Application of cavitation system to accelerate the endogenous enzymatic hydrolysis of baicalin and wogonoside in Radix Scutellariae

Baicalinase is an endogenous enzyme present in the roots of Scutellaria baicalensis that can hydrolyse baicalin and wogonoside into baicalein and wogonin directly, which have higher bioactivity and bioavailability than their glycosides. Baicalinase-catalysed hydrolysis with cavitation was examined as a highly efficient and green alternative method for large scale production of baicalein and wogonin. Effect of cavitation system on the transformation and extraction efficiency was investigated by using reaction kinetic and mass transfer kinetic models at three levels of temperature (20-50 °C). Experimental results showed that cavitation had a significant effect (p < 0.01) on the biocatalysis process; the transformation of baicalin and wogonoside reached 98.21% and 96.60% at 50 °C for 60 min, and the yields of baicalein and wogonin were about 4.47- and 2.86-fold higher than those of the raw sample without enzymatic hydrolysis, respectively.     

蛋白基体系美拉德反应在荔枝皮原花青素作用下的抑制研究

本试验分别研究了在乳清蛋白-葡萄糖和牛血清白蛋白-葡萄糖模拟生理体系中,荔枝果皮原花青素(Litchi pericarp procyanidins,LPPC)对美拉德反应和晚期糖基化终末产物(advanced glycation end products,AGEs)的抑制作用,以荧光性AGEs的强度为表征依据。结果表明,在乳清蛋白-葡萄糖模拟体系中孵育35 d时,LPPC对AGEs的抑制效果最强,达60.21±1.34%。LPPC浓度为1 mg/m L时,相对抑制率最大可达85.33±9.02%(显著高于维生素C(Vc),p0.05)。在牛血清白蛋白-葡萄糖体系中,孵育35 d后LPPC对AGEs的相对抑制率最高达70.01±1.32%,且与LPPC浓度呈现正相关。当LPPC浓度为0.5 mg/m L时,抑制率最大为95.46±10.12%(显著高于氨基胍(AG),p0.05)。不同p H值下蛋白质的稳定性研究表明,乳清蛋白在与LPPC长期孵育后,其热稳定性明显降低,再次提示了LPPC对美拉德反应的抑制作用。LPPC可作为一种天然的食品基质的AGEs抑制剂深度开发。     

Fractionation of lipids and purification of γ-linolenic acid (GLA) from Spirulinaplatensis

The microalgae, Spirulinaplatensis, is an excellent source of γ-linolenic acid (GLA), an essential polyunsaturated fatty acid and a potent nutraceutical. The fatty acid composition of S.platensis ARM 740 was determined after transmethylation by gas chromatography (GC). Lipid fractionation was achieved on silica gel column chromatography and preparative TLC. Neutral lipids, glycolipids and phospholipids accounted for 77.0%, 15.6% and 7.4%, respectively, of the total lipid fraction. S.platensis ARM 740 was found to contain 94% of the total GLA in the glycolipid fraction. Attempts were made to purify GLA methyl ester by using urea to form inclusion complexes with the saturated and the less unsaturated FAMEs (fatty acid methyl esters), which enhanced the purity of GLA methyl ester to 84%. A further approach to isolate GLA methyl ester with higher purity involved the use of argentated silica gel chromatography. An initial PUFA concentration step frequently adopted by most researchers to increase GLA purity was not necessary in the isolation of GLA from S.platensis. A GLA methyl ester with a purity of >96% and a recovery of 66% was obtained. Purity of the isolated GLA methyl ester was confirmed by GC and IR analysis with respect to authentic standard.     

Selective enrichment of n−3 polyunsaturated fatty acids with C18–C20 acyl chain length from sardine oil using Pseudomonas fluorescens MTCC 2421 lipase

An extracellular lipase purified from Pseudomonas fluorescens MTCC 2421 was used to enrich sardine oil triglycerides with eicosapentaenoic acid (20:5 n−3) and linolenic acid (18:3 n−3) to 35.28% and 8.25%, respectively, after 6 h of hydrolysis. The corresponding n−6 fatty acids (18:2 n−6 and 20:4 n−6) exhibit a reduction (54.93% and 50%, respectively). Structure–bioactivity relationship analyses revealed that the lower hydrophobic (log P values) constants of 18:3 n−3 and 20:5 n−3 (5.65 and 5.85, respectively) result in their higher hydrolytic resistance towards lipase, leading to their enrichment in the triglyceride fraction after lipase-catalysed hydrolysis. Lipase-catalysed hydrolysis of sardine oil for 6 h followed by urea fractionation at 4 °C with methanol provided free fatty acids containing 42.50% 20:5 n−3 and 10.31% 18:3 n−3, respectively. Argentation neutral alumina column chromatography, using n-hexane/ethylacetate (2:1, v/v) resulted in 20:5 n−3 of high purity (83.62%), while 18:3 n−3 was found to be eluted with n-hexane/dichloromethane (4:1, v/v) as eluting solvent with a final purity of 75.31%.     

Preparative isolation and purification of xanthohumol from hops (Humulus lupulus L.) by high-speed counter-current chromatography

Xanthohumol (XN) and related prenylflavonoids are the main bioactive components of hops (Humulus lupulus L.). The current work is to investigate the use of high-speed counter-current chromatography (HSCCC) in search for high isolation of xanthohumol from hops. A solvent system consisted of n-hexane-ethyl acetate-methanol-water at a volume ratio of 5:5:4:3 was employed. The results demonstrated that the constructed method could be well applied for the isolation of xanthohumol from hops extract. After HSCCC isolation procedure, the purity of xanthohumol was over 95% assayed by HPLC and the yield of extraction was 93.60%. The chemical structure identification of xanthohumol was carried out by UV, ¹H NMR and ¹³C NMR. The present results demonstrated that xanthohumol could be efficiently obtained using a single HSCCC step from H. lupulus L. extract.     

荔枝皮原花青素与VC、VE的协同抗氧化研究

选取荔枝皮原花青素(LPPC),采用DPPH自由基清除实验与等辐射分析法(Isobologram)相结合,探讨荔枝皮原花青素、VC、VE单独和复配后的抗氧化作用,结果显示：荔枝皮原花青素、VC和VE的IC50值分别为9.98、9.21、27.96mg/L,表明荔枝皮原花青素对DPPH自由基有明显的清除作用。Isobologram分析图中荔枝皮原花青素与VC、VE复配后的效应点都在相加线及95%可信限的下方,理论IC50add值与实验IC50mix值有显著性差异,并且相互作用指数都小于1,证实荔枝皮原花青素与VC,荔枝皮原花青素与VE之间存在协同抗氧化效应。     

基于一测多评法对小儿解感颗粒中5个成分的质量控制

目的 建立基于一测多评法(QAMS)测定小儿解感颗粒中黄芩苷、汉黄芩苷、黄芩素、汉黄芩素、甘草苷5个成分的含量的方法 .方法 采用高效液相色谱(HPLC),色谱柱为Waters SunFire C18柱(4.6 mm×250 mm,5μm);流动相为乙腈(A)-0.1％磷酸水溶液(B),梯度洗脱;流速1.0 ml/mi...     

龙葵果花色苷的分离与鉴定

采用硅胶柱层析技术分离制备龙葵果花色苷。分离得到的2 个花色苷馏分经紫外-可见光谱、高效液相色谱-电喷雾串联质谱（high performance liquid chromatography electrosprary ionization-mass spectrometry,HPLC-DADESI-MS/MS）进行结构鉴定,并结合酸水解分析糖苷种类。最终确定馏分Ⅰ为飞燕草素-3-琥珀酰阿拉伯糖苷,根据峰面积归一化法计算其纯度为94%;馏分Ⅱ为矢车菊素-3-半乳糖苷和矢车菊素-3-乙酰半乳糖苷,峰面积归一化法计算其纯度分别为45.67%和13.97%。     

二次柱层析制备高纯度表没食子儿茶素没食子酸酯(EGCG)的工艺研究

以商品荼多酚为原料,先用聚酰胺柱层析预分离表没食子儿茶素没食子酸酯(EGCG),再用硅胶柱层析制备高纯度的EC-CG.采用此工艺条件,利用较低等级的商品茶多酚(70%TP)可以制备出纯度达98%以上的EGCG.     

Effects of limited enzymatic hydrolysis with trypsin on the functional properties of hemp (Cannabis sativa L.) protein isolate

Effects of limited enzymatic hydrolysis induced by trypsin on the physicochemical and functional properties of hemp (Cannabis sativa L.) protein isolate (HPI) were investigated. The enzymatic hydrolysis was confirmed by sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and size exclusion chromatography (SEC). SEC and differential scanning calorimetry (DSC) analyses confirmed the presence of aggregates in the corresponding hydrolysates (with the degree of hydrolysis of 2.3–6.7%). Functional properties, including protein solubility (PS), thermal properties, emulsifying and foaming properties, and water holding and fat adsorption capacities (WHC and FAC) were evaluated. The PS was remarkably improved by the limited enzymatic hydrolysis at all tested pH values. However, the enzymatic hydrolysis led to the marked decreases in emulsifying activity index, foaming capacity and foam stability, WHC and FAC. These decreases were to a great extent related to the presence of aggregates in the hydrolysates.     

Purification and identification of novel antioxidant peptides from enzymatic hydrolysates of sardinelle (Sardinellaaurita) by-products proteins

In order to utilise sardinelle (Sardinellaaurita) protein by-products, which is normally discarded as industrial waste in the process of fish manufacturing, heads and viscera proteins were hydrolysed by different proteases to obtain antioxidative peptides. All hydrolysates showed different degrees of hydrolysis and varying degrees of antioxidant activities. Hydrolysate generated with crude enzyme extract from sardine (Sardinapilchardus) displayed high antioxidant activity, and the higher DPPH radical-scavenging activity (87 ± 2.1% at 2 mg/ml) was obtained with a degree of hydrolysis of 6%. This hydrolysate was fractionated by size exclusion chromatography on a Sephadex G-25 into eight major fractions (P₁–P₈). Fraction P₄, which exhibited the highest DPPH scavenging activity, was then fractionated by reversed-phase high performance liquid chromatography (RP-HPLC). Seven antioxidant peptides were isolated. The molecular masses and amino acids sequences of the purified peptides were determined using ESI-MS and ESI-MS/MS, respectively. Their structures were identified as Leu-His-Tyr, Leu-Ala-Arg-Leu, Gly-Gly-Glu, Gly-Ala-His, Gly-Ala-Trp-Ala, Pro-His-Tyr-Leu and Gly-Ala-Leu-Ala-Ala-His. The first peptide displayed the highest DPPH radical-scavenging activity (63 ± 1.57%; at 150 μg/ml) among these peptides.     

Acid hydrolysis of kappa‐carrageenan as a way of gaining new substances for freezing process modification and protection from excessive recrystallisation of ice

The effects of κ‐carrageenan and its hydrolysates on modification of the freezing process and also on inhibition of excessive recrystallisation of ice in sucrose solutions during storage were compared. Acid hydrolysis of κ‐carrageenan was carried out using sulphuric (H₂SO₄) and hydrochloric acid (HCl). Most effective in the hydrolysis process turned out to be H₂SO₄, which degraded κ‐carrageenan to a molecular mass of around 3 × 10⁶ Da, after 1.5 h of hydrolysis. Addition of 0.005% of the new poligeenan (degraded carrageenan), to a sucrose solution (30%), frozen at ?20 °C, caused a nearly 50% reduction in the phase‐change stages, and consequently, the total time of freezing was shorter. Significant retardation of recrystallisation was observed for both types of poligeenan, but a stronger effect was observed for the oligosaccharides obtained after HCl hydrolysis, and after 96 h of storage at ?8 °C, the equivalent diameter of ice crystals was not greater than 11 μm.     

Isolation and purification of four flavone C-glycosides from antioxidant of bamboo leaves by macroporous resin column chromatography and preparative high-performance liquid chromatography

The present study investigated a robust method for the preparation of four flavone C-glycosides, i.e. orientin, homoorientin, vitexin and isovitexin, which were prepared from an ethanol aqueous extract, i.e. antioxidant of bamboo leaves (AOB), by AB-8 resin-based column chromatography and preparative high-performance liquid chromatography (HPLC) using a mobile phase consisting of 10% and 15% (v/v) of acetonitrile and 1% acetic acid. These flavone C-glycosides were further purified by the drowning-out crystallization method using methanol and water as drowning-out anti-solvents and salting-out agents, respectively. The purity was assessed by analytical HPLC and the confirmation of chemical structures was performed by IR, MS, NMR and UV spectroscopy. Orientin (49 mg), homoorientin (142 mg), vitexin (15 mg) and isovitexin (62 mg) were prepared from 6.5 g of crude column chromatography fraction obtained from 5 L of AOB concentrated solution. The present method is robust and suitable for preparing available quantities of pure flavone C-glycosides and the quantification of orientin, homoorientin, vitexin and isovitexin in bamboo leaves.     

Hypocholesterolaemic effect of Bifidobacterium animalis subsp. lactis (Bb12) and trypsin casein hydrolysate

Hypercholesterolaemia is one of the most important risk factors in the development of cardiovascular disease. In this study, we report the in vitro potential of Bifidobacterium animalis subsp. lactis (Bb12) cultures and bovine casein hydrolysates formed by trypsin and Bb12 culture to reduce cholesterols levels. Cholesterol levels in vitro were reduced by up to 48% after incubation with Bb12 and up to 87% after incubation with trypsin hydrolysates, whereas unhydrolysed bovine casein did not affect cholesterol levels. Individual peptide fractions, obtained from size-exclusion chromatography, from casein hydrolysates formed by trypsin after a 48 h hydrolysis, reduced cholesterol levels by 2.7–50%. The molecular masses of these fractions, containing hypocholesterolaemic peptides, were below 1200 Da, as determined by LC-MS.     

小麦麸皮中β-葡聚糖的分离纯化及组成研究

以小麦麸皮为原料,碱法提取β-葡聚糖。采用硫酸铵沉淀法、DEAE Sepharose CL-6B阴离子交换柱层析法、Sepharose CL-4B凝胶过滤层析法对β-葡聚糖的粗提物进行逐步纯化,得到组分A1和A2。并经紫外分光光度法、高效凝胶渗透色谱法鉴定其纯度,最终获得相对分子质量为4.87×105（纯度为98.57%）的β-葡聚糖A1组分、6.13×104左右（纯度为97.03%）的A2组分。可见,经过一系列的纯化实验,可以获得纯度较高的β-葡聚糖单一组分。采用特异性β-葡聚糖酶对经阴离子交换柱层析获得的β-葡聚糖组分A进行水解,经高效液相色谱法分析其寡糖的组成。结果表明,特异性酶解后的β-葡聚糖组分A主要由麦芽三糖和麦芽四糖组成,除了含有少量葡萄糖外,还含有麦芽糖、麦芽五糖。      

Bioethanol production from ball milled bagasse using an on-site produced fungal enzyme cocktail and xylose-fermenting Pichia stipitis

Sugarcane bagasse is one of the most promising agricultural by-products for conversion to biofuels. Here, ethanol fermentation from bagasse has been achieved using an integrated process combining mechanical pretreatment by ball milling, with enzymatic hydrolysis and fermentation. Ball milling for 2 h was sufficient for nearly complete cellulose structural transformation to an accessible amorphous form. The pretreated cellulosic residues were hydrolyzed by a crude enzyme preparation from Penicillium chrysogenum BCC4504 containing cellulase activity combined with Aspergillus flavus BCC7179 preparation containing complementary β-glucosidase activity. Saccharification yields of 84.0% and 70.4% for glucose and xylose, respectively, were obtained after hydrolysis at 45 °C, pH 5 for 72 h, which were slightly higher than those obtained with a commercial enzyme mixture containing Acremonium cellulase and Optimash BG. A high conversion yield of undetoxified pretreated bagasse (5%, w/v) hydrolysate to ethanol was attained by separate hydrolysis and fermentation processes using Pichia stipitis BCC15191, at pH 5.5, 30 °C for 24 h resulting in an ethanol concentration of 8.4 g/l, corresponding to a conversion yield of 0.29 g ethanol/g available fermentable sugars. Comparable ethanol conversion efficiency was obtained by a simultaneous saccharification and fermentation process which led to production of 8.0 g/l ethanol after 72 h fermentation under the same conditions. This study thus demonstrated the potential use of a simple integrated process with minimal environmental impact with the use of promising alternative on-site enzymes and yeast for the production of ethanol from this potent lignocellulosic biomass.     

Antioxidant activity of hydrolysates obtained from scallop (Patinopecten yessoensis) and abalone (Haliotis discus hannai Ino) muscle

Scallop (Patinopecten yessoensis) and abalone (Haliotis discus hannai Ino) muscle were hydrolysed with commercially available food-grade proteases. The resulting hydrolysates showed DPPH and hydroxyl radicals scavenging abilities, reducing power, and ferrous ion chelating capacity. The antioxidant activities of hydrolysate of abalone foot muscle (HAFM) increased with increasing incubation time during the whole hydrolysis process in 180 min. Whereas, the antioxidant activities of hydrolysate of scallop adductor muscle (HSAM) increased at initial stage and peaked after 25-30 min of hydrolysis, and then gradually decreased thereafter. Compared with HAFM, HSAM with comparable hydrolysis time contained more free amino acids (FAA) and small-sized peptides (below 500 Da), which may account for the differences in antioxidant activities versus hydrolysis time curves of the two hydrolysates. The above results indicate that limited hydrolysis of proteins can increase their antioxidant activity, whereas extensive hydrolysis can decrease it.     

白酒糟制备高纯度微晶纤维素工艺优化及其结构表征

白酒糟含有丰富的纤维素和半纤维素组分,是制备食品添加剂微晶纤维素(MCC)的良好原料来源。在硝酸-乙醇法提取酒糟粗纤维的基础上,探索酸法水解制备高纯度酒糟微晶纤维素(GSMCC)的工艺参数并进行结构表征。着重考察盐酸浓度、温度、液固比和时间4个因素对GSMCC纯度和得率的影响,并通过响应面优化法确定最优工艺参数。结果表明,酒糟粗纤维在温度72.3℃,盐酸质量分数7.5%,液固比25∶1(mL/g)的最优条件下水解2 h,可制得纯度92.57%、聚合度276.39的GSMCC,得率高达89.25%;经漂白后纯度略增至93.31%,聚合度降为255.86。扫描电镜(SEM)、傅里叶变换红外光谱(FTIR)和X衍射(XRD)分析发现,GSMCC的微观形貌呈不规则颗粒状,具有典型纤维素的红外光谱特征,纤维素晶型为Ⅰ型,结晶度为67.49%。热重分析(TGA)表明GSMCC热稳定性良好。本试验结果说明利用白酒糟制备食品添加剂微晶纤维素的工艺可行且具有良好结构特性。     

基于组合色谱技术的蓝莓果实中花色苷元制备

首先采用液液萃取及大孔树脂层析技术研究花色苷的制备方法,萃取剂为乙酸乙脂,大孔树脂为Amberlite XAD-7HP,在此基础上制得花色苷粗提液。花色苷经酸水解、浓缩后,采用固相萃取技术（C18小柱）对花色苷元进行除酸、除糖处理,制得了纯度为70.2%的花色苷元混合物。最后采用半制备型高效液相色谱技术实现4 种主要花色苷元单体的分离,分别为飞燕草素（纯度98.2%）、矢车菊素（纯度96.3%）、牵牛花素（纯度92.6%）、锦葵色素（纯度90.5%）。本研究可为花色苷元的规模化制备及后期功能活性研究提供技术参考和依据。