound-Vacuum Assisted Extraction of Polyphenols from Apple Pomace
TIAN Li, LI Haiping, YUAN Yahong, YUE Tianli, WEI Jianping
2017, 38(14):  233-239.  doi:10.7506/spkx1002-6630-201714036
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One-factor-at-a-time method and response surface methodology were used in combination to optimize the ultrasound-vacuum assisted extraction of polyphenols from apple pomace. At the same time, antioxidant assays in vitro were also carried out on the polyphenols and the polyphenol composition was analyzed by HPLC. The optimal extraction parameters that provided the maximum yield of polyphenols of 6.46?mg/g were determined as follows: 50% ethanol as extraction solvent, a solid-to-solvent ratio of 1:30 (g/mL), an extraction time of 13?min, an extraction temperature of 50 ℃, an ultrasound power of 420?W, and a vacuum degree of 0.08 MPa. Compared with traditional ultrasonic assisted extraction, the extraction time was considerably shortened by using ultrasound-vacuum assisted extraction while yielding polyphenol extracts containing more epicatechin and rutin and having stronger antioxidant activities.
Optimization of Enzymatic Hydrolysis of Oysters and Amino Acid Composition and Nutritional Quality of Oyster Hydrolysates
LIU Haimei, CHEN Jing, AN Xiaoyu, ZHAO Qin, GUO Qingjun, CHE Xinxin, CUI Yun
2017, 38(14):  240-244.  doi:10.7506/spkx1002-6630-201714037
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The enzymatic hydrolysis of oysters with Protamex was optimized using one-factor-at-a-time and orthogonal array design methods. The independent variables were hydrolysis time, enzyme dosage and temperature, and the response was degree of hydrolysis. The solubility characteristics of oyster hydrolysates were evaluated by nitrogen solubility index (NSI), trichloroacetic acid-nitrogen solubility index (TCA-NSI), and the amino acid score (AAS), chemical score (CS) and essential amino acid index (EAAI) were measured for nutritional quality evaluation. The optimal hydrolysis parameters were determined as follows: enzyme dosage, 30 AU/kg; temperature, 55 ℃; and time, 4 h, yielding a degree of hydrolysis of up to 27.28%. The obtained hydrolysate contained 18 amino acids, being high in umami amino acids, with Glu being the most abundant and essential amino acids accounting for 39.5% of the total amino acids, suggesting high nutritional value. At pH 4.0–7.0, the NSI of the hydrolsate was over 83%, and TCA-NSI was as high as 88.64% indicating good solubility.
Extraction and Structural Characterization of Hemicellulose from Coconut Coir Fiber
WANG Zheng, HUANG Guangmin, ZHAO Bin, LI Shandan, LU Daxing, HU Dan
2017, 38(14):  245-249.  doi:10.7506/spkx1002-6630-201714038
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In this investigation, hemicellulose was extracted from crude coconut coir fiber with aqueous sodium hydroxide solution, and its structure was analyzed. It turned out that the highest yield (14.33%) of hemicellulose was obtained when the extraction was performed at 65 ℃ for 4.0 h in 5% sodium hydroxide solution with a solid-to-solvent ratio of 1:50 (g/mL). The extracted product was confirmed as hemicelluloses using infrared spectroscopy. By gel permeation chromatography, the number-average molecular weight of the hemicellulose was 49 049 D and weight-average molecular weight 368 354 D. The hemicellulose was hydrolyzed with sulfuric acid and analyzed by high performance liquid chromatography, and the results showed that the retention time was completely consistent with that of standard xylose.
Optimization of Blasting Extrusion Processing for Production of Lentinus edodes Powder and Its Physicochemical Properties
ZHU Hui, LI Yuntong, CHEN Guiyun, CHEN Ye
2017, 38(14):  250-255.  doi:10.7506/spkx1002-6630-201714039
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This work reports on the production of Lentinus edodes powder to be used as ingredient of seasonings by blasting extrusion processing. The processing conditions were optimized using a combination of one-factor-at-a-time and orthogonal array design methods. Physiochemical properties of Lentinus edodes powder were evaluated and compared before and after blasting extrusion and we also determined the conditions and scope of application of Lentinus edodes powder and its nutritional changes. The optimal processing parameters that provided the highest total content of inosinic acid (IMP) and guanylic acid (GMP) were determined as follows: moisture content of raw material 50%, temperature 150 ℃, and mold hole diameter 3 mm. Fat content of Lentinus edodes powder was decreased after extrusion. In addition, the extruded Lentinus edodes powder had stable thermal properties, its surface became more smooth and compact, and its micromeritic properties were improved without significant changes in active groups. The extruded Lentinus edodes powder could be used as an ingredient of seasonings or functional food additive in food industry.
Optimization of Adsorption Removal of Alternaria Mycotoxin TeA from Citrus Juice by Inactive Lactic Acid Bacteria
GE Na, PENG Bangzhu, XU Xiaoyun, PAN Siyi
2017, 38(14):  256-262.  doi:10.7506/spkx1002-6630-201714040
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Citrus fruit is susceptible to pollution by Alternaria during growth and post-harvest storage. Tenuazonic acid (TeA) is considered to be one of the most toxic mycotoxins produced by Alternaria, and constitutes a serious threat to consumers’ health and safety. Therefore, this study aimed to obtain the optimal conditions for adsorption removal of TeA from citrus juice using inactive lactic acid bacteria (LAB) as adsorbent. The optimization was implemented using a combination of one-factor-at-a-time method and response surface methodology with Box-Behnken design. The independent variables were adsorbent dosage, adsorption time, initial concentration of TeA and pH of citrus juice. The response variable was the percentage removal of TeA. An adsorption time of 11.43 h, 1.20 g/20 mL of inactive LAB, an initial TeA concentration of 250 μg/L and pH of 3.15 were determined as to be optimal. Under these optimal conditions, the percentage adsorption of TeA from citrus juice was 86.98%. A quadratic polynomial regression model was established and validated between the independent variables and the response. It turned out that the model had a high goodness?of?fit and good prediction performance.
Optimization of Enzymatic Hydrolysis Conditions for the Preparation of Antioxidant Peptides and Amino Acid Composition Analysis from Alfalfa Leaf Protein
LIU Long, ZHANG Wei, CHEN Yuantao, LIU Haibin, ZHANG Qi, GAO Zhongchao, LEI Lei
2017, 38(14):  263-269.  doi:10.7506/spkx1002-6630-201714041
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In order to develop and utilize alfalfa leaves as a good resource of protein, antioxidant peptides were prepared from alfalfa leaf protein by alkaline protease hydrolysis. Through one-factors-at-a-time experiments, hydrolysis time, enzyme concentration, hydrolysis temperature and pH were selected as independent variables for further optimization using response surface methodology, and the response variable was antioxidant activity. The optimum conditions that provided the maximum percentage scavenging of DPPH radical of 64.25% were found to be hydrolysis time of 240 minutes, enzyme concentration of 4.80%, hydrolysis temperature of 60 ℃ and pH of 11.60. Then the most potent antioxidant peptide was purified and collected by preparative liquid chromatography, having stronger antioxidant activity than the crude hydrolysate. The amino acid composition of the antioxidant peptide was determined by high performance chromatography (HPLC) to consist of Asp, Gly, His, Tyr, Val, Leu, Phe, Trp and Lys.
Optimization of Apple Pomace Pectin Extraction by Steam Explosion Employing Response Surface Methodology
SUN Junliang, DU Hanmei, LIANG Xinhong, JIAO Zhonggao, RAN Junjian, HE Hongju, ZHU Mingming
2017, 38(14):  270-275.  doi:10.7506/spkx1002-6630-201714042
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Pre-soaking in aqueous Na2CO3 solution followed by steam explosion was employed in the extraction of apple pomace pectin for the purpose of improving the product yield. Processing parameters were optimized using a combination of one-factor-at-a-time method and response surface methodology. The results showed that a Na2CO3 concentration of 6% and an explosion pressure of 0.6 MPa maintained for 174 s were found to be optimal the maximum pectin yield of 21.42%, 10.96% higher than that obtained without steam explosion. In addition, the esterification degree, emulsifying activity and emulsion stability of pectin were increased by 12.25%, 20.47 m2/g, and 36.37 min, respectively by steam explosion. Scanning electron microscopy results revealed that the surface of both apple pomace and pectin extracted without steam explosion was smooth and complete, which became loose after steam explosion. The application of steam explosion in pectin extraction from apple pomace could improve the yield, esterification degree, emulsifying activity and emulsion stability of pectin. This research can provide theoretical and practical evidence for the production of apple pectin.
Safety Detection
Positional Distribution of Trans Fatty Acids in Triglycerides and Phospholipids of Partially Hydrogenated Soybean Oil and Ruminant Animal Fat
HU Shengben, ZHUO Chengfei, ZOU Guyue, DENG Zeyuan, LI Jing
2017, 38(14):  276-283.  doi:10.7506/spkx1002-6630-201714043
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Lipids from partially hydrogenated soybean oil and ruminant animal fat were extracted by the Folch methods, and then separated using a NH2 solid phase extraction (SPE) cartridge to obtain triglycerides and phospholipids. The analyses were carried out after using Sn-1,3-specific lipase to hydrolyze the acyl group Sn-1 and Sn-3 in triglycerides, and using phospholipase A2 to hydrolyze the acyl group at the Sn-2 position of phospholipids, which were finally converted to fatty acid methyl esters and analyzed by gas chromatography. Results showed that trans fatty acids accounted for 41.466% and 2.451%–3.179% of the total fatty acids in triglycerides from hydrogenated soybean oil and ruminant animal fat, respectively. Trans fatty acids were selectively located at the Sn-2 position of hydrogenated soybean oil triglycerides and at the Sn-1,3 position of ruminant animal fat triglycerides. For ruminant animal fat phospholipids, trans fatty acids were preferentially located at the Sn-1 position. Dietary intake of trans fatty acids from different sources and their different positional distribution preferences in triglycerides may be the cause of the fact that trans fatty acids from different sources have different impacts on human health.
Quantification of Added Egg White in Surimi Products by Competitive Enzyme Linked-Immunosorbent Assay
ZHAO Yongjuan, WENG Ling, YAN Longjie, LIU Mengqi, ZHANG Lingjing, CAO Minjie,
2017, 38(14):  284-289.  doi:10.7506/spkx1002-6630-201714044
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In this study, a competitive enzyme linked-immunosorbent assay (c-ELISA) using the polyclonal antibody against purified ovalbumin (OVA), the major protein in egg white, was proposed to quantify added egg white in surimi products. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot results showed the high specificity of the anti-OVA polyclonal antibody. The limit of detection (LOD) of the c-ELISA method was 1.82 mg/kg, and the intra- and inter-assay coefficients of variation were 3.1% and 7.8%, respectively. Recovery rates of egg white added to fish meatballs ranged from 87.4% to 97.2%. The c-ELISA method was successfully applied to quantify added egg white in commercial surimi products, and results demonstrated that the added egg white content was 2.7–83.5 g/kg in six samples from different manufacturers. Our results indicated that the method established can be applied to quantify added egg white in surimi products.
Spectrophotometric Determination of Cyanide Content in Rubber Seeds Using Isonicotinic Acid-Pyrazolone after Exogenous β-Glucosidase Pretreatment
ZHANG Ye, WANG Ke, LIU Shisheng
2017, 38(14):  290-296.  doi:10.7506/spkx1002-6630-201714045
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This study investigated the determination of the residual cyanide content of rubber seeds detoxified by different methods by a spectrophotometric method using isonicotinic acid-pyrazolone with and without exogenous β-glucosidase pretreatment. The results showed that cyanide was undetectable or detectable at a level lower than the actual one by the spectrophotometric method without exogenous β-glucosidase pretreatment, while it was detected at a significantly higher level with the sample pretreatment. The optimized pretreatment conditions were 3.35 h soaking at 45.06 ℃ and pH 5.56 with 24.11 U/100 mL of β-glucosidase added, under which, the predicted cyanide content of rubber seeds was 28.79 μg/g. The recovery of spiked samples ranged from 85.02% to 92.04%. The proposed method was accurate and reliable.
Non-Destructive Detection of Moldy Core in Apple Fruit Based on Deep Belief Network
ZHOU Zhaoyong, HE Dongjian, ZHANG Haihui, LEI Yu, SU Dong, CHEN Ketao
2017, 38(14):  297-303.  doi:10.7506/spkx1002-6630-201714046
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Apple moldy core is a major disease affecting the internal quality of apple fruit. However, due to the lack of effective means to accurately detect moldy core in apple fruits, detection of moldy core in apples has become a major problem to be solved in the apple industry. To date, there have been no reports on the use of spectroscopy for distinguishing various degrees of moldy core decay in apple fruits. The objective of this study was to develop a non-destructive method for the detection of various degrees of moldy core decay in apple fruits using near infrared transmittance spectroscopy, successive projections algorithm (SPA), and multi-class classification algorithms partial least square-discriminant analysis (PLS-DA), back propagation artificial neural network (BP-ANN), support vector machine (SVM) and deep belief network (DBN). For developing a model to determine the degree of moldy core in apples, 225 samples were selected including a training set of 150 samples and a test set of 75 samples. The model consisted of several layers of restricted Boltzmann machine (RBM) network, which achieved eigenvector projection, and one layer of BP network, which allowed the classification of various degrees of moldy core based on the output eigenvector. The Sd value was calculated by dividing the lesion area by the total cross-sectional area. It was proposed that Sd = 0, 0 < Sd ≤ 10%, 10% < Sd < 30% and Sd ≥ 30 indicated health, mild, moderate and severe degrees, respectively. The classification accuracy of the DBN model was 88.00%, suggesting good performance of the model, and it was compared with those of the BP-ANN and SVM models.
Rapid Identification of Lipid Oxidation Levels in Fish by Thin Layer Chromatography Image Analysis Technique
YU Hui, SATO Minoru, YAMAGUCHI Toshiyasu, NAKANO Toshiki
2017, 38(14):  304-309.  doi:10.7506/spkx1002-6630-201714047
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In this study, we developed a rapid method to determine the amount of polar compounds generated from lipid oxidation in fish based on thin layer chromatography (TLC) and image analysis in order to identify the extent of lipid oxidation in fish. The chromatographic separation of yellowfin tuna (Thunnus albacores) oil (0.5 μL) was performed on TLC plates using hexane-diethyl ether-acetic acid (30:70:1, V/V) as a mobile phase for 12 min. Color development was carried out by spaying the plates with 3% copper acetate in phosphoric acid and heating them at 150 ℃ for 15 min. The polar compounds derived from lipid oxidation during the heating process were semiquantitatively determined by area calculation with a special software. It was indicated that the amount of polar compounds measured by TLC was highly correlated with two common lipid oxidation parameters of peroxide value (POV) and carbonyl value (CV) with a correlation coefficient (R2) of 0.993 8 and 0.972 5, respectively. Furthermore, a good positive correlation between the measured amount of polar compounds and POV with a correlation coefficient of 0.962 5 was observed for salted Pleurogrammus azonus stored at 4 ℃, while salted sockeye salmon (Oncorhynchus nerka) stored at ?5 ℃ showed a good negative correlation between this measured parameter and color parameter a* value with a correlation coefficient of ?0.822 9. These results strongly suggest that TLC image analysis has the advantages of simplicity, rapidity, microscale quantitation and easy visualization without the need for expensive equipment.
Analysis of Adulterated Oilseed Rape Honey Using Differential Scanning Calorimetry Combined with Principal Component Analysis and Two-Dimensional Correlation Analysis
CHEN Guiyun, WU Wei, CHEN Kunjie
2017, 38(14):  310-315.  doi:10.7506/spkx1002-6630-201714048
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Differential scanning calorimetry (DSC) curves of adulterated oilseed rape honey in the temperature range of 30–100 ℃ were carefully analyzed by principal component analysis (PCA) and two-dimensional (2D) correlation analysis to reveal the effect of adulteration on DSC characteristics of honey. The results showed that 2D-DSC profiles under temperature perturbation and PCA could support each other and both methods could detect small changes that were difficult to detect in the original curve. Adulteration affected heat flow rate irregularly by and large. The former method was more intuitive in showing the irregular characteristics while the latter could reveal the hidden weak regularity. The two methods could be used in combination to further reveal the DSC properties of adulterated honey samples.
Purification Efficiency of Four Fusarium Toxins from Wheat Flour with Different Purification Columns
ZHANG Jianing, DING Ke, HAN Tao, CHEN Xiangning
2017, 38(14):  316-322.  doi:10.7506/spkx1002-6630-201714049
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A method for the simultaneous determination of four Fusarium toxins in wheat flour including zenralenone, deoxynivalenol, T-2 and HT-2 toxins was established using high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Samples were cleaned up with a MycoSep?226 multifunctional column, a DZT MS-PREP immunoaffinity column, which were both commercial products, and a polystyrene-divinylbenzene (PS-DVB) immunoaffinity column which was prepared in our laboratory, respectively. The recoveries of Fusarium toxins purified with MycoSep?226 ranged from 69.42% to 111.12% with relative standard deviations (RSDs) of 1.46%–13.24%, those obtained with PS-DVB ranged from 70.05% to 112.80% with RSDs of 3.42%–12.25%, and those obtained with DZT MS-PREP ranged from 88.01% to 107.31% with RSDs of 1.09%–14.42%. Immunoaffinity columns had the advantages of repeated use, less organic solvent consumption and high specificity. Moreover, PS-DVB could be a promising new immunoaffinity carrier.