酸法和酶法提取牦牛骨胶原蛋白的特性分析

以牦牛骨为原料,分别采用酸法和酶法提取胶原蛋白并进行理化性质分析。通过氨基酸组成、紫外光谱、傅里叶变换红外光谱、热收缩温度、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（sodium dodecyl sulfate-polyacrylamide gelelectrophoresis,SDS-PAGE）和扫描电子显微镜图像分析对胶原蛋白进行比较。结果表明：2?种方法的提取物都为典型的胶原蛋白,在230?nm波长左右出现最大紫外吸收峰;酸溶胶原蛋白（acid-soluble collagen,ASC）和酶溶胶原蛋白（pepsin-soluble collagen,PSC）的热收缩温度分别为40.12?℃和40.94?℃,变性焓值分别为0.25?J/g和0.22?J/g;红外光谱及SDS-PAGE分析表明,ASC和PSC主要由α、β、γ三种亚基组分组成,属于I型胶原蛋白,且三股螺旋结构完整;扫描电子显微镜结果表明,2?种方法提取的胶原蛋白都保留了较为完整的纤维网状结构,但酸法提取的胶原蛋白结构分布相对均匀。综合来看,2?种方法所提胶原蛋白在理化特性上并无太大差别,但酶法提取的胶原蛋白得率较高,酸法提取成本较低。     

乙酸溶胀-挤压提取对罗非鱼真皮胶原蛋白理化性质的影响

以磷酸预处理辅助实现罗非鱼皮真皮与表皮的分离,采用低温乙酸溶胀法提取真皮中的胶原蛋白,并对所得胶原蛋白样品进行理化性质分析。结果表明,在4℃环境中,以0.006 mol/L磷酸、50 g/L料液处理鱼皮3 h,可获得最佳真皮、表皮分离效果。所得真皮在4℃环境中,以0.035 mol/L乙酸溶液(料液质量浓度为100 g/L)溶胀处理6 h,挤压后可得羟脯氨酸质量分数为7.9%的真皮胶原蛋白样品,提取得率达27.6%(干基)。凝胶电泳结果分析表明,所得样品主要含有α1、α2和β链,由紫外光谱分析结果可知其最大吸收在230 nm波长附近,符合典型Ⅰ型胶原蛋白特征;衰减全反射傅里叶变换红外光谱分析结果显示其三螺旋结构得到较好地保留。样品热变性温度及热收缩温度分别为29.2℃和55.0℃。研究结果为鱼皮资源的高效开发利用以及高品质胶原蛋白的制备提供了理论参考。     

Biochemical and thermo-mechanical analysis of collagen from the skin of Asian Sea bass (Lates calcarifer) and Australasian Snapper (Pagrus auratus), an alternative for mammalian collagen

Australasia has a large fish industry, and fish skin by-products from the processing industry could be used for the commercial production of fish collagen. The aim of this study was to characterize collagen extracted from the Asian sea bass (Australian barramundi) (Lates calcarifer) and snapper (Pagrus auratus) skin as an alternative to mammalian-derived collagen in gelatin products. The acid-soluble fractions of collagen from Asian sea bass and snapper skin were extracted and yielded about 8 and 7.5 % collagen (on a dry weight basis), respectively. The electrophoretic and chromatography patterns indicated that both collagens comprise of α1, α2, α3, and β chains, corresponding to the properties of calf skin collagen type I. Amino acid analysis and peptide mapping of digested collagen suggested differences in their amino acid sequences and collagen primary structure. Fourier transform infrared spectroscopy demonstrated that the helical structure of collagen was completely maintained in Asian sea bass and partially in snapper. Transition temperatures for the completion of the melting process in the two collagen networks were confirmed with differential scanning calorimetry and dynamic oscillatory rheology to be about 29 °C. Zeta potential analysis identified the isoelectric points (pI values) of collagen from Asian sea bass and snapper skin at pH 6.90 and 7.75, respectively. Thus, Asian sea bass and snapper skin could be an important alternative source of collagen to replace mammalian collagen for industrial applications.     

超声波辅助酶法提取牛皮胶原蛋白及其结构表征

以牛皮为原料,优化超声波辅助酶提取牛皮中胶原蛋白的工艺。在单因素实验的基础上设计响应面试验,以牛皮胶原蛋白提取率为响应值,优化得到胶原蛋白的最佳提取工艺,并对其进行结构表征。结果表明:牛皮中胶原蛋白的最佳提取工艺条件为超声波功率161 W、超声波处理时间64 min、胃蛋白酶添加量109 U/g、料液比1:16 g/mL,在此条件下胶原蛋白的提取率为63.77%。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)、紫外光谱(UV)和傅立叶红外光谱(FTIR)分析表明,超声波辅助酶提取的胶原蛋白符合Ⅰ型胶原蛋白的特征,保持了其完整的三螺旋结构,氨基酸组成和扫描电镜(SEM)分析得到超声波辅助酶提取的胶原蛋白三螺旋稳定程度略微下降。本研究为牛皮中胶原蛋白的提取优化了一种有效的工艺,且很大程度上缩短了胶原蛋白的提取时间,并且具有较高的应用价值。     

Structural characteristics of camel-bone gelatin by demineralization and extraction

Camel bone was demineralized through HCl acidulation process at different concentrations (0.0%, 1.5%, 3.0%, and 6.0%) over 1–5 days. The level of demineralization was acid concentration and soaking time dependent. Highest demineralization (62.0%) was recorded in bone sample treated with 6.0% dilute acid for 5 days. Energy dispersive X-ray spectroscopy (EDX) elemental analysis revealed reduction in Ca and increase in N and H, while O remains unaffected. Particulate characteristics by scanning electron microscope showed an increased surface roughness of bone after demineralization. Fourier transform infrared (FT-IR) analysis of ossein depicted the presence of functional group similar to that of bone protein (collagen). Statistical optimization by central composite design (CCD) revealed a significant quadratic model for optimum values of extraction temperature, pH, and extraction time. The highest gelatin yield from camel bone was 23.66% at optimum extraction condition (71.87°C, pH 5.26, and 2.58 h) and the bloom was 205.74 g. Camel bone is suitable for production of gelatin with good potentials in food and nonfood applications.     

两种方法提取的鸡肺胶原蛋白性质及保湿性研究

本文以鸡肺为原材料,利用酸提和超声辅助酸提法提取胶原蛋白,并对其性质及保湿性进行研究。结果表明,所提取的鸡肺胶原蛋白经氨基酸分析显示组成基本相似,甘氨酸含量最高,分别占24.12%和23.93%,亚氨基酸含量仅次之,分别占19.74%和22.28%。酸提和超声辅助酸提胶原蛋白在229.0和224.9 nm处有胶原蛋白的特征紫外吸收峰。SDS-PAGE显示两种鸡肺胶原蛋白中均含有α₁链、α₂链、少量的β链和γ链,傅里叶变换红外光谱及X-射线衍射图谱共同表明两种方法提取的鸡肺胶原蛋白分子排列紧密,均保持了原有的三螺旋结构,说明在超声辅助酸提过程中胶原蛋白的结构保持完好,三螺旋结构未被破坏。酸提的胶原蛋白等电点为7,粒径分布在340 nm左右;超声辅助酸提的胶原蛋白等电点为6,粒径分布在295 nm左右,结合胶原蛋白的溶解性说明了超声辅助酸提可破坏胶原蛋白的氢键,改变蛋白质的颗粒大小和聚集形态。扫描电镜结果表明超声辅助酸提的胶原蛋白结构呈连续纤维状并多孔,且颗粒直径明显减小。在放置24 h时,超声辅助酸提胶原蛋白的吸水性和保水性分别为常规酸提胶原蛋白的1.25倍和1.17倍。综上所述,超声辅助酸提未破坏胶原蛋白特有的三螺旋结构,通过改变氢键和蛋白的颗粒大小使得胶原蛋白具有更好的溶解性和吸湿保湿性。     

Study on the self‐assembly property of type I collagen prepared from tilapia (Oreochromis niloticus) skin by different extraction methods

Type I collagen was prepared from tilapia (Oreochromis niloticus) skin by acetic acid and pepsin process at 4 °C, respectively (ASC and PSC), and hot‐water method separately at 25, 35 and 45 °C (C‐25, C‐35 and C‐45). Their structure and self‐assembly property were discussed. SDS‐PAGE patterns suggested that pepsin hydrolysis and the 35 and 45 °C extraction produced collagen with much reduced proportions of α‐ and β‐chains. Fourier transform infrared spectroscopy spectra revealed that pepsin hydrolysis did not change the conformation of collagen, but higher extraction temperature did. Self‐assembly curves and atomic force microscopy (AFM) observations showed that only ASC, PSC and C‐25 could self‐assemble into fibrils with D‐periodicity, but the reconstruction rate of C‐25 was lower. Besides, PSC had relatively higher resolution ratio compared with others. Overall, pepsin‐extracted collagen displayed higher solubility and better fibril‐forming capacity, having the potential of applying in biomaterials and food‐packaging materials.     

不同提取工艺下牛骨胶原蛋白的结构特性

为提高牛骨资源的利用率,以新鲜牛骨为原料,分别采用乙酸、柠檬酸、乙酸和胃蛋白酶、柠檬酸和胃蛋白酶提取得到四种牛骨胶原蛋白（ASC1、ASC2、PSC1和PSC2）并通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）,傅里叶红外光谱（FTIR）,扫描电镜（SEM）,表面疏水性等指标对其理化性质和结构特征进行研究。结果表明:PSC1的得率最高,可达3.23%,四种提取方法得到的胶原蛋白均于230 nm附近出现最大紫外吸收峰;SDS-PAGE和红外光谱分析表明牛骨胶原蛋白主要由α₁、α₂、β三种亚基成分组成,属于 I型胶原蛋白,且胶原蛋白的空间结构保留完整;扫描电镜直观表明了四种胶原蛋白的纤维网络结构均保留较完整,其中酸法提取的胶原蛋白（ASC1和ASC2）网络结构分布较为均匀;酶法制得的胶原蛋白（PSC1和PSC2）纤维化较为明显,四种胶原蛋白理化特性无显著差异,但微观结构存在一定差异。本研究为提高畜禽副产物高值化应用发展及深加工技术提供了一定的理论基础。     

不同部位鲽鱼皮酸溶性胶原蛋白的提取及其特性

以新鲜的鲽鱼黑皮和白皮为研究对象,从其中提取酸溶性胶原蛋白(acid-soluble collagens,ASC),并对其理化性质和乳化活性及乳化稳定性进行比较研究,为鲽鱼皮的开发利用提供一定的理论依据。通过傅里叶变换红外光谱(Fourier transform infrared spectroscopy,FTIR)技术和扫描电子显微镜(scanning electron microscopy,SEM)初步研究了黑皮和白皮ASC的结构特征;同时对其溶解性、流变学特性、等电点、乳化活性指数(emulsifying activity index,EAI)及乳化稳定性指数(emulsifying stability index,ESI)进行了研究。FTIR扫描结果表明,黑皮和白皮ASC都存在酰胺A、酰胺B、酰胺Ⅰ、酰胺Ⅱ、酰胺Ⅲ,均保持了胶原蛋白三维螺旋结构的完整性;SEM观察结果表明,黑皮和白皮ASC均呈现多空的网络结构,且分布比较均匀,说明酸法提取胶原蛋白的过程基本上保留了胶原纤维原有的空间网状结构。EAI及ESI结果表明,黑皮ASC的EAI明显低于白皮ASC(P0.05),分别为(2.60±0.53)、(7.30±0.41)m~2/g,而黑皮ASC的ESI明显高于白皮ASC(P0.05),分别为(49.12±5.18)、(27.56±4.91)min。以上结果显示,从黑皮和白皮中提取的ASC的理化性质和功能特性存在一定的差异。     

Collagen content in farmed Atlantic salmon (Salmo salar,L.) and subsequent changes in solubility during storage on ice

To elucidate whether collagen is an important factor for fish flesh quality, the collagen content and its changes in solubility during storage on ice in muscle of farmed Atlantic salmon (Salmo salar, L.) were measured. The contents of acidsoluble, pepsin-soluble and insoluble collagen in white muscle were determined in fresh fish muscle and at the end of 5, 10 and 15 days storage on ice. Total collagen was found to be 0.66% of fresh weight, with a relative distribution of 6% acidsoluble, 93% pepsin-soluble and 1% insoluble collagen. During storage on ice, a progressive change in solubility of muscle collagen was found. For insoluble collagen, significantly lower values were detected at day 15 compared to day 0. A minor, but even increase in acid-soluble collagen was found from day 0, while no changes were seen in pepsin-soluble collagen during storage. These results show that collagen fibres of farmed Atlantic salmon have a high solubility in acid and salt solutions and contain few cross-links. Some cleavage of intermolecular cross-links seems to occur during storage on ice.     

提取温度对罗非鱼皮明胶理化性质的影响

本研究以罗非鱼皮为原料,研究在不同温度(30、40、50、60、70、80、90℃)下提取的明胶所表现的得率、凝冻强度、聚丙烯酰胺凝胶电泳分析、傅里叶红外光谱分析、热变性温度、流变性性能等理化特征。结果表明,提取温度对鱼皮明胶的紫外吸收无明显影响;随着提取温度的升高,鱼皮明胶的得率逐渐增大,在提取温度为60~80℃时差异不显著(p>0.05),在90℃达到最大值(62.60%±0.84%);聚丙烯酰胺凝胶电泳图谱显示,较低温度(30、40、50℃)下提取的鱼皮明胶由α₁、α₂、β 3条肽链组成,提取温度升高后电泳条带强度降低;提取温度为50℃时,罗非鱼皮明胶的凝冻强度最大,高达(884.33±26.76)g;FTIR光谱显示,不同温度下提取的鱼皮明胶在酰胺区都具有特征吸收峰,在光谱中显示出微小的差异;不同温度下提取的鱼皮明胶的热变性温度分别为(101.05±2.97)、(104.35±3.54)、(107.59±0.37)、(97.80±4.21)、(95.35±2.60)、(92.52±3.15)、(89.66±1.23)℃;提取温度通过改变明胶的分子量大小影响明胶溶液内部贮藏能量的多少,进而对明胶溶液的流变性能产生影响。     

梅花鹿鹿茸胶原的理化特性研究

对胃蛋白酶法在酸性条件下制备的梅花鹿鹿茸胶原的理化性质进行研究。通过紫外扫描(UV)、傅里叶红外光谱(FTIR)、氨基酸组成测定、聚丙烯酰胺凝胶电泳(SDS-PAGE)和示差量热扫描(DSC)等方法对其理化性质进行分析。紫外扫描图谱可知,胶原纯度较高,在230nm波长处有强吸收峰;红外光谱表明其酰胺A、酰胺B、酰胺Ⅰ谱带等主要吸收峰与Ⅰ型胶原标品一致,具有三螺旋结构;凝胶电泳表明鹿茸胶原含有α1、α2和β链,符合Ⅰ型胶原特征;氨基酸分析说明其具有典型胶原的氨基酸组成;示差量热扫描测得其热收缩温度(ts)为84.05℃。梅花鹿鹿茸具有典型的I型胶原的理化特性。     

Characterization of acid- and pepsin-soluble collagens from flatfish skin

Acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC) from flatfish skin were characterized. The yield of PSC was 85.5%, which was higher than that of ASC at 57.3%. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns showed that both the ASC and PSC were consisted of α1 and α2 chains, and β-chain. According to the results of Fourier transform infrared (FTIR) spectroscopy and hydroxylation, no difference in the helical structure between ASC and PSC was found. Thermal denaturation temperature (TDT) of flatfish skin collagen was 26.6°C for ASC and 26.7°C for PSC, which were similar to those of temperate fishes, while were lower compared to tropical fishes, such as the conger eel and Nile perch. There was no solubility difference between ASC and PSC at various pH and NaCl concentrations. Therefore, PSC will be preferable compared to ASC for commercial uses because of its higher yield.     

罗非鱼胶原蛋白肽补血微囊制备工艺及其理化特性研究

为了研究胶原蛋白肽补血微囊工艺条件以及包合前后理化性质及结构变化,采用阿拉伯胶和明胶对黄芪甲苷、阿魏酸和鱼胶原蛋白肽进行包合,采用热重分析、傅里叶红外光谱和X射线衍射对包合后产物的理化性质和结构特性进行检测。结果显示,采用Box-Behnken响应面试验对包合工艺进行优化,得到最佳包合工艺条件为芯壁比1:8.333、壁材浓度0.91%、超声时间15.94 min,得到平均包合率75.35%;修正为:芯壁比1:8、壁材浓度0.9%、超声时间16 min,得到黄芪甲苷包合率73.59%、阿魏酸包合率88.76%、鱼胶原蛋白肽包合率65.80%,包合工艺的平均包合率76.05%。X-射线衍射(XRD)、热重分析(TG)、傅里叶红外光谱(FI-TR)表明包合物已经形成,而且包合物显著增加芯材稳定性。     

超声波辅助提取骆驼皮胶原蛋白的工艺及结构表征

以骆驼皮为原料,研究超声波辅助提取胶原蛋白的最优工艺,并对提取得到的胶原蛋白进行表征.以胶原蛋白的提取率为指标,探讨酶添加量、料液比、超声波功率、超声波处理时间及总提取时间5个因素对骆驼皮胶原蛋白的影响,确定超声波功率后,在单因素实验的基础上,结合响应曲面分析法得到最佳提取条件为:酶添加量4.5％、料液比1:23、超声...     

Effects of Pretreatment on Properties of Gelatin from Perch (Lates Niloticus) Skin

Fish gelatins obtained from perch fish skin pretreated with various solutions containing acetic acid, sodium hydroxide (NaOH) and sodium chloride (NaCl) were successfully characterized for their nanostructure pattern using field emission scanning electron microscopy. Each pretreatment transformed collagen to gelatin with fibril, zigzag cracks, straight rods, and cross-linked rods nanostructure patterns. Pretreatment solutions also affect the gel yield, gel strength, amino acid profile, and functional groups in perch gelatin as analyzed by Fourier transform infrared spectroscopy. Samples pretreated with NaCl, NaOH, and acetic acid solution showed the highest gel yield (22.84%) and gel strength (179.84 g). Fourier transform infrared spectra for perch gelatins also revealed weak C–N amide II and III bond stretches as well as weak C=O bond stretch.     

淡水鱼皮胶原蛋白提取工艺研究(Ⅰ)

研究了罗非鱼、鳙鱼、草鱼和鲫鱼4种淡水鱼皮胶原蛋白的提取工艺。结果表明,鱼皮用乙醚2次除脂效果显著,除脂率依次为92.4%、95.1%、66.8%和76.7%,除脂后胶原蛋白的提取率和得率都略有增加。除非胶原蛋白的工艺为:先用NaOH浸泡6h,再用NaCl浸泡6h,4种鱼皮胶原蛋白对应的提取率分别为34.4%、28.7%、31.5%和42.5%,得率依次为8.4%、8.0%、8.2%和6.1%,比其他处理相应高出0.4-2.3倍、0.04-1.6倍、0.1-0.9倍和0.07-0.5倍。乙酸的提取效果优于柠檬酸,对应提取率分别为34.8%、23.6%、33.6%、43.0%,得率依次为8.5%、6.6%、8.63%、6.19%,比柠檬酸相应高2.4-4.3倍、1.7-4.0倍。温度对罗非鱼、鳙鱼和草鱼鱼皮胶原蛋白提取影响较大,最佳提取温度都为20℃,其对应提取率依次为34.05%、27.05%、33.38%,得率分别为8.32%、7.54%、8.58%;温度对鲫鱼鱼皮胶原蛋白的提取影响较小,在520℃范围内提取率和得率分别约为42%和6%左右。     

海蜇Ⅰ型胶原蛋白的提取及结构特性研究

以海蜇为原料，采用盐酸-胃蛋白酶法提取胶原蛋白，并通过聚丙烯酰胺凝胶电泳（SDS-Polyacrylamide gel electrophoresis，SDS-PAGE）、紫外光谱（UV-visible spectroscopy，UV）、氨基酸分析、红外光谱（Fourier-transform infrared spectroscopy，FTIR）、圆二色谱（Circular dichroism，CD）以及扫描电镜（Scanning electron microscope，SEM）对其结构特性进行了系统全面的研究。结果显示，海蜇胶原蛋白在135 kDa左右有一条α-链，在245 kDa以上有一条β-链和γ-链，紫外吸收峰在233 nm处，符合I型胶原蛋白的特征，其亚基组成可能为[α₁(I)]₃。海蜇胶原蛋白中含量最高的氨基酸为甘氨酸，占氨基酸总量的25.99%，且含有15.94%的亚氨基酸。红外光谱和圆二色谱均表明海蜇胶原蛋白分子排布紧凑，具有主要由氢键构成的完整三螺旋结构。扫描电镜结果显示海蜇胶原蛋白具有多层聚集、以纤维为主的无规则网状结构。     

Comparison of collagen characteristics of sea- and freshwater-rainbow trout skin

Proximate composition, volatile basic nitrogen content, and concentrations of collagen in skin samples from either sea- (S-RT) or freshwater-rainbow trout (F-RT) were characterized and compared, to assess the effect of the sea or freshwater habitat on these parameters. Results of amino acid composition, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, thermal denaturation assay and Fourier transform infrared (FT-IR) spectroscopy of acid-soluble collagens were comparable between the two sample sets. Both acid-soluble collagens from sea- and freshwater-rainbow trout skins contained glycine as the major amino acid and high alanine, proline, and hydroxyproline contents, and was found to be predominantly composed of α1-, α2-, and β-chains. FT-IR spectra of ASCs from S-RT and F-RT skins were quite similar. These findings suggest that different rainbow trout habitats (seawater and freshwater) do not affect amino acid composition and molecular weight properties of ASCs from S-RT and F-RT skins.     

Chemical composition and thermal properties of Tilapia oil extracted by different methods

Fish oil is beneficial to health and its quality is significantly impacted by the extraction method. To determine the effect of various methods used to extract the oil on its chemical composition and thermal properties, tilapia oil extracted by dilute alkali hydrolysis, enzymatic hydrolysis, and microwave-assisted methods were investigated in this study. Nutrient analyses showed that freeze-dried tilapia contained fat (36.69 g/100 g freeze-dried tilapia), protein (49.95 g/100 g freeze-dried tilapia), ash (12.04 g/100 g freeze-dried tilapia) and moisture (1.21 g/100 g freeze-dried tilapia). Chemical analysis showed that the fish oil extracted by dilute alkali hydrolysis had the lowest acid value (AV) and peroxide value (POV) (AV: 0.77 mg KOH/g oil and POV: 3.76 mmol/kg oil) compared to the oil extracted using the other methods. The main fatty acid components were palmitic acid (22.78–23.16%), oleic acid (25.73–26.19%), and linoleic acid (15.69–15.98%). Furthermore, the oil extracted by dilute alkali hydrolysis showed significantly higher content of DHA+ EPA (2.30%). The thermal behaviors indicated that the aforementioned oils had significantly different crystal structures. The main endothermic peaks of fish oil extracted by dilute alkali hydrolysis, enzymatic hydrolysis and microwave-assisted methods were located at ?2.03℃, ?3.20℃ and ?4.37℃, respectively. However, infrared spectroscopic analyses indicated them had the same characteristic functional groups. Consequently, different extraction methods significantly affect the oil’s physical and chemical properties without affecting spectral characteristics. Furthermore, these results advocated the use of alkali hydrolysis for fish oil extraction.