α-淀粉酶产生菌的分离、筛选及鉴定

通过富集培养、平板筛选、摇瓶复筛从40 ℃的温泉底泥中分离筛选出一株产α-淀粉酶菌株，通过菌株的生理生化试验、16S rRNA基因序列遗传分析对该菌株进行鉴定。结果表明，菌株FA4被鉴定为苏云芽孢杆菌（Bacillus thuringiensis），该菌株具有较强的热稳定性，在55 ℃下仍能生长。     

Sources, properties and suitability of new thermostable enzymes in food processing

Investigations concerning recombinant a-amylases from Pyrococcus woesei and thermostable a-glucosidase from Thermus thermophilus indicate their suitability for starch processing. Furthermore, the study of recombinant ss-galactosidase from Pyrococcus woesei suitable for purpose of low lactose milk and whey production are also presented. The activity of this enzyme in a wide pH range of 4.3-6.6 and high thermostability suggests that it can be used for processing of dairy products at temperatures which restrict microbial growth during a long operating time of continuous-flow reactor with an immobilized enzyme system. Preparation of recombinant a-amylase and ss-galactosidase was facilitated by cloning and expression of genes from Pyrococcus woesei in Escherichia coli host. Satisfactory level of recombinant enzymes purification was achieved by thermal precipitation of native proteins originated from Escherichia coli. The obtained a-amylase has maximal activity at pH 5.6 and 93 degrees C. The half-life of this preparation (pH 5.6) at 90 degrees C and 110 degrees C was 11 h and 3.5 h, respectively, and retained 24% of residual activity following incubation for 2 h at 120 degrees C. An advantageous attribute of recombinant a -amylase is independence of its activity and stability on calcium salt. a-Glucosidase from Thermus thermophilus also not require metal ions for stability and retained about 80% of maximal activity at pH range 5.8-6.9. Thus, this enzyme can be used together with recombinant a-amylase.     

Nutrient digestibility,ruminal fermentation,and milk yield in dairy cows fed a blend of essential oils and amylase

Two experiments were carried out to evaluate a blend of essential oils (EO) combined with amylase as an alternative to ionophores and its potential for reducing the use of antibiotics in the dairy industry. In experiment 1, 8 rumen-cannulated Holstein cows (576 ± 100 kg of body weight, 146 ± 35 d in milk, and 35.1 ± 4.0 kg/d of milk yield at the start of the experiment) were assigned to a 4 × 4 Latin square experiment with 21-d periods to determine the influence of feed additives on total apparent digestibility of nutrients, ruminal fermentation, N utilization, microbial protein synthesis, blood glucose and urea concentrations, and milk yield and composition in dairy cows. Treatment sequences assigned to cows in each block included no feed additives (control; CON); monensin (MON) added at 13 mg/kg of diet dry matter (DM); a blend of EO supplemented at 44 mg/kg of diet DM; and EO treatment combined with α-amylase at 330 kilo novo units/kg of diet DM (EOA). Differences among treatments were studied using orthogonal contrasts as follows: CON versus feed additives (MON, EO, and EOA), MON versus EO and EOA, and EO versus EOA. No differences were detected in nutrient intake and digestibility in cows. In general, feed additives decreased ruminal NH₃-N concentration of cows, notably when diet was supplemented with MON. Furthermore, feed additives increased ruminal concentrations of acetate, butyrate, and branched-chain fatty acids. Cows fed treatments containing EO and EOA exhibited lower pH, higher NH₃-N, and a trend to greater total volatile fatty acid concentration in the ruminal fluid compared with cows fed MON. Treatments containing EO increased ruminal butyrate concentration compared with MON. No treatment × time interaction effect was observed on ruminal fermentation measurements. Cows fed diets supplemented with feed additives had greater efficiency of N transfer into milk (milk N:N intake), whereas cows fed EOA exhibited greater N transfer into milk than those fed EO. Treatments had no effect on milk yield and composition, but feed additives increased the milk yield efficiency (milk yield divided by dry matter intake), whereas treatments containing EO had similar milk yield efficiency compared with MON. For experiment 2, 30 multiparous Holstein cows (574 ± 68 kg of body weight, 152 ± 54 d in milk, and 30.9 ± 4.1 kg/d of milk yield at the start of the experiment) were enrolled to a randomized complete block design experiment. The MON, EO, and EOA treatments were randomly assigned to cows within blocks (n = 10), and feed additives were provided throughout a 9-wk period. No differences were found in nutrient intake and digestibility, but cows fed EOA tended to exhibit greater dry matter intake than those fed EO. Blood metabolites and milk production were not affected by treatments. However, cows fed MON or EOA had greater milk protein content than those cows fed treatments containing EO. Feeding EO with or without amylase had similar response to feeding MON in terms of feed intake and milk yield, with a small negative effect on milk protein yield when feeding EO alone. Feed additives increased the concentrations of acetate, butyrate, and branched-fatty acids in ruminal fluid, whereas treatments containing EO had greater ruminal butyrate and NH₃-N concentrations. Therefore, either EO or EOA can replace MON in diets of dairy cows while maintaining performance.     

Polyphenol nanoparticles of millet,rice and wheat: extraction,identification, functional and morphological characteristics

Polyphenols complexes were extracted from Common millet (CM), Long-grain rice (LGR) and Huaihe wheat (HW), purification method, components and functional characteristics were studied. Higher phenol content (18.7 in CM, 12.9 in LGR and 22.3 mg g⁻¹ in HW) was obtained by the high-speed shear, with high antioxidants (DPPH 89.2% in HW, ABTS 81.6% in LGR, FRAP 0.83 in HW) after purified. Thirteen typical polyphenol complexes were identified from 21 peaks by UHPLC-LTQ-Orbitrap-MS/MS. Gingerenone A was first detected in CM, HW and LGR; (+/−)-Gingerol was first detected in CM and HW. Morphology by TEM displayed a rough spherical structure with 200–500 nm. Peculiarly contained hydroxyl polyphenol (5-(3,4-Dihydroxyphenyl) valeric acid, Apigenin-8-C-xylosyl-6-C-glucoside), polyphenols with B-rings (Trans-cinnamic acid, Gingerenone A) and the highest contents (+/−)-Gingerol with hydroxyl and Sinapinic acid with B-rings, CM had a higher α-amylase inhibition rate (80.5%) than LGR and HW, which providing a foundation for the development of hypoglycaemic drugs.     

Production,milk fatty acid profile,and nutrient utilization in grazing dairy cows supplemented with ground flaxseed

Flaxseed has been extensively used as a supplement for dairy cows because of its high concentrations of energy and the n-3 fatty acid (FA) cis-9,cis-12,cis-15 18:3. However, limited information is available regarding the effect of ground flaxseed on dry matter intake (DMI), ruminal fermentation, and nutrient utilization in grazing dairy cows. Twenty multiparous Jersey cows averaging (mean ± standard deviation) 111 ± 49 d in milk in the beginning of the study were used in a randomized complete block design to investigate the effects of supplementing herbage (i.e., grazed forage) with ground corn-soybean meal mix (control diet = CTRL) or ground flaxseed (flaxseed diet = FLX) on animal production, milk FA, ruminal metabolism, and nutrient digestibility. The study was conducted from June to September 2013, with data and sample collection taking place on wk 4, 8, 12, and 16. Cows were fed a diet formulated to yield a 60:40 forage-to-concentrate ratio consisting of (dry matter basis): 40% cool-season perennial herbage, 50% partial total mixed ration, and 10% of ground corn-soybean meal mix or 10% ground flaxseed. However, estimated herbage DMI averaged 5.59 kg/d or 34% of the total DMI. Significant treatment by week interactions were observed for milk and blood urea N, and several milk FA (e.g., trans-10 18:1). No significant differences between treatments were observed for herbage and total DMI, milk yield, feed efficiency, concentrations and yields of milk components, and urinary excretion of purine derivatives. Total-tract digestibility of organic matter decreased, whereas that of neutral detergent fiber increased with feeding FLX versus CTRL. No treatment effects were observed for ruminal concentrations of total volatile FA and NH₃-N, and ruminal proportions of acetate and propionate. Ruminal butyrate tended to decrease, and the acetate-to-propionate ratio decreased in the FLX diet. Most saturated and unsaturated FA in milk fat were changed. Specifically, milk proportion of cis-9,cis-12,cis-15 18:3, Σn-3 FA, and Σ18C FA increased, whereas that of cis-9,cis-12 18:2, Σn-6 FA, Σ odd-chain FA, Σ<16C FA, and Σ16C FA decreased with feeding FLX versus the CTRL diet. In conclusion, feeding FLX did not change yields of milk and milk components, but increased milk n-3 FA. Therefore, costs and industry adoption of premiums for n-3-enriched milk will determine the adoption of ground flaxseed in pasture-based dairy farms.     

具有α-葡萄糖苷酶和二肽基肽酶IV抑制作用降糖益生菌的筛选

以实验室保藏的13 株乳酸杆菌为研究对象,研究其细胞代谢物（cell-free excretory supernatants,CFS）和细胞内容物（cell-free extracts,CFE）对α-葡萄糖苷酶和二肽基肽酶IV（dipeptidyl peptidase IV,DPP-IV）的抑制活性;然后对酶抑制率较高的菌株进行益生特性评价,以期筛选出具有潜在降糖作用的益生菌。结果表明,13 株乳酸杆菌的CFS对α-葡萄糖苷酶具有一定的抑制作用,抑制率为0%～12.13%;CFE对α-葡萄糖苷酶无抑制作用。13 株乳酸杆菌CFS和CFE对DPP-IV均表现出一定的抑制作用,抑制率分别为0%～7.13%和0%～55.42%。选取酶抑制率较高的嗜酸乳杆菌KLDS1.1003、KLDS1.0901和KLDS1.0902进行益生特性的研究。其中嗜酸乳杆菌KLDS1.0901表现出较高的酸耐受性,嗜酸乳杆菌KLDS1.1003表现出较高的胆盐耐受性和疏水性。主成分分析表明嗜酸乳杆菌KLDS1.1003的综合性能最佳：其对α-葡萄糖苷酶的抑制活性可达10.29%;对DPP-IV的抑制率为CFS 7.13%,CFE 50.14%;于pH 2.0的条件下孵育1 h后,存活率达到51.55%;在0.3%的胆盐条件下孵育,滞后时间为2.26 h;在二甲苯、乙酸乙酯和氯仿溶剂中的疏水率分别为134.33%、20.51%和344.08%,总体上嗜酸乳杆菌KLDS1.1003具有较高的酶抑制活性和良好的益生特性。     

Major triterpenes,cycloeucalenone and 31-norcyclolaudenone as inhibitors against both α-glucosidase and α-amylase in banana peel

The inhibitory activities of banana peel extract against carbohydrate digestive enzymes were investigated. Cycloeucalenone and 31-norcyclolaudenone were obtained by bioassay-guided fractionation as effective inhibitors of α-glucosidase and α-amylase. The structure–activity relationships of four triterpenes isolated from banana peel, cycloeucalenone, 31-norcyclolaudenone and cycloeucalenol and its isomer were investigated. The IC₅₀ values of cycloeucalenone and 31-norcyclolaudenone against α-glucosidase were 31.83 ± 2.46 μm and 38.85 ± 1.54 μm , respectively, and their IC₅₀ values against α-amylase were 20.33 ± 0.59 μm and 27.63 ± 0.83 μm , respectively. In contrast, cycloeucalenol and its isomer had insufficient inhibitory activity against either enzyme. The primary active sites of cycloeucalenone and 31-norcyclolaudenone are the carbonyl group at C-3 and the double bond in the side chain. Cycloeucalenone induced a parabolic mixed-type inhibition with a K_i value of 73.86 μm in the α-glucosidase inhibitory assay. This study provides new evidence for the potential use of banana peel triterpenes as antidiabetic agents.     

Fatty Acid Composition,Total Sugar Content and Anti-Diabetic Activity of Methanol and Water Extracts of Nine Different Fruit Tree Leaves Collected from Mediterranean Region of Turkey

In this research, the total sugar content, fatty acid compositions, and α-amylase and α-glucosidase inhibitory activity of methanolic and water extracts of nine different fruit tree leaves were determined. α-amylase and α-glycosidase inhibitory activity were determined by using Caraway-Somogyi iodine/potassium iodide and 4-nitrophenyl-α-D-glucopyranoside as substrate, respectively. The total sugar content of the nine different fruit tree leaves varies from 281.02 mg GE/g (glucose equivalents) to 643.96 mg GE/g. Methanolic extract from avocado leaves had the strongest in α-amylase and α-glucosidase inhibitory activity, 69.21 and 96.26%, respectively. Fatty acid composition of nine fruit tree leaves was characterized by gas chromatography and 24 components were identified. Among the tested fruit tree leaves, the main component was linolenic acid (49.09%). The level of essential fatty acids are over 50% in mulberry, grape, and loquat leaves. Polyunsaturated fatty acids were the major group of fatty acids present in oils of mulberry, fig, pomegranate, grape, and loquat leaves. Therefore, these oils can be considered as a good source of polyunsaturated fatty acids. Furthermore, avocado can be regarded as a new source for diabetic therapies.     

Inhibition of the in vitro activities of α-amylase, α-glucosidase and pancreatic lipase by yellow field pea (Pisum sativum L.) protein hydrolysates

The aim of this work was to produce yellow field pea protein-derived peptides as inhibitors of α-amylase, α-glucosidase and pancreatic lipase activities. A pea protein concentrate was hydrolysed with alcalase, chymotrypsin, pepsin or trypsin and the hydrolysates separated into different fractions (<1, 1–3, 3–5, 5–10 kDa) by membrane ultrafiltration. Peptide sequence analysis showed that the alcalase hydrolysate had higher levels of di- and tripeptides when compared with the chymotrypsin, pepsin and trypsin hydrolysates. The peptide fractions inhibited α-amylase and α-glucosidase activities at levels that were similar to the unfractionated hydrolysates. The peptides were more active against α-amylase (inhibition at μg level) than α-glucosidase (mg level). In contrast, the fractionated peptides had reduced ability (IC₅₀ >4.2 mg mL⁻¹) when compared with the unfractionated hydrolysate (IC₅₀ <4.2 mg mL⁻¹) to inhibit lipase activity. Enzyme kinetic studies revealed that the peptides reduced α-amylase activity through competitive inhibition. However, inhibition of α-glucosidase activity was non-competitive.     

Effect of dietary flax seed and oil on milk yield,gross composition,and fatty acid profile in dairy cows: A meta-analysis and meta-regression

Several experiments were conducted over the past few years to evaluate the feeding value of flax seed and oil in dairy cow diets. The current meta-analysis and meta-regression was undertaken to assess the overall effect of different forms of flax, as a source of trienoic (cis-9,cis-12,cis-15 18:3) fatty acids (FA), on lactation performance and on transfer efficiency of its constituent n-3 FA from diet to milk fat. Comparisons were first conducted with nonsupplemented controls or with diets containing either saturated (mainly 16:0 or 18:0 or both), monoenoic (mainly cis-9 18:1), or dienoic (mainly cis-9,cis-12 18:2) FA. Results indicate that supplementing flax seed and oil decreased dry matter intake, as well as actual and energy-corrected milk yield without affecting the efficiency of utilization of dietary dry matter or energy as compared with nonsupplemented iso-energetic controls. When compared with the other 3 types of dietary fat evaluated, flax rich in trienoic FA supported a yield of energy-corrected milk similar to supplements rich in saturated, monoenoic, or dienoic FA. Greater milk fat concentration and feed efficiency were observed with saturated supplements. However, milk fat concentration and yield were lower with dienoic FA than with flax supplements. Further analyses were conducted to compare the effect of different forms of flax oil, seed, or fractions of seed. Among the 6 categories evaluated, mechanically processed whole seed (rolled or ground) allowed the greatest yield of energy-corrected milk and the best feed efficiency when compared with free oil, intact or extruded whole seed, protected flax, and flax hulls. Feeding protected flax and flax hulls allowed the greatest milk fat concentration of cis-9,cis-12,cis-15 18:3. Moreover, the greatest transfer efficiencies of this fatty acid from diet to milk were recorded with the same 2 treatments, plus the mechanically processed whole seed. These results make this last category the most suitable treatment, among the 6 flax forms evaluated, to combine optimum lactation performance and protection of flax constituent cis-9,cis-12,cis-15 18:3.