Loading...

Table of Content

25 February 2018, Volume 39 Issue 4
Food Chemistry
Structural Changes of Muscle Sarcoplasmic Protein from Crisp Grass Carp (Ctenopharyngodon idellus C. et V) during Crispness Formation
FENG Jing, LIN Wanling, LI Laihao, YANG Xianqing, WANG Jinxu, WU Yanyan, HUANG Hui, HU Xiao, HAO Shuxian
2018, 39(4):  1-5.  doi:10.7506/spkx1002-6630-201804001
Asbtract ( 303 )   HTML ( 8)   PDF (2276KB) ( 204 )  
Related Articles | Metrics
The structural changes of muscle sarcoplasmic protein from crisp grass carp during crispness formation were investigated by measuring total sulfhydryl content, surface hydrophobicity, fluorescence spectrum and infrared spectrum. The results indicated that total sulfhydryl content was decreased along with crispness formation, showing a decrease of 35.02% at the end of the process (P < 0.05). Surface hydrophobicity showed an increase of 65.83% until the end of crispness formation (P < 0.05). The peak position in the fluorescence spectrum was significantly blue-shifted by 5.24% at the end of crispness formation (P < 0.05). Furthermore, infrared spectroscopy showed that characteristic absorption peaks (amides A, B, Ⅰ, Ⅱ and Ⅲ) were observed for muscle sarcoplasmic protein during crisp formation, the position of each of which was fluctuated. The secondary structure of sarcoplasmic protein showed that the relative contents of α-helix were decreased whereas those of β-sheet, β-turn and random coil were increased along with crispness formation. The structure of muscle sarcoplasmic protein changed gradually during crispness formation in grass carp, thereby affecting crispness formation.
Structures and Angiotensin Converting Enzyme (ACE) Inhibitory Activity of in Vitro Digests of Animal and Plant Proteins
LIANG Tingting, TONG Litao, PU Huayin, WANG Lili, ZHOU Xianrong, JU Zhiyuan, ZHOU Sumei, HUANG Junrong
2018, 39(4):  6-12.  doi:10.7506/spkx1002-6630-201804002
Asbtract ( 453 )   HTML ( 10)   PDF (2749KB) ( 312 )  
Related Articles | Metrics
This study aimed to reveal whether the difference in hypotensive activity between animal and plant proteins could be caused by the angiotensin conversion enzyme (ACE) inhibitory activity of peptides present in protein digests. The ACE inhibitory activity of peptides derived from in vitro digestion of plant and animal proteins was assessed. The in vitro digestion of plant proteins (rice, oat, soybean, and pea), red meat proteins (pork and beef), and white meat protein (chicken) was carried out by using a two-step process of hydrolysis with pepsin and trypsin. The hydrolysis degree, relative molecular weight distribution, amino acid composition and ACE inhibition rate of the obtained peptides were determined. The results showed that the inhibition percentage of ACE activity by protein digests increased with the increase in the degree of hydrolysis and consequently the decrease in the relative molecular weight. Among all protein digests, the ACE inhibition rate of plant protein digests was the highest (60.41%), while the lowest value (40.13%) was observed for red meat protein digests. The amino acid composition of protein digests showed that the levels of hydrophobic amino acids in both plant protein and white meat protein digests were significantly higher than in red meat protein digests. The higher the concentration of hydrophobic amino acids in protein digests was, the higher the ACE inhibitory activity was. It was suggested that the high content of hydrophobic amino acids may be one of the main causes of the ACE inhibitory activity of plant protein-derived peptides.
Relationship of Cathepsin with Free Amino Acids and Flavor Substances during Salted Decapterus maruadsi Processing
WU Yanyan, CAO Songmin, LI Laihao, YANG Xianqing, LIN Wanling, CHEN Shengjun
2018, 39(4):  13-19.  doi:10.7506/spkx1002-6630-201804003
Asbtract ( 493 )   HTML ( 9)   PDF (2409KB) ( 167 )  
Related Articles | Metrics
This study aimed to ascertain the relationships of cathepsin with proteolysis and flavor development in dry-salted Decapterus maruadsi. The fish were processed by two methods to obtain traditional high-salted fish (HS) and low-salted and lactic acid bacteria-fermented fish (LS). The changes in cathepsin B, L, and H activities, proteolysis index (PI) and free amino acid (FAA) contents were measured at different stage of processing. Furthermore, correlation analysis was performed using Pearson’s correlation test. The results showed a similar trend in cathepsin B, L and H activities between HS and LS samples. During the salting stage, cathepsin B, L and H activities in LS fish were higher than in HS fish; however, during drying cathepsin B and L activities in HS fish were higher than in LS fish, indicating HS method is more conducive to cathepsin B and L activities. LS method gave higher cathepsin than HS method during the whole process. The evolutionary trends of PI in LS and HS samples were identical. For both samples, PI and the contents of total free amino acids and total taste-active amino acids significantly increased during drying (P < 0.05); PI and total free amino acid content were higher in HS sample, while higher total content of taste-active amino acids was found in LS sample. Proteolysis by cathepsin B was more pronounced in HS sample. Both cathepsin B and L played an important role in promoting protein degradation and FAA formation, with the former being more effective. Compared with HS method, LS method could improve the percentage of taste-active amino acids as flavor substances while reducing the rate of protein degradation.
Effect of Slightly Acidic Electrolyzed Water on the Bioactive Compounds and Antioxidant Activity of Tartary Buckwheat Sprouts
ZHENG Chenxi, HAO Jianxiong, SONG Shuhui, JIANG Zhengqiang, LIU Haijie,
2018, 39(4):  20-25.  doi:10.7506/spkx1002-6630-201804004
Asbtract ( 319 )   HTML ( 5)   PDF (2645KB) ( 283 )  
Related Articles | Metrics
The bioactive components and antioxidant activity of tartary buckwheat sprouts treated with slightly acidic electrolyzed water (SAEW) with different available chlorine concentrations (ACC) were examined in this study. The results demonstrated that the content of bioactive components in buckwheat sprouts treated with SAEW was higher than that in sprouts treated with tap water (TW). The contents of total phenols, total flavonoids and rutin in sprouts treated with SAEW (pH 5.5, with 20 mg/L ACC) were increased by 28.69%, 26.19% and 29.08% respectively, as compared with those in the control group. The phenylalanine ammonia-lyase (PAL) activity in SAEW-treated sprouts was higher than that in the control sample throughout the whole period of germination. The contents of total phenolics, total flavonoids and rutin showed a significant correlation with values of the antioxidant capacity assayed by 3 different methods.
Effect of Fat Content on Physicochemical Properties of Milk
CHI Xuelu, TONG Lingjun, PAN Minghui, Nueraliya·ALIFU, AI Nasi, WANG Bei, SUN Baoguo
2018, 39(4):  26-31.  doi:10.7506/spkx1002-6630-201804005
Asbtract ( 323 )   HTML ( 8)   PDF (2722KB) ( 474 )  
Related Articles | Metrics
In this paper, milk samples with different fat contents (0.5%, 1.5%, 2.5% and 3.5%) (n = 12 for each fat content) were prepared and comparatively tested for their chemical composition, microstructure, particle size, color and stability. The results indicated that both the casein and total solid contents of milk were significantly increased (P < 0.05) with the increase of fat content. Yet, there was no significant difference (P > 0.05) in protein content among milk samples. The acidity of fresh milk samples was in the range of 16–18 °T, which was directly proportional?to fat content. In addition, the higher the amount of fat was, the less stable and more yellowish the milk system became. Hopefully, this study will provide basic data for exploring the relationship between fat content and physicochemical properties of milk, and provide a technical basis for the development and production process optimization of dairy products.
Moisture Migration and Pasting Profiles of Noodles during Steaming
CHEN Jie, YU Han, WANG Yuanhui, BIAN Ke
2018, 39(4):  32-36.  doi:10.7506/spkx1002-6630-201804006
Asbtract ( 367 )   HTML ( 8)   PDF (2969KB) ( 273 )  
Related Articles | Metrics
Low field nuclear magnetic resonance (LF-NMR), magnetic resonance imaging (MRI), a rapid viscosity analyzer (RVA) and an optical microscope were used to study the moisture migration, water status, pasting properties and microstructure of noodles during steaming. The correlation among moisture content, transverse relaxation time (T2), proton density (M2) and pasting properties was also analyzed. The results showed that less tightly bound water was the major component of the total moisture in steamed noodle. During steaming, moisture migrated from the surface to the interior until reaching the core. Moisture content and the proportions of less tightly bound water and tightly bound water increased until reaching a plateau; the proportion of tightly bound water showed a slower increase and the mobility of less tightly bound water gradually rose. Both peak viscosity and breakdown decreased with the increase of steaming time. The microscopic investigation showed that starch granules swelled, and became greater in size. Moreover, starch granules in the central region of steamed noodles had a lower degree of gelatinization than in the external region. Correlation analysis showed that moisture content was highly significantly correlated with T22, M22 and Mtotal (P < 0.01), and each of these was significantly (P < 0.05) or extremely significantly (P < 0.01) with peak viscosity, breakdown and final viscosity.
Effects of Hydrocolloids on the Properties of Starch Paste from Lycoris radiate Bulbs
ZHANG Fan, ZHOU Shoubiao, LI Ning, ZOU Jinyou
2018, 39(4):  37-42.  doi:10.7506/spkx1002-6630-201804007
Asbtract ( 623 )   HTML ( 4)   PDF (2699KB) ( 171 )  
Related Articles | Metrics
In order to improve the quality of starch from Lycoris radiate bulbs, the effects of different hydrocolloids on properties of starch paste, including swelling capacity, pasting properties, freeze-thaw stability and rheological properties, were studied in this paper. After adding hydrocolloids, the swelling power of starch was decreased, and the solubility of mixed systems was improved except for the addition of konjac gum. All hydrocolloids, especially xanthan gum, suppressed starch retrogradation effectively so that the freeze-thaw stability was enhanced. The pasting properties showed that addition of hydrocolloids increased the peak time and paste temperature, decreased peak viscosity, breakdown and setback, and improved the thermal stability and cold stability of starch. Static rheological studies suggested that the rheological curves could be well fitted to the Herschel-Bulkley equation and that the starch and hydrocolloid blends exhibited a typical pseudoplastic ?ow behavior. Addition of all colloids except gum arabic reduced the thixotropy of starch gels to varying degrees. In dynamic rheological experiments it was found that G’ values of all the systems were larger than their G” values and exhibited a frequency dependence, indicating a typical weak gel properties. Addition of hydrocolloids could keep the internal structure of the starch system stable and enhance its shear resistance. Therefore, hydrocolloids can improve the properties of starch paste to a certain extent, depending on the type and structure of hydrocolloids.
Effects of Clarifying Agents on Active Ingredients and Characteristics of Haskap (Lonicera caerulea L.) Juice
ZENG Deyong, ZHANG Ligang, ZHAO Yuhong
2018, 39(4):  43-48.  doi:10.7506/spkx1002-6630-201804008
Asbtract ( 297 )   HTML ( 7)   PDF (2584KB) ( 211 )  
Related Articles | Metrics
This study was designed to compare the effect of 8 different clarifying agents on active components and physicochemical properties of haskap (Lonicera caerulea) juice with naturally clarified and original juices as control. The results showed that there were significant differences in effectiveness between 8 clarifying agents. Polyvinylpyrrolidone (PVPP) showed the maximum transmittance (74.3%, P < 0.05), together with the lowestc ontents of polyphenols (0.452 g/L), flavonoids (0.035 mg/L) and anthocyanins (16.69 mg/L) in haskap juice as compared with other clarifying agents (P < 0.05). Pectinase and cellulose did not show significantly different effects on transmittance or the contents of active components (P > 0.05). Besides, no significant difference in pH was seen between juices treated with 8 clarifying agents (P > 0.05). The highest L* value of 31.50 was recorded after treatment with PVPP, while the lowest L* value of 12.50 was recorded in untreated juice (P < 0.05). Pectinase and cellulose resulted in the lowest browning index, which did not show significant difference as compared with that before treatment (P > 0.05). To sum up, various clarifying agents had different effects on active components, transmittance and other physicochemical characteristics of haskap juice. Therefore, the appropriate clarifying agent should be chosen taking into consideration various factors.
Bioengineering
Effect of Probiotic Lactobacillus plantarum on Volatile Flavor Formation in Cheddar Cheese
HAO Xiaona, ZHANG Jian, YANG Yawei, YU Zhijian, CAO Yongqiang, YANG Zhennai
2018, 39(4):  49-58.  doi:10.7506/spkx1002-6630-201804009
Asbtract ( 213 )   HTML ( 4)   PDF (2810KB) ( 168 )  
Related Articles | Metrics
This study examined the effect of probiotics on flavor formation in cheese. Lactobacillus plantarum 1-2, isolated from Tibetan kefir, was added at a level of 8.0 (lg(CFU/mL)) or 9.0 (lg(CFU/mL)) to pasteurized milk and added at 8.0 (lg(CFU/g)) to milk curds after whey draining, separately, which were then processed into Cheddar cheese. The aim was to determine the effect of inoculation methods, inoculum size and ripening time on the volatile flavor composition of cheese. By using solid phase microextraction coupled to gas chromatography-mass spectrometry (SPME-GC-MS), a total of 26 and 30 flavor compounds were detected in the control and probiotic cheeses, respectively. Addition of the probiotic L. plantarum 1-2 resulted in the formation of four unique volatile flavors, namely ethylbenzene, dodecane, hexanol and acetone. Ripening time had the greatest effect on cheese flavor. Benzene content in the probiotic cheese group was increased significantly with ripening time, but in the control group, benzene was not detected until after 12 weeks of ripening. The viable count of L. plantarum 1-2 also had a significant impact on cheese volatile flavor. Different addition methods and quantities of the probiotic strain had similar effects on the volatile flavor compounds in cheese, with the largest effect being observed on butyric acid. In the probiotic cheese group, butyric acid content reached the maximum after 12 weeks of maturation, showing a 3.96-fold increase compared with the control group. Cheeses made from pasteurized milk with different quantities of L. plantarum 1-2 added had different compositions of volatile compounds. However, addition of the probiotic strain at 9.0 (lg(CFU/mL)) to pasteurized milk and at 8.0 (lg(CFU/g)) to milk curds after whey draining had a similar effect on the composition of volatile compounds (P < 0.05). These results can provide an experimental basis for improving the processing and flavor quality of probiotic cheese.
Volatile Flavor Compounds of Fermented Grapefruit Juice with Probiotics
SHU Wenxiu, WU Zufang, LIU Lianliang, WENG Peifang
2018, 39(4):  59-65.  doi:10.7506/spkx1002-6630-201804010
Asbtract ( 341 )   HTML ( 7)   PDF (2550KB) ( 345 )  
Related Articles | Metrics
The aim of this study was to evaluate the volatile flavor compounds of grapefruit juice fermented with two isolates, Lactobacillus plantarum L1 and L. fermentum L2, which were previously isolated in our laboratory. The volatile flavor compounds were measured by static-headspace solid phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS) and analyzed by principal component analysis (PCA). The aroma compounds were detected by electronic nose. Results indicated that a total of 79 volatile flavor compounds were identified, including 20 alcohols, 15 olefins, 12 alkanes, 10 ketones, 5 aldehydes, 2 esters and 15 other compounds, 59, 36 and 35 of which were present in fermented grapefruit juices with L1, L2 and both, respectively. Fermented grapefruit juice with probiotics contained more volatile compounds than fruit grapefruit juice; some esters were detected only in the former. In addition, aldehydes accounted for a much lower percentage of the total volatiles, while alcohols, ketones and alkanes accounted for a significantly higher percentage after fermentation as compared with fresh grapefruit juice. All four samples were clearly discriminated by PCA on the basis of their characteristic flavor components. The results of electronic nose revealed fermented grapefruit juices with different Lactobacillus species to be significantly different in flavor. In conclusion, these results provide insight into the flavor characteristics of fermented grapefruit juice and also provide a basis for product quality evaluation.
Optimization of Preparation of Water Extract from Roselle and Fermentation of Roselle Wine and Variations in Organic Acids and Phenolic Acids Concentrations
LI Yuzhu, LONG Mou, TANG Yanyan, JIANG Xi, DU Muying,
2018, 39(4):  66-75.  doi:10.7506/spkx1002-6630-201804011
Asbtract ( 263 )   HTML ( 7)   PDF (4348KB) ( 292 )  
Related Articles | Metrics
The preparation of water extract from roselle grown in Yunnan and the fermentation of roselle wine were optimized using one-factor-at-a-time method and response surface methodology. Results showed that a solid-to-solvent ratio of 1:28 (g/mL), an extraction temperature of 84.0 ℃ and 78 min extraction were found to be the optimal conditions to obtain a higher yield of anthocyanins and polyphenols of (0.060 ± 0.009)% and (0.201 ± 0.05)%, respectively. The optimal fermentation conditions were as follows: temperature 24.5 ℃, yeast inoculum size 1.6 g/L, and sugar 22.5%. The wine produced under these conditions contained (10.6 ± 0.12)% alcohol (V/V), (121.25 ± 0.35) mg/L anthocyanins, and (629.58 ± 0.22) mg/L polyphenols. It also contained a variety of amino acids with a balanced amino acid composition. Seven organic acids and eight phenolic acids were identified in the water extract, with citric acid, hibiscus acid and protocatechuic acid being the major constituents. However, hibiscus acid and lactic acid were the main organic acid and protocatechuic acid was similarly the main phenolic acid in the wine. During fermentation, the contents of citric, hydroxycitric and tartaric acid decreased while the opposite was observed for pyruvic, lactic and succinic acid. In addition, an decrease in protocatechuic, gentisic and ferulic acid was observed, accompanied by an increase in gallic, coumaric and syringic acid.
Cloning, Expression and Characterization of Sortase A from Lactobacillus acidophilus ATCC4356
WU Jing, WANG Wenwen, XIONG Ronglu, WU Zhen, PAN Daodong, ZENG Xiaoqun, GUO Yuxing
2018, 39(4):  76-81.  doi:10.7506/spkx1002-6630-201804012
Asbtract ( 262 )   HTML ( 6)   PDF (3062KB) ( 146 )  
Related Articles | Metrics
Sortase (Srt) plays an important role in the adhesion of Gram-positive bacteria to host cells. The focus of this study was to recombine the sortase A (SrtA) of Lactobacillus acidophilus (Lap-SrtA) in E. coli and to evaluate the differences between Lap-SrtA with other pathogenic sortase. The srtA gene was cloned from L. acidophilus ATCC4356 and expressed in E. coli Transetta (DE3). The recombinant protein was purified and analyzed using Dabcyl-QALPTTGEE (Edans) as substrate. The results showed that the molecular weight of the recombinant protein was about 20 kDa with an Lap-SrtA activity, and the activity could be enhanced by Mg2+, Zn2+, and Mn2+. However, Ca2+ had no visible effect on the activity of SrtA, making it distinct from other pathogenic sortase. Chalcone could significantly inhibit SrtA activity. Furthermore, there were eight segments of β sheet structure in Lap-SrtA as analyzed by homology modeling using SWISS MODEL, and the active site residues of Lap-SrtA were found to be His 137, Cys 198 and Arg 205.
Effect of Lactic Acid Bacteria and Yeast on the Quality of Yoghurt during Fermentation by Tibetan Kefir
LONG Xingyao, LI Jian, XIE Jie, ZHAO Xin, WANG Hongwei, ZHANG Yu, SUO Huayi,
2018, 39(4):  82-89.  doi:10.7506/spkx1002-6630-201804013
Asbtract ( 344 )   HTML ( 8)   PDF (3187KB) ( 205 )  
Related Articles | Metrics
Tibetan kefir is a fermented milk product produced by mixed cultures of lactic acid bacteria, yeasts, acetobacter and moulds. Its quality characteristics are significantly different from those of ordinary yogurt. In order to determine the roles of lactic acid bacteria and yeast in yoghurt fermentation by Tibetan kefir, these microbes were separately inhibited by cycloheximide and penicillin during fermentation and the flavor, taste and texture of normally fermented yogurt and fermented yogurts with inhibition of lactic acid bacteria and yeasts were determined and compared. The results showed that the alcohol content of yoghurt was increased from 1.85 to 2.77 g/kg and the VB2 content was decreased from 168 to 157 μg/100 g during fermentation with yeast inhibition, which showed a significant difference compared with normal fermentation. During fermentation with inhibition of lactic acid bacteria, the acidity of yoghurt was increased from 18 to 28 °T, along with a decrease in amino acid nitrogen from 65 to 53 mg/100 g, acetic acid from 1 to 0.7 mg/g and VB1 from 20 to 18 μg/100 g, while lactic acid content was continuously maintained at 2 mg/g; all these parameters were considerably different from those of normally fermented yogurt. During yogurt fermentation, the lactic acid bacteria in Tibetan kefir made great contributions to the acidity, amino acid nitrogen, texture, lactic acid, acetic acid and VB1 while the yeasts made great contributions to alcohol and VB2.
Optimization of Fermentation Medium for Glutamate Decarboxylase Production by Enterococcus faecium by Bidirectional One-Factor-at-a-Time and Taguchi Methods
YANG Shengyuan, LI Yun
2018, 39(4):  90-98.  doi:10.7506/spkx1002-6630-201804014
Asbtract ( 204 )   HTML ( 6)   PDF (3657KB) ( 165 )  
Related Articles | Metrics
The fermentation medium for glutamate decarboxylase (GAD) production by Enterococcus faecium was optimized by bidirectional one-factor-at-a-time and Taguchi methods. The results indicated that the peptone-sucrose-beef extract (PSB) medium was found to be the most suitable for GAD production among four tested media. However, CaCl2, K2HPO4 and ammonium citrate in the PSB medium were unfavourable for GAD production. Peptone, sucrose, sodium acetate and monosodium glutamate (MSG) had significant effects on GAD production (P < 0.01), which was not significantly affected by beef extract (P > 0.10). The optimal culture medium predicted by Minitab software consisted of 15 g/L peptone, 10 g/L beef extract, 12.5 g/L sucrose, 6.0 g/L sodium acetate, 10 g/L MSG, and 1.0 g/L Tween 80. The predicted total GAD activity obtained by using the optimized medium was (399.30 ± 12.51) U, which was not significantly different from the experimental value ((384.26 ± 10.32) U). The constituents of the modified PSB medium were much fewer compared with the initial one. However, the modified PSB medium was more suitable for GAD production, in which the total GAD activity produced by E. faecium was improved by 45.71% as compared to the initial one.
Effects of Torulaspora debrueckii on Wine Aroma
YUAN Miaomiao, JIANG Kaikai, SUN Yuxia, WANG Shiping
2018, 39(4):  99-105.  doi:10.7506/spkx1002-6630-201804015
Asbtract ( 342 )   HTML ( 13)   PDF (1441KB) ( 232 )  
Related Articles | Metrics
In this paper, Cabernet sauvignon grapes from the north foot of Tianshan Mountain in Xinjiang and Rushan region in Shandong were fermented with Saccharomyces cerevisiae and Torulaspora debrueckii alone and in combination, respectively. Physicochemical properties and volatile contents of the resulting wines were analyzed. The results showed that a total of 60 volatile compounds related to the metabolism of yeasts were detected in all wines, including 23 alcohols, 26 esters, 8 acids and 3 ketones. The aroma components of wines fermented with different yeast strains were significantly different. The wine produced using Torulaspora debrueckii alone showed significantly higher contents of esters, alcohols and acids, whereas that produced by mixed culture fermentation showed lower contents of volatile acids. Torulaspora debrueckii produced a wine that was richer in ethyl esters, especially fatty acid ethyl esters such as ethyl octanoate, ethyl decanoate and ethyl dodeconoate, increasing the fruity flavor. Mixed culture fermentation could enhance the fermentation aroma of wines and increase its complexity irrespective of the geographical origin of grapes.
Dynamic Succession of Bacterial Community Structure during Fermentation Process of Pixian Bean Paste Analyzed by High Throughput Sequencing
GUAN Tongwei, XIANG Huiping, WANG Penghao, DENG Aoyu, DONG Dan, ZHAO Shunxian, ZHANG Xichao
2018, 39(4):  106-111.  doi:10.7506/spkx1002-6630-201804016
Asbtract ( 198 )   HTML ( 4)   PDF (2852KB) ( 156 )  
Related Articles | Metrics
Illumina MiSeq high-throughput sequencing technology was used to study the structure, abundance and dynamic succession of the bacterial community in Pixian bean paste during the fermentation process. A total of 731 188 effective sequences were obtained, which were classified into 11 phyla (Firmicutes, Proteobacteria, Actinobacteria, Chlorobi, Chloroflexi, Gemmatimonadetes, Fusobacteria, Cyanobacteria, Bacteroidetes, Acidobacteria and Deferribacteres) and unclassified bacteria, and these were assigned into 37 082 operational taxonomic units?(OTUs). The results showed that the bacterial community richness in Pixian bean paste was high. The species and quantities of bacteria were changed with environmental change and fermentation time. The bacterial community abundance of sample peijiao (three-month fermented hot pepper) was largely different from that of samples from five other processing stages (BZ1Y, BZ5Y, BZ10Y, HE1Y and HE5Y). Our data proved high bacterial diversity in Pixian bean paste and some novel microbial species, which need further identification. High-throughput bacterial genome sequencing revealed the presence of a total of 185 genera with the predominance of Staphylococcus, Weissella, Pediococcus, Lactobacillus, Corynebacterium and Bacillus during the entire fermentation process, which played an important role in flavor formation of Pixian bean paste.
Enzymatic Preparation and Activity of Fucosylated Oligosaccharides
WANG Fengwu, QUE Fei, KOU Lingyun, CHI Fang, WANG Ying
2018, 39(4):  112-117.  doi:10.7506/spkx1002-6630-201804017
Asbtract ( 318 )   HTML ( 8)   PDF (2947KB) ( 193 )  
Related Articles | Metrics
Fucoidanase was used to degrade fucoidan to obtain fucosylated oligosaccharides. A total of 122 experiments were designed and implemented using uniform design and orthogonal array design. A three-layer BP network model was built to simulate the hydrolysis process, and it was optimized by genetic algorithm using Matlab software. A mouse model of D-galactose-induced aging was used to test the antioxidant activities of fucosylated oligosaccharides and fucoidan. The results showed that the molecular weight of hydrolysates was changed significantly during 4 h hydrolysis at 20–40 ℃ with 0–24 U/mg of fucoidanase. On the basis of this finding, the optimization was carried out and the result showed that an enzyme dosage of 20.7 U/mg, a temperature of 26 ℃ and a hydrolysis time of 2.3 h were found to be the optimal conditions to obtain a hydrolysate with the highest antioxidant activity. The hydrolysate scavenged 85.1% 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical with a molecular weight of 18.2 kDa. The results of animal experiments showed that the antioxidant activity of fucosylated oligosaccharides were significantly higher than that of fucoidan in terms of hydroxyl free radical scavenging activity, catalase (CAT) activity and malonaldehyde (MDA) content (P < 0.05). This paper can provide experimental support for the industrial preparation of fucoidan hydrolyzate with high antioxidant activity.
Screening, Cloning and Characterization of a Novel β-Glucosidase from Soil Metagenomic Library
TANG Leli, WANG Xiaomeng, WU Xiuling, HUANG Qing, LI He
2018, 39(4):  118-124.  doi:10.7506/spkx1002-6630-201804018
Asbtract ( 622 )   HTML ( 6)   PDF (3655KB) ( 214 )  
Related Articles | Metrics
A novel β-glucosidase gene, bgl2238, was isolated from a metagenomic library by functional screening (GenBank: KU320675.1). The bgl2238 gene was sequenced and expressed in Escherichia coli BL21(DE3). The biochemical properties of the resulting protein were further examined. Amino acid sequence analysis showed that the full length of the gene was 2 238 bp, which encoded a protein of 745 amino acids. The β-glucosidase had a molecular mass of 80.67 kDa with an isoelectric point (pI) of 4.95, and it was identified as an acidic protein with stable structure and high hydrophobicity and without signal peptide sequence. Phylogenetic analysis of the bgl2238 gene indicated 58% similarity to the β-glucosidase from Bacteroides thetaiotaomicron, and it belonged to the GH3 and GH3C superfamily. bgl2238 was ligated into the vector pET-32a(+) and then transformed into Escherichia coli BL21 (DE3) for heterologous expression. The maximal activity (29.1 U/mg) of the enzyme was observed at 44 ℃ and pH 6.10 with 4-nitrophenyl-β-D-glucopyranoside (pNPG) as the substrate; furthermore it retained 70% of its activity within the temperature range of 4–50 ℃ for 4 h. The Km and Vmax values of bgl2238 were 0.296 mmol/L and 576 μmoL/(L·min), respectively. Its activity was enhanced significantly by K+, Na+, Fe2+, Mg2+, Mn2+, Ca2+, Co2+ and Ni2+ at various concentrations; among these metal ions, Fe2+ was found to have the biggest promoting effect and a relative activity up to 260% was attained at a Fe2+ concentration of 10 mmol/L. On the other hand, the presence of heavy metal ions (Ag+, Hg2+ and Cu2+) inhibited its activity. To sum up, these characteristics were beneficial for the potential application of bgl2238 in industrial cellulose degradation.
Optimization of Hydrolysis Conditions of Xylanase and α-Glucuronidase Mixture for Birch Xylan by Response Surface Methodology
DONG Yuanyuan, LI Yaxian, SHEN Yihong, XUE Yemin
2018, 39(4):  125-131.  doi:10.7506/spkx1002-6630-201804019
Asbtract ( 362 )   HTML ( 5)   PDF (4001KB) ( 184 )  
Related Articles | Metrics
Objective: This study aimed to optimize the hydrolysis of birch xylan with a mixture of recombinant thermostable xylanase (XynB) and glucuronidase (AguA) from an engineered strain of Escherichia coli. Methods: The hydrolysis conditions were optimized using combination of one-factor-at-a-time method and response surface methodology. Results: The optimized conditions were obtained as follows: substrate concentration, 4.2 g/100 mL; temperature, 80.66 ℃; pH, 7.65; and XynB/AguA dosage, 60/9 U/g, yielding a reducing sugar concentration of 17.82 mg/mL. Conclusion: The reducing sugar and xylose concentrations were respectively 17.91 mg/mL and 13.66 mg/mL after 4 h of hydrolysis with the enzyme mixture.
Effects of Diammonium Phosphate on the Fermentation Performance and Volatile Composition of Jujube Wine
YIN Xiaojie, CHEN Gang, JIAN Suping, ZHENG Shudan, HUANG Lishan, SHI Jianxin
2018, 39(4):  132-137.  doi:10.7506/spkx1002-6630-201804020
Asbtract ( 321 )   HTML ( 5)   PDF (4453KB) ( 185 )  
Related Articles | Metrics
This research studied the effects of different amounts of diammonium phosphate on the fermentation performance and volatile composition of jujube wine fermented by Saccharomyces cerevisiae. The results showed that diammonium phosphate could increase the specific growth rate and fermentation activity of yeast, shorten the fermentation time, and enrich the fruity and floral aroma components of jujube wine. GC-MS analysis showed that at a diammonium phosphate concentrate of 320 mg/L, ester concentration and odor activity value (OAV) of jujube wine were the highest; furthermore, the sensory score of the wine was 90 and it not only had a rich jujube flavor, but also possessed a pleasant floral and fruity aroma.
Component Analysis
Determination of 12 Oligosaccharides in Human Milk by Ultra-Performance Liquid Chromatography-Mass Spectrometry
CHEN Xinxin, LU Jing, LIU Lu, PANG Xiaoyang, ZHANG Shuwen, LI Cheng, Lü Jiaping
2018, 39(4):  138-143.  doi:10.7506/spkx1002-6630-201804021
Asbtract ( 279 )   HTML ( 7)   PDF (2408KB) ( 469 )  
Related Articles | Metrics
A qualitative and quantitative method for the determination of 12 oligosaccharides in human milk was established. The conditions for gradient elution were set as follows: mobile phase A, 10 mmol/L ammonium formate solution; mobile phase B, acetonitrile; flow rate, 0.3 mL/min; and column temperature, 50 ℃. The gradient elution procedure was programmed as follows: 0–10 min, 95%–75% B; 10–15 min, 75% B; 15–20 min, 75%–65% B; 20–21 min, 65%–10% B; 21–24 min, 10% B, 24–25 min, 10%–95% B; and 25–35 min, 95% B. The results showed that the standard curves of 12 oligosaccharides had good linearity within the concentration range investigated (R2 > 0.98) and the recoveries were between 80.00% and 120.00%. The repeatability and instrumental precision (expressed as elative standard deviations) were less than 10%.
Correlation Analysis between Taste Compounds and Sensory Attributes of Cherry Wine Based on Partial Least Squares Regression
NIU Yunwei, KONG Jiali, XIAO Zuobing,
2018, 39(4):  144-148.  doi:10.7506/spkx1002-6630-201804022
Asbtract ( 321 )   HTML ( 4)   PDF (2288KB) ( 233 )  
Related Articles | Metrics
In this study, six cherry wines from different producers were evaluated by sensory evaluation and 33 characteristic taste compounds were identified in these samples using high performance liquid chromatography (HPLC). Principal component analysis (PCA) and agglomerative hierarchical clustering (AHC) were used to attempt to discriminate the wines. The results demonstrated that wines from the same geographical areas were clustered into one group and samples from different geographical origins were clearly discriminated. Correlation analysis between characteristic taste compounds and sensory attributes using partial least squares regression (PLSR) indicated a significant positive association between the sweet taste and glucose, sucrose and fructose; between the astringent taste and serine and phenylalanine; and between the bitter taste and caffeic acid, glutamic acid, serine, glycine, threonine, alanine, valine, methionine, phenylalanine, isoleucine, leucine and lysine.
Nutritional Characteristics and Umami Assessment of Euphausia superba and Exopalaemon carinicauda
CAO Rong, ZHAO Ling, SUN Huihui, LIU Qi
2018, 39(4):  149-153.  doi:10.7506/spkx1002-6630-201804023
Asbtract ( 377 )   HTML ( 5)   PDF (2271KB) ( 177 )  
Related Articles | Metrics
This study aimed to scientifically evaluate the potential value of Euphausia superba as an emerging resource, and to provide useful data for the processing and utilization of small-sized shrimps into high value products. E. superba and Exopalaemon carinicauda were selected to analyze nutritional composition and umami taste characteristics. Moisture, crude protein, crude fat, ash, amino acid, and fatty acid composition were determined to analyze the nutritional characteristics. Free amino acids and flavor nucleotides were determined to calculate taste active value (TAV) and equivalent umami concentration (EUC), which indicated the umami taste. Results showed that there were significant differences in crude protein (13.25% vs. 16.41%) and fat (3.12% vs. 1.10%) contents between E. superba and E. carinicauda (P < 0.05). Their fatty acid compositions were significantly different as well. The content of unsaturated fatty acids in E. superba was relatively higher than in E. carinicauda. The total content of amino acids in E. superba protein hydrolysate was significantly lower than in E. carinicauda protein hydrolysate (P < 0.05), but the proportion of essential amino acids was higher in the former, meaning that the nutritional value of protein in E. superba was accordant with the FAO/WHO pattern. Inosine monophosphate (IMP) and glutamic acid (Glu) played a major role in contributing to the umami taste, and the EUC of E. superba and E. carinicauda was 11.01 and 12.98 g MSG/100 g respectively. Conclusively, E. superba and E. carinicauda have high nutritional values and desirable umami taste characteristics, indicating promising prospects for use as ordinary foods or condiments.
Characteristics of Phenolic Accumulation in Muscadine Seeds during Berry Development
WEI Zheng, GUO Wenfeng, HUANG Yu, YIN Ling, QU Junjie, ZHANG Yali, LU Jiang,
2018, 39(4):  154-164.  doi:10.7506/spkx1002-6630-201804024
Asbtract ( 213 )   HTML ( 3)   PDF (4042KB) ( 132 )  
Related Articles | Metrics
The characteristics of phenolic accumulation in seeds of two muscadine grape cultivars, ‘Noble’ and ‘Carlos’ at different stages of berry development were studied by using ultra performance liquid chromatography tandem triple quadrupole time-of-flight mass spectrometry (UPLC-Triple-TOF-MS/MS) in comparison with two Vitis vinifera cvs, ‘Cabernet Sauvignon’ and ‘Riesling’. A total of 106 phenolic compounds were identified and quantified in grape seeds collected from four developmental stages investigated, including 48 ellagic acids and precursors, 24 flavonols, 20 hydroxy benzoic acids, 9 flavan-3-ols, 3 stilbenes and 2 hydroxy cinnamic acids. Among them, ellagic acids, precursors (mostly precursors) and flavan-3-ols (mainly gallates and monomers) were the predominant phenolics detected in seeds of muscadine and Vitis vinifera grapes, respectively; however, there were great differences in their contents between years and among cultivars. The precursors in ‘Noble’ seeds accumulated abundantly at the young berry stage, but was decreased gradually until veraison, and increased again at the end of veraison, while for ‘Carlos’, these compounds were dominant before veraison, and then slightly decreased. The flavan-3-ols in ‘Riesling’ seeds were increased progressively with berry development in the year 2013, reaching the highest level at veraison, and declined thereafter, while their content remained constant during the growth season in 2014. For ‘Cabernet Sauvignon’, the highest content was found in young and fully ripe berries in 2013 and 2014, respectively.
Determination of Amino Acid Contents in Cereals and Ileal Digesta by Reversed-Phase High Performance Liquid Chromatography with AccQ-Tag Pre-column Derivatization Method
HAN Fenli, HAN Fei, LI Aike, CHEN Xi, SONG Ge, FAN Liuping, YU Zhiquan
2018, 39(4):  165-170.  doi:10.7506/spkx1002-6630-201804025
Asbtract ( 412 )   HTML ( 7)   PDF (2620KB) ( 209 )  
Related Articles | Metrics
The contents of amino acids in different cereals and ileal digesta were determined by reversed-phase high performance liquid chromatography (RP-HPLC) according to the AccQ-Tag method. The ratio of material to acidolysis agent was optimized in this paper. Quantification was performed using an internal standard method after pre-column derivatization with 6-aminoquinolinyl-N-hydroxysuccinimidate (AQC). The analysis procedure was carried out using a Waters 2695 HPLC system fitted with a Waters AccQ-Tag Nova-PakTMC18 column (3.9 mm × 150 mm, 4 μm) and a Waters 2475 fluorescence detector (Waters Corp., USA) at an excitation wavelength of 250 nm and emission wavelength of 395 nm at 37 ℃. Sodium acetate (pH 4.95), acetonitrile and water were used respectively as mobile phases A, B and C for gradient elution and the injection volume was 10 μL. The results showed that the optimum ratio of material to acidolysis agent was 1:100 (mg/μL). The calibration curves of 17 amino acids were linear within the range of 25?500 μmol/L (12.5–250 μmol/L for cystine) with correlation coefficients (r) of 0.999 6?0.999 9. The average recoveries were in the range of 97.56%–103.92% with relative standard deviation (RSD) of 0.31%?2.75% (n = 3). This method is rapid, simple and accurate for the determination of amino acids in cereals and ileal digesta.
Analysis of Differential EUC in Dried Fruiting Bodies of Different Cultivars of Lentinus edodes
YU Hailong, LI Yu, CHEN Wanchao, LI Wen, SONG Chunyan, SHANG Xiaodong, ZHANG Lujun, ZHANG Meiyanl, YANG Yan
2018, 39(4):  171-175.  doi:10.7506/spkx1002-6630-201804026
Asbtract ( 371 )   HTML ( 7)   PDF (2393KB) ( 173 )  
Related Articles | Metrics
The dried fruiting bodies of 12 main shiitake (Lentinula edodes) cultivars in China obtained under identical conditions of medium formulation, spawn running, coloring, fruiting and postharvest treatment were analyzed for free amino acids, 5’-nucleotides and equivalent umami concentration (EUC) by high performance liquid chromatography (HPLC). The results showed that the contents of various amino acids in different cultivars were different while the contents of total amino acids in the 939 and Shen 8 cultivars were similar. Asp was 3.7 times more abundant in 939 than in Shen 8. The content of umami amino acids in Wuxiang was the highest among the cultivars evaluated ((5.76 ± 0.13) mg/g), which was 1.2 times higher than that of 135. The 605 cultivar contained the highest content of sweet amino acids (14.35 ± 0.32) mg/g, which was 2.6 times compared with Cr04. The content of flavor nucleotides in 605 was the highest ((5.24 ± 0.27) mg/g), which was 3.3 times compared with 808. Overall, Shenxiang 10 showed the highest EUC ((342.1 ± 4.61) g MSG/100 g).
Simultaneous Determination of Twenty-Three Phenolic Acids in Cauliflower (Brassica oleracea L. var. botrytis L.) and Broccoli (B. oleracea?L. var. italica) from Five Producing?Places by Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry
MA Shuai, WANG Jihua, GAO Yuan, WANG Meng, FENG Xiaoyuan
2018, 39(4):  176-187.  doi:10.7506/spkx1002-6630-201804027
Asbtract ( 187 )   HTML ( 7)   PDF (5329KB) ( 260 )  
Related Articles | Metrics
A method for the simultaneous determination of twenty-three phenolic acids in cauliflower (Brassica oleracea L. var. botrytis L.) and broccoli (B. oleracea L. var. italica) was developed by ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The target analytes were separated on an Acquity UPLC HSS T3 column (2.1 mm × 150 mm, 1.8 μm) under a gradient elution mode using a mobile phase consisting of 0.1% formic acid and 0.1% formic acid acetonitrile. The flow rate was 0.3 mL/min and the column temperature was 45 ℃. The target compounds were analyzed in the negative ion multiple reaction monitoring (MRM) mode. Good linear relationships were obtained for all the analytes in the concentration range of 1.0–500.0 μg/L were obtained with high correlation coefficients over 0.992. The precision, repeatability and stability, expressed as relative standard deviation (RSD), were lower than 5.10%. The recoveries from spiked samples ranged from 81.32% to 100.39%, with RSD below 6.21%. The proposed method was applied to the determination of phenolic compounds in broccoli and cauliflower samples collected from five different cities. The method showed time saving, high sensitivity, and good repeatability, and was suitable for the simultaneous determination of phenolic compounds in cauliflower, broccoli and other vegetables.
Optimization of Refining and Volatile Composition Analysis of Vanilla Extract
XU Fei, LI Na, CHU Zhong, ZHANG Yanjun,
2018, 39(4):  188-193.  doi:10.7506/spkx1002-6630-201804028
Asbtract ( 217 )   HTML ( 6)   PDF (2853KB) ( 244 )  
Related Articles | Metrics
Vanilla extract was refined by organic solvent extraction. Different solvents were tested for their extractability using electronic nose and sensory evaluation. Solvent composition and number of extraction cycles were optimized for higher retention rates of major components. The volatile components before and after the extraction were compared by gas-chromatography-mass spectrometry (GC-MS). The results showed that the optimized conditions that provide maximum vanillin content (9.48%) were found to be extraction performed 3 times using a mixture (3:7, V/V) of ethyl acetate and n-butanol as extraction solvent. After the extraction, the proportion of aromatic compounds was increased from 35.58% to 51.00%, along with a decrease in the proportion of alkanes from 4.42% to 0.78%. These results can provide a theoretical basis and technical support for the comprehensive utilization of vanilla extract and product development.
Determination and Comparison of α-Carotene and β-Carotene of Plant-Derived?Foods
CUI Yajuan, KONG Fanhua, LIU Guojie, LU Fei, LI Dong
2018, 39(4):  194-198.  doi:10.7506/spkx1002-6630-201804029
Asbtract ( 400 )   HTML ( 4)   PDF (2825KB) ( 636 )  
Related Articles | Metrics
The study was conducted to develop an analytical method for simultaneous online determination of α-carotene and β-carotene in plant-derived?foods by high performance liquid chromatography (HPLC). Furthermore, the method was validated with freeze-dried carrot powder. The results showed that the calibration curves for α-carotene and β-carotene were linear in the ranges of 0.8–20 and 0.4–8.2 μg/mL, respectively. The precision expressed as relative standard deviation (RSD) were 4.45% and 4.71%, respectively. The recoveries at three spiked levels were 96%–112% and 83.5%–103% with RSD values of 2.67%–5.83% and 4.90%–7.42%, respectively. These data indicated good repeatability and accuracy. Comparative analysis of various plant-derived?foods by this method showed that β-carotene was more abundant than α-carotene in all samples except for pumpkin, which gave the opposite result.
Analysis of the Volatile Components of Fermented Hot Pepper from Different Varieties Grown in Guizhou by Electronic Nose Combined with SPME-GC-MS
LU Kuan, WANG Xueya, SUN Xiaojing, HUO Xin, GAO Yuqiong, FENG Fajin, DU Chengxing
2018, 39(4):  199-205.  doi:10.7506/spkx1002-6630-201804030
Asbtract ( 285 )   HTML ( 8)   PDF (2981KB) ( 395 )  
Related Articles | Metrics
In this research, electronic nose (E-nose) and solid phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) were used to evaluate and compare the volatile components of fermented hot pepper from different varieties grown in Guizhou. The results showed that the E-nose could distinguish among five different varieties of fermented hot pepper. In principal component analysis (PCA) and linear discriminant analysis (LDA), the cumulative contribution of the first 2 principal components accounted for greater than 95% (99.93% and 99.19% , respectively) of total variance, suggesting that the E-nose sensors had good recognition performance and discrimination between samples on the basis of their flavor characteristics was excellent with clear distinctions. A total of 124 volatile compounds were identified, 97 of which were found in fermented Huangping line pepper, with esters being the most abundant constituents (36.82%); 88 and 94 of which in fermented Shibing line pepper and Dafang wrinkle pepper, respectively, the predominant ones being alcohols (31.88% and 28.99%, respectively); and 89 and 71 of which in fermented Baiyi plane pepper and Huaxi Dangwu pepper, respectively, hydrocarbons being the major ones (31.88% and 28.99%, respectively). These results were consistent with PCA and LDA. Therefore, the electronic nose is useful for clear discrimination of fermented pepper from different varieties grown in Guizhou.
Main Physicochemical Quality Indicators of Base Spirit of Geographical Indication Product Yibin Spirit
LIU Qian, LI Hua, YANG Minmin, YANG Zhijie, CHEN Li
2018, 39(4):  206-211.  doi:10.7506/spkx1002-6630-201804031
Asbtract ( 225 )   HTML ( 7)   PDF (2755KB) ( 218 )  
Related Articles | Metrics
This study aimed to clarify the main physicochemical quality indicators of base spirit of the geographical indication product Yibin spirit for the purpose of providing a basis for the amendment of relevant standards. This investigation determined the alcohol (%, V/V), total acid and total ester contents of 524 samples and the contents of individual compounds including acetaldehyde, acetal, n-propyl alcohol, sec-butyl alcohol, isobutanol, n-butyl alcohol, isoamyl alcohol, ethyl acetate, ethyl butyrate, ethyl caproate and ethyl lactate in 267 of these samples according to the national standard (GB/T 10345-2007). Statistical analysis of data was performed with the SPSS software package, version 17.0. The results showed that the higher the alcohol content was, the greater the differences in total acids, total esters, ethyl acetate and ethyl caproate contents were. Overall, Yibin base spirit had appropriate content of total acids and high levels of total esters and ethyl caproate. A high proportion of these 524 samples were recognized as premium products according to the Technological Requirement for Solid-State Fermentation of Traditional Multi-Grain Luzhou-Flavor Liquor (Yibin liquor) (DB511500/T10-2014).
Quality and Flavor Analysis of Different Hot Pepper Varieties Grown in Guizhou
WANG Xueya, LU Kuan,, SUN Xiaojing, PENG Guihua
2018, 39(4):  212-218.  doi:10.7506/spkx1002-6630-201804032
Asbtract ( 293 )   HTML ( 11)   PDF (2827KB) ( 519 )  
Related Articles | Metrics
In this paper, the proximate nutritional composition, volatile flavor substances and texture properties of five well-known hot pepper varieties grown in Guizhou were measured. The results showed that the nutritional components of chili pepper varied among varieties. The contents of moisture, crude protein and polyphenols in Huangping line pepper were higher than those in the other varieties, as well as the contents of total ash, soluble sugar, reducing sugar and VC in Baiyi flat pepper, the contents of crude fiber and capsorubin in Huaxi Dangwu pepper, and the contents of total acid and total capsaicin in Dafang wrinkle pepper. Texture parameters showed that the hardness and resilience of Baiyi flat pepper were the highest, while the cohesiveness, adhesiveness and chewiness of Huangping line pepper were the highest among the five pepper varieties. Hardness was significantly positively correlated with resilience (r = 0.812), but negatively with cohesiveness, adhesiveness and chewiness (r = ?0.741, ?0.720 and ?0.600, respectively). There was a significant positive correlation of adhesiveness with cohesiveness and chewiness (r = 0.630 and 0.752, respectively). A total of 128 volatile components were detected in these pepper varieties, with Huangping line pepper and Shibing line pepper containing the largest number of volatile compounds (93). The major volatile compounds were terpenes (20.57%–48.60%), followed by sulfur-containing compounds (5.23%–27.02%). The types and amounts of volatile compounds in hot pepper varied greatly among varieties. Overall, the pepper varieties grown in Guizhou showed differences in the nutrient components, texture characteristics and flavor components. Baiyi flat pepper could be used as a raw material for fermented pepper. Huaxi Dangwu pepper was suitable for dry pepper processing. Huangping line pepper could be used as a fresh pepper variety. Selection of suitable pepper varieties is useful to obtain high quality pepper products.
Chemical Composition and Antioxidant Activity of Chinese Wolfberry Bee Pollen
ZHOU Wangting, MI Jia, LU Lu, LUO Qing, YAN Yamei, CAO Youlong, ZENG Xiaoxiong
2018, 39(4):  219-224.  doi:10.7506/spkx1002-6630-201804033
Asbtract ( 479 )   HTML ( 9)   PDF (3064KB) ( 190 )  
Related Articles | Metrics
In the present study, the main chemical components of Chinese wolfberry bee pollen including total soluble sugar, free mono- and disaccharides, proteins, free amino acids, total polyphenols, total flavonoids and minerals were analyzed. In addition, the antioxidant activity was evaluated by using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging, superoxide anion radical scavenging, 2,2’-azinobis-(3-ethylbenzthiazoline-6-sulphonate) cation (ABTS+·) radical scavenging and metal iron chelating assays. The results showed that the content of total soluble sugars was 44.8%, the contents of fructose, glucose and sucrose were 18.4%, 15.2% and 3.7%, respectively, protein content was 25.0%, and the contents of total polyphenols and total flavonoids were 22.95 mg gallic acid equivalent/g and 21.17 mg rutin equivalent/g, respectively. Chinese wolfberry bee pollen exhibited significant antioxidant activity. Moreover, the antioxidant activity of different extracts from Chinese wolfberry bee pollen was in the decreasing order of supernatant from ethanol precipitation of water extract > water extract > crude polysaccharides.
Speciation Analysis and Bioavailability Assessment of Selenium in Two Selenium-Enriched Foods by in Vitro Biomimetic Gastrointestinal Tract Models
CHEN Shanglong, LIU Enqi, CHEN Anhui, LIU Hui, WU Yonghua, QIN Xu
2018, 39(4):  225-232.  doi:10.7506/spkx1002-6630-201804034
Asbtract ( 256 )   HTML ( 6)   PDF (3919KB) ( 157 )  
Related Articles | Metrics
An effective method was developed for the speciation analysis of selenium in digests of Se-enriched Cordyceps militaris and selenium-enriched yeast prepared using in vitro biomimetic gastrointestinal tract models by microwave digestion-high resolution-continuum source graphite furnace atomic absorption spectrophotometry (HR-CS GFAAS). The aim was to study the influences of gastric and intestinal digestion on the speciation and bioavailability of Se in selenium-enriched food. The gastric and intestinal digests were separated through a 0.45 μm membrane filter into suspended and soluble Se, and inorganic and organic Se from the soluble fraction were further separated using D101 macroporous resin. Moreover, Se-binding protein and polysaccharide were precipitated by acetone and ethanol, respectively. The partition behaviors of Se from Se-enriched food in the stomach and gut were simulated with n-octyl alcohol and monolayer liposome as cell membrane, respectively. The optimal analysis wavelength for Se was 196.026 7 nm. A mixture of 1 g/L Pd(NO3)2 and 0.5 g/L Mg(NO3)2 (5 μL) was selected as the best matrix modifier, and an ashing temperature of 1 100 ℃ and an atomization temperature of 2 200 ℃ were used. The results showed that Se contents in selenium-enriched Cordyceps militaris and selenium-enriched yeast were determined to be 2.129 and 27.75 μg/g, respectively under these conditions. The contents of all Se forms except suspended Se in the intestinal digest were higher than in the gastric digest. n-Octyl alcohol-soluble Se and monolayer liposome-bound Se were more abundant than water-soluble Se during simulated intestinal digestion.
Effects of Different Fermentation Methods on the Quality of Wines Made from Wild Kiwifruit
CHEN Hongmei, WANG Shasha, YIN Henan, DONG Zhe, YUAN Chunlong,
2018, 39(4):  233-240.  doi:10.7506/spkx1002-6630-201804035
Asbtract ( 248 )   HTML ( 9)   PDF (3770KB) ( 348 )  
Related Articles | Metrics
The aroma compounds of different wines made from whole fruits, cloudy and clear juice of wild kiwifruit were extracted by headspace solid-phase micro-extraction (HS-SPME) and analyzed by gas chromatography-mass spectrometry (GC-MS). Additionally, physicochemical characterization and sensory evaluation were also performed to comparatively the quality of three different wines. Results showed that wines made from whole kiwifruits and cloudy juice had decreased total acid and volatile acid concentrations and consequently increased pH as well as reduced VC loss than wine from clear juice. The total aroma in wine fermented with skins was significantly higher than that in wine fermented without skins (P < 0.05), with higher levels of esters, terpenes and ketones while significantly lower levels of acids being found in the former (P < 0.05). The characteristic aroma compounds of three wines were distinct and their sensory aroma characteristics were also distinct. Kiwifruit wine fermented with skins had higher sensory scores with outstanding aroma for quality, intensity and authenticity of taste and aroma as well as a better balance of various aroma attributes than two other wines. Therefore, fermentation with skins can improve the quality of kiwifruit wines.
Processing Technology
Optimization of Process Parameters for Immobilization of Phospholipase A1 onto Macroporous Resin and Its Application for Degumming Rapeseed Oil
SHUI Longlong, CAO Lili, PANG Min, PAN Lijun, BAO Sai, LI Jinhong, JIANG Shaotong
2018, 39(4):  241-247.  doi:10.7506/spkx1002-6630-201804036
Asbtract ( 259 )   HTML ( 3)   PDF (2832KB) ( 210 )  
Related Articles | Metrics
Eight kinds of macroporous resins were comparatively used to immobilize phospholipase A1, and the best carrier was found to be D001. The optimum conditions for immobilization were as follows: pH 5.0, phospholipase dosage 1.5 mL/g and 4 h reaction, yielding an activity of immobilized enzyme as high as 665.8 U/g. The degumming of rapeseed oil with the immobilized enzyme was optimized using response surface methodology. The optimum conditions were determined as follows: immobilized enzyme dosage 1.8 g/kg, pH 5.5, reaction time 3.6 h, and reaction temperature 51 ℃. The phosphorus content of degummed oil prepared under these condition was 5.82 mg/kg. After fifth repeated use, the immobilized enzyme still retained 47.9% of its initial activity and the phosphorus content of degummed oil was 9.78 mg/kg.
Formulation Optimization of Extruded Sesame-Based Food Products Using Mixture Design and Fuzzy Evaluation
ZHANG Xun, ZHANG Lixia, LU Xin, ZHANG Yingnan, GAO Wenchao
2018, 39(4):  248-253.  doi:10.7506/spkx1002-6630-201804037
Asbtract ( 225 )   HTML ( 7)   PDF (2759KB) ( 206 )  
Related Articles | Metrics
Sesame flour, starch, soybean protein isolate (SPI) and sucrose were mixed and extruded in this study, and the formulation was optimized to improve the eating quality using fuzzy evaluation and mixture design. The results showed that there was a significant correlation of fuzzy evaluation scores with crispness and hardness (P < 0.01). Fuzzy evaluation scores could be considered as an indicator to evaluate the quality of extruded products. The effects of the four components and their interaction on the sensory quality were significant (R2 = 0.997 1, P < 0.01). The optimal combination that provided maximum fuzzy evaluation score (89.4 ± 0.86) was starch 65%, sesame flour 12.5%, SPI 8% and sucrose 14.5%. There was no significant difference between the sensory evaluation and the predicted value (P > 0.05). Scanning electron microscope (SEM) images showed that the surface of the product was structurally compact, even and smooth and without obvious bulges. These results can provide a basis for the comprehensive utilization of sesame and the development of new products.
Optimization of Extraction and Antitumor Activity of Polysaccharide from Tanyang Congou Black Tea
SUN Sujun, JI Haiyu, BAI Yun, YU Juan, FENG Yingying, DONG Xiaodan, LIU Anjun
2018, 39(4):  254-260.  doi:10.7506/spkx1002-6630-201804038
Asbtract ( 234 )   HTML ( 7)   PDF (3156KB) ( 177 )  
Related Articles | Metrics
Optimization of the extraction of polysaccharide from Tanyang Congou black tea for higher polysaccharide yield was carried out using one-factor-at-a-time method and response surface methodology. The optimal conditions that provided maximum polysaccharide yield (10.52%) were found as follows: temperature, 60 ℃; extraction time, 70 min; and solvent-to-solid ratio, 30:1 (mL/g). The purified polysaccharide (76% purity) contained uronic acid and pyranose rings, and consisted of arabinose (34%), galacturonic acid (30%), glucose (14%), galactose (13%), fucose (5%), and rhamnose (4%). Furthermore, the polysaccharide showed a remarkable inhibitory effect on the growth of solid tumors in H22-bearing BALB/c mice and protected spleen and thymus against tumor damage. These results provide an experimental basis for the extraction and antitumor activity evaluation of polysaccharide from black tea.
Optimization of Maillard Reaction Conditions for Enzymatic Hydrolysate of Duck Bone Protein
HAN Keyan, HUANG Jichao, LIU Dongmei, ZHOU Xinghu, HUANG Ming
2018, 39(4):  261-267.  doi:10.7506/spkx1002-6630-201804039
Asbtract ( 279 )   HTML ( 9)   PDF (2766KB) ( 213 )  
Related Articles | Metrics
For the purpose of finding the optimal conditions for Maillard reaction of the enzymatic hydrolysate of duck bone protein, this study aimed to determine the effect of temperature, time and pH on browning degree and flavor of Maillard reaction products (MRPs). Firstly, experiments were designed and conducted using one-factor-at-a-time approach and the results were statistically analyzed by using the Friedman test. Then optimization of the reaction conditions was carried out using an orthogonal array design. The overall flavor profile was evaluated using a sensory panel based on fuzzy mathematics. Furthermore, the volatile flavor compounds of MRPs were determined by solid phase microextraction (SPME) combined with gas chromatography-mass spectrometry (GC-MS) to validate the optimized conditions. The optimal conditions were found to be reaction at 115 ℃ and pH 7.0 for 60 min. The reaction product obtained under these conditions contained a variety of volatile flavor components, including aldehydes, ketones, alkanes, thiophenes, thiazoles and esters. This study can provide a useful guidance for the industry production of high quality meat-flavor essence.
Optimization of Subcritical Butane Extraction and Chemical Composition of Wheat Bran Oil
ZHANG Zhiguo, TIAN Xin
2018, 39(4):  268-274.  doi:10.7506/spkx1002-6630-201804040
Asbtract ( 222 )   HTML ( 6)   PDF (2901KB) ( 191 )  
Related Articles | Metrics
The subcritical butane extraction of wheat bran oil was optimized using response surface methodology with a Box-Behnken design. The optimum extraction conditions were determined as follows: extraction temperature 40 ℃, solvent-to-solid ratio 1:3 (g/mL), extraction time 30 min, and 3 extraction cycles. The chemical composition of the oil extracted under these conditions was detected by gas chromatography-mass spectrometry (GC-MS); qualitative analysis was performed by comparing the mass spectra of analytes with the standard spectra in a mass spectral database and quantitative analysis by area normalization method. A total of 23 components were identified, including 17 fatty acid esters (78.17%) and 5 sterol compounds (19.35%). The main fatty acid methyl esters were methyl ester (11.22%), methyl linoleate (41.77%), methyl oleate (13.36%), methyl stearate (2.87%) and methyl 9-eicosanoate (1.99%). The major sterol compounds were β-sitosterol (8.13%), stigmastanol (4.73%), ergostanol (3.42%) and campesterol (2.30%).
Optimization of Extraction and Antioxidant Activity of Polysaccharides from Cordyceps cicadae by Response Surface Methodology
SONG Jiamin, WANG Hongfei, SUN Meng, WANG Kaikai, XU Feng, SHAO Xingfeng, LI Hesheng
2018, 39(4):  275-281.  doi:10.7506/spkx1002-6630-201804041
Asbtract ( 426 )   HTML ( 13)   PDF (3168KB) ( 442 )  
Related Articles | Metrics
The extraction of water-soluble polysaccharides (WSP) from Cordyceps cicadae was optimized using one-factor-at-a-time method and response surface methodology. Solvent-to-solid ratio, extraction time and temperature were used as independent variables while polysaccharide yield was taken as response variable. The in vitro antioxidant activity of WSP was investigated by measuring total reducing power and scavenging activities against 1,1-diphenyl-2-trinitrophenylhydrazine (DPPH), hydroxyl and superoxide anion radicals. The optimum extraction conditions were found to be extraction at 80 ℃ for 130 min with a solvent-to-solid ratio of 50:1 (mL/g). Experiments conducted under these conditions gave a polysaccharide yield of 26.14 mg/g. WSP possessed strong antioxidant activity with half inhibitory concentration (IC50) for scavenging of DPPH, hydroxyl and superoxide anion radicals 28.99 μg/mL, 0.19 mg/mL and 0.30 mg/mL, respectively.
Optimization of Base Seasoning Preparation from Decapterus maruadsi Based on Electronic Nose and Electronic Tongue
CHEN Xiaoting, WU Jingna, LU Haixia, LIU Zhiyu,, CHEN Yihui
2018, 39(4):  282-289.  doi:10.7506/spkx1002-6630-201804042
Asbtract ( 217 )   HTML ( 6)   PDF (3333KB) ( 214 )  
Related Articles | Metrics
This study aimed to optimize the enzymatic hydrolysis of Decapterus maruadsi muscle to produce base seasoning with improved taste and flavor. Optimization of hydrolysis conditions was done using combination of one-factor-a-time method and response surface methodology. The taste and aroma profiles of protein hydrolysates were evaluated using an electronic nose, an electronic nose and a sensory panel, and the amino acid composition and the content of disodium 5’-ribonucleotide, a?taste-active?compound were determined. A solid-to-solvent ratio of 1:2 (g/mL), an enzyme dosage of 8 000 U/g, a temperature of 60 ℃ and a hydrolysis time of 6.4 h were found to be the optimal conditions to obtain a higher degree of hydrolysis of 27.52% using papain. The protein hydrolysate prepared using the optimized conditions had a good flavor, with essential amino acids and umami plus sweet amino acids accounting for 57.93% and 20.68% of the total amino acids, respectively, and its disodium 5’-ribonucleotide content was 3.47%, suggesting good nutritional value and quality. The hydrolysate possessed a unique flavor of Decapterus maruadsi. Hopefully, this study will provide theoretical foundation for further development of Decapterus maruadsi into high value products.
Safety Detection
Shake-and-Read Test Strips for Discriminating Synthetic Food Colorants from Natural Colorants in Soft Drinks
WU Hongjing, WANG Weiya, YE Weijia, YANG Puning, HUANG Zhibing, TU Zhui
2018, 39(4):  290-295.  doi:10.7506/spkx1002-6630-201804043
Asbtract ( 616 )   HTML ( 6)   PDF (1596KB) ( 218 )  
Related Articles | Metrics
A test strip method was used to detect three synthetic colorants commonly used in soft drinks. Under the optimal conditions, the entire detection procedure was completed within 10 min, and the limits of detection (LODs) of three synthetic food colorants (SFCs), namely, Ponceau 4R (E124), Tartrazine (E102), and Sunset Yellow (E110), were 0.4, 0.3, and 0.4 mg/L, respectively. The proposed method was compared with thin-layer chromatography and high-performance liquid chromatography. The results of validation with 14 commercial samples indicated the reliability and sensitivity of the strip test method to the presence of the three SFCs in soft drinks. This study indicated that the polyamide film-based test strip exhibited the potential to monitor illegal or improper addition of SFCs.
Recognition of Beef Adulterated with Pork Using Electronic Nose Combined with Statistical Analysis
ZHANG Juan, ZHANG Shen, ZHANG Li, WANG Qian, DING Wu
2018, 39(4):  296-300.  doi:10.7506/spkx1002-6630-201804044
Asbtract ( 216 )   HTML ( 8)   PDF (2412KB) ( 170 )  
Related Articles | Metrics
Adulterated beef mixed with pork was qualitatively and quantitatively studied by using an electronic nose combined with statistical analysis. The feature values were extracted by cluster analysis and mean value method. Principal component analysis (PCA) and linear discriminant analysis (LDA) were used for qualitative analysis. Quantitative models were established by partial least squares (PLS), multivariate linear regression (MLR) and BP neural network (BPNN) to quantitatively predict pork adulteration in beef. The results showed that the characteristic value extracted by cluster analysis could reflect the response signal of the electronic nose more comprehensively, while LDA was more suited for qualitatively detecting adulterated beef. The correct classification rates for the training and verification sets were 98.8% and 97.4%, respectively in the multi-layer perceptron neural network analysis, indicating that the classification results are good. The coefficient of determination (0.999 3 and 0.993 0) and root mean square error (0.90% and 2.50%) of the BPNN model were significantly better than those of the other models. Thus, the BPNN model allowed better prediction of the content of pork in adulterated beef. These results conclusively show that an electronic nose is an feasible approach for the detection of adulterated beef mixed with pork.
Distribution and Metabolism Patterns of Sulfamethoxazole and Sulfadiazine in Yellow Grouper (Epinephelus awoara) Tissues
DENG Jianchao, DING Junwei, YANG Xianqing, LI Laihao, HU Xiao, WU Yanyan, CHEN Shengjun, YANG Shaoling, ZHAO Qingzhi
2018, 39(4):  301-306.  doi:10.7506/spkx1002-6630-201804045
Asbtract ( 227 )   HTML ( 4)   PDF (2247KB) ( 107 )  
Related Articles | Metrics
In this study, yellow grouper (Epinephelus awoara) were administered with a single dose (200 mg/kg) of sulfamethoxazole (SMZ) and sulfadiazine (SDZ) incorporated in the diet to investigate the distribution and elimination patterns of SMZ and SDZ in various fish tissues. Both sulfonamides were detected by high performance liquid chramotography-tandem mass spectrometry (HPLC-MS/MS) and quantified by the internal standard method. The results showed that SMZ concentrations in fish tissues were in the following order: liver 827.97 (μg/kg) > muscle (776.70 μg/kg) > blood (610.29 μg/L) > kidney (432.14 μg/kg) > gill (345.18 μg/kg), while SDZ concentrations in fish tissues were in the following order: liver (895.30 μg/kg) > muscle (660.55 μg/kg) > gill (431.88 μg/kg) > blood (419.56 μg/L) > kidney (310.67 μg/kg). The elimination half-life of SMZ in fish tissues was in the following order: kidney (26.65 h) > gill (21.00 h) > muscle (20.38 h) > blood (18.73 h) > liver (16.90 h), while that of SDZ in fish tissues was in the following order: kidney (31.50 h) > blood (27.72 h) > gill (24.75 h) > muscle (21.66 h) > liver (18.24 h). The elimination half-lives of two sulfonamides in liver were much shorter than all other tissues and they were metabolized at a faster rate in liver. On the contrary, the sulfonamides showed the longest elimination half-life and were metabolized at the fastest rate in kidney among the tissues tested. After simultaneous administration of single-dose SMZ and SDZ to fish living at (25 ± 2) ℃, the withdrawal period should last at least 3 days. In conclusion, our study provides a theoretical and experimental basis for reasonable application of SMZ and SDZ in fish farming.
Temporal and Spatial Distribution Characteristics of Mold Communities during Storage of Corn as Studied by Denaturing Gradient Gel Electrophoresis
YUE Xiaoyu, ZHANG Hua, CHEN Weifeng, ZOU Jian, LI Xin, YANG Na
2018, 39(4):  307-311.  doi:10.7506/spkx1002-6630-201804046
Asbtract ( 206 )   HTML ( 7)   PDF (2389KB) ( 171 )  
Related Articles | Metrics
In this paper, denaturing gradient gel electrophoresis (DGGE) was used to research the community composition of molds in corn grains stored for different years at different spatial locations in the grain depot. The aim of this investigation was to understand the pattern of mold contamination during grain storage for the purpose of reducing mold and mycotoxin contamination and improving the safety of grains. The results revealed that Penicillium, Aspergillus and Mucor were the main molds found in stored corn. The mold community composition was closely correlated with storage duration but less correlated with spatial location in the depot. The corn samples stored for 1 year and 3 years showed great differences in mold community composition, while storage for 2 years were in a period of transition. The results of this study can provide a theoretical basis and useful data to construct an accurate model for predicting mold communities in stored grain.
An Universal Quantitative Detection Method for Genetically Modified Soybean Event MON89788 Using Duplex Digital PCR
LIU Jin, LI Ting, XIAN Yuyin, WU Xiyang, LING Li, GAO Dongwei
2018, 39(4):  312-319.  doi:10.7506/spkx1002-6630-201804047
Asbtract ( 250 )   HTML ( 6)   PDF (2686KB) ( 246 )  
Related Articles | Metrics
An universal quantitative detection method for genetically modified (GM) soybean event MON89788 using duplex digital PCR (dPCR) with satisfying specificity, stability and sensitivity was established in this paper. This method could quantify both the endogenous and exogenous genes in a single reaction system and was demonstrated to be suitable for both droplet digital PCR (ddPCR) and chip digital PCR (cdPCR) platforms. The method then was used to qualify blind samples by a food analysis performance assessment scheme (FAPAS) test. The absolute limits of quantitation (LOQs) of ddPCR for MON89788 event-specific gene and lectin endogenous gene were 8.0 copies/μL and 8.2 copies/μL, respectively, while those of cdPCR were 7.443 copies/μL and 7.646 copies/μL, respectively. The relative LOQs of MON89788 were both 0.1% by ddPCR and cdPCR.
Development of a Bioluminescence Method Combined with ATP Amplification for Detection of Bacteria in Dairy Products
CHANG Chao, WANG Ling, WU Jin’e,
2018, 39(4):  320-324.  doi:10.7506/spkx1002-6630-201804048
Asbtract ( 223 )   HTML ( 5)   PDF (2376KB) ( 133 )  
Related Articles | Metrics
A bioluminescence method combined with adenosine triphosphate (ATP) amplification through regeneration from pyrophosphate (PPi) was developed to detect bacteria in dairy products. Experimental conditions were optimized by?one-factor-at-a-time method. The sensitivity, precision and accuracy of this method were studied. The results showed that adenosine phosphosulfate (APS) concentration of 10 μmol/L, ATP sulfurylase (ATPS) activity of 0.15 U/mL and pH of 7.8 were found to be the optimum conditions. The method showed a good linearity within the range of 102–107 CFU/mL with lowest limits of detection (LODs) of 10?17 mol/mL, 102 CFU/mL and 102 CFU/mL for ATP, Escherichia coli and Pseudomonas aeruginosa, respectively. The recoveries of E. coli in dairy at three spiked levels ranged from 81.33% to 97.78%, with coefficients of variation between 14.24% and 22.17%. Good correlation between this method and the standard plate count method was obtained using spiked dairy samples. The proposed method is fast, simple, sensitive, stable, and thus suited for fast detection of bacteria in dairy products.
Determination of Bisphenol Compounds in Infant Formula by QuEChERS-SPE Rapid Sample Preparation and Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry
WANG Shengli, ZHONG Guowei
2018, 39(4):  325-330.  doi:10.7506/spkx1002-6630-201804049
Asbtract ( 264 )   HTML ( 4)   PDF (2751KB) ( 251 )  
Related Articles | Metrics
A robust analytical method was established for the determination of eight bisphenol compounds in infant formula by using quick, easy, cheap, effective, rugged, safe-solid phase extraction (QuEChERS-SPE) for sample preparation and ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) for compound identification and quantitation. Bisphenol compounds from samples were extracted with acetonitrile, and the extract was subjected to sequential QuEChERS-SPE treatment to remove background interferences. The matrix-matched standard calibration method was used for quantitation. The results showed great reduction of the matrix effect with corrected matrix factor ranging 0.80–1.20 for the 8 bisphenols. within the linear range, the correlation coefficients (R2) of the calibration curves for the compounds were greater than 0.999. The method recovery was 50.2%?116% (n = 6) with relative standard deviations of 0.5%?14%. The limits of detection (LODs) and the limits of quantification (LOQs) were 0.017?0.78 and 0.055?2.6 μg/kg for milk powder, and 0.001 1?0.055 and 0.003 5?0.18 μg/L for liquid milk, respectively. The method was applied to different brands of infant formulas in Chinese and Canadian markets. Based on measured concentrations and infant daily consumption of milk, the potential dietary intakes of BPA, BPF and BPS (calculated from the highest concentrations) were estimated to be 0.065, 0.050, and 0.005 9 μg/(kg·d), respectively. The estimated intakes did not exceed the tolerable daily intake (4 μg/(kg·d)) set by the European Food Safety Association.