香菇多糖磷酸化修饰工艺条件的研究

研究了制备磷酸化香菇多糖的工艺条件。通过单因素实验,考察磷酸化试剂、温度、时间、pH值对修饰后香菇多糖磷酸根接枝量、粘度的影响,确定最佳磷酸化工艺条件为:质量分数分别为5%三聚磷酸钠(STPP)和2%三偏磷酸钠(STMP)混合液作为磷酸化试剂,温度90℃,时间5h,pH 9.0,所得磷酸化香菇多糖衍生物的磷酸根接枝量为6.826%。红外光谱分析结果证实,修饰后的香菇多糖含有磷酸酯基团。粘度测定表明,香菇多糖经磷酸化修饰后的粘度基本没有变化。     

磷酸化香菇多糖制备工艺的研究

研究制备磷酸化香菇多糖的工艺条件.以修饰后香菇多糖的磷酸根接枝量、黏度为考察指标,通过单因素和正交实验,考察磷酸化试剂、温度、时间、pH 值时修饰后香菇多糖磷酸根接枝量、黏度的影响.结果确定最佳磷酸化工艺条件为:8%混合磷酸化试剂、温度 80℃、时间 5 h、pH值8.0,所得磷酸化香菇多糖衍生物的磷酸根接枝量为 7.77%,黏度测定表明香菇多糖经磷酸化后黏度几乎没有变化.     

酸法降解浒苔多糖及其清除羟自由基活性研究

以自制的浒苔多糖为原料,利用三氟乙酸(TFA)对其降解,制得低分子质量的降解浒苔多糖。以对.OH清除率为指标表征降解浒苔多糖的抗氧化活性,并进行降解工艺优化。利用正交试验确定最佳降解条件为TFA浓度1.0mol/L、反应时间5h、温度80℃、料液比1:200(g/mL),降解浒苔多糖得率为40.05%,对.OH清除率达61.83%。降解和未降解浒苔多糖的.OH清除率IC50分别为0.3521mg/mL和1.8940mg/mL,降解浒苔多糖的抗氧化活性明显高于未降解浒苔多糖。通过黏度法测得降解前后浒苔多糖的特性黏度值由60.618mL/g降至22.789mL/g,平均相对分子质量由1.19×105降为6.41×104。     

磷酸化香菇多糖的工艺优化

研究了香菇多糖的磷酸化优化工艺条件,并对修饰后的磷酸根接枝量和粘度进行测定.采用正交实验,考察磷酸化试剂用量、温度、时间、pH对修饰后香菇多糖磷酸根接枝量和粘度的影响.结果表明,最佳工艺条件为:8%混合磷酸化试剂(5%三聚磷酸钠和2%三偏磷酸钠的混合试剂),温度80℃,时间5h,pH8.0.在该反应条件下,磷酸化香菇多糖衍生物的磷酸根接枝量为7.77%,粘度测定表明,香菇多糖经磷酸化后,粘度几乎没有变化.     

磷酸化香菇多糖的制备及其部分理化性质的研究

研究了制备磷酸化香菇多糖的工艺条件及其部分理化性质.通过单因素实验,考察磷酸化试剂、温度、时间、pH值对修饰后香菇多糖磷酸根接枝量、黏度的影响,确定最佳磷酸化工艺条件为:5%三聚磷酸钠(STPP)和2%三偏磷酸钠(STMP)混合液作为磷酸化试剂,温度90℃,时间5h,pH值9.O,所得磷酸化香菇多糖衍生物的磷酸根接枝量为6.826%.同时对修饰前后香菇多糖的理化性质进行分析,红外光谱表明磷酸基团已被接枝上且多糖性质无较大变化.     

若羌灰枣多糖的磷酸化修饰及其抗氧化活性研究

用三聚磷酸钠和三偏磷酸钠对纯化后的若羌灰枣多糖(RQGJPs)进行磷酸化修饰，以磷酸化修饰多糖(M-RQGJPs)的取代度作为评价指标，优化磷酸化修饰的反应参数，并分析M-RQGJPs对DPPH·、·O^-₂、·OH的清除率。结果表明：RQGJPs磷酸化修饰的最佳反应参数为反应时间5 h、反应温度80℃、pH 9.5,在此条件下磷酸化取代度为0.092±0.004;在试验质量浓度范围内，M-RQGJPs对DPPH·、·O^-₂、·OH的清除率分别提高了45.2%、68.9%和72.0%,说明磷酸化修饰对RQGJPs的抗氧化活性有明显提升作用。     

响应面法优化超声辅助提取浒苔多糖的工艺

为优化浒苔多糖的提取工艺条件,采用超声辅助提取技术提取浒苔多糖,考察3 个变量(超声温度、超声时间和液料比)对浒苔多糖收率的影响,并通过响应面设计法确定浒苔多糖超声辅助提取技术的最佳工艺条件。结果表明：最佳工艺条件为超声温度80℃、超声时间28min、液料比63:1(mL/g),按此工艺条件提取浒苔多糖,收率为25.84mg/g;验证实验表明,实际浒苔多糖收率与模型预测值相近。采用响应面法优化浒苔多糖超声辅助提取工艺可行。     

黑木耳多糖的磷酸化修饰及其抗氧化活性研究

本研究对黑木耳胞外多糖(Auricularia auricular polysaccharide,AAP)进行磷酸化修饰,通过响应面法优化了磷酸化修饰黑木耳多糖的工艺条件,并对磷酸化黑木耳多糖(phosphorylated auricularia auricula polysaccharide,P-AAP)进行抗氧化活性研究。结果显示磷酸化修饰黑木耳多糖的最佳条件为磷酸化试剂中三聚磷酸钠(sodium tripolyphosphate,STPP)与三偏磷酸钠(sodium trimetaphosphate,STMP)质量比为5:2,反应温度88℃,反应时间5 h,反应pH为8.6,此条件下多糖中磷酸根含量为9.93%。抗氧化活性试验结果表明,与AAP相比,经过DEAE-Sepharose Fast Flow柱及葡聚糖凝胶G-100柱纯化后的P-AAP1对DPPH自由基的清除率提高了34.37%,半抑制浓度(IC₅₀)为1.07 mg/mL;对羟基自由基的清除率提高了30.39%,IC₅₀值为0.91 mg/mL;对超氧阴离子自由基的清除率提高了26.40%,IC₅₀值为0.41 mg/mL。因此,黑木耳胞外多糖通过磷酸化修饰后其抗氧化活性能被显著提高。     

浒苔多糖脱蛋白脱盐方法的优选

目的：探讨浒苔多糖脱蛋白脱盐的方法。方法：利用蛋白酶水解浒苔多糖的蛋白质,通过正交试验优选蛋白酶水解工艺条件;利用透析袋流水透析脱盐,通过正交试验优选透析工艺条件。结果：中性蛋白酶脱蛋白效果优于Sevag法和三氯乙酸法,中性蛋白酶脱蛋白较佳工艺条件为：加酶量0.12g/10mL、pH6.0、反应温度45℃、反应时间1h,在此条件下,蛋白质脱除率达52.36%。透析脱盐的较佳条件为：7000Da、透析时间8h、浒苔多糖溶液浓度6%,在此条件下,灰分脱除率为76.95%。（除杂后,浒苔多糖蛋白质含量为3.43%,灰分含量为8.04%,多糖损失率为18.96%）。结论：蛋白酶法脱蛋白质技术和透析袋流水透析脱盐技术,对浒苔多糖的脱蛋白脱盐效果较好,有一定的推广潜力。     

浒苔多糖铁的制备工艺优化及其结构表征

通过响应面法优化浒苔多糖铁的螯合工艺,并利用红外光谱法和X衍射分析浒苔多糖铁的结构。结果表明,浒苔多糖铁的最佳制备工艺为反应温度54 ℃,螯合时间3.8 h,浒苔多糖与三价铁比例3.9∶1,pH7.5,在此条件下螯合率为19.6%。红外光谱表明浒苔多糖中的O-H和C=O与Fe3+发生了螯合。X射线衍射结果进一步证实了浒苔多糖铁的生成。从而为拓展浒苔资源的高值利用及开发新型多糖补铁剂提供了一定的科学依据。     

磷酸化对鲢小清蛋白抗原性降低的作用

通过建立鱼类主要过敏原小清蛋白（parvalbumin,PV）磷酸化反应的条件,研究磷酸化对PV的抗原性及结构影响。将PV和葡萄糖-6-磷酸二钠盐（D-glucose-6-phosphate disodium salt hydrate,G6P-Na2）混合,在不同质量比、反应时间以及反应温度条件下进行干法磷酸化反应。采用Tricine-十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和Dot-blotting检验其聚合类型以及抗原性变化。通过扫描电镜对聚合物的微观结构进行分析,利用圆二色谱仪和ANS探针法分析磷酸化产物的二级结构和疏水性变化。结果显示,在PV与G6P-Na2质量比为1∶4、反应温度80?℃、反应时间80?min条件下生成的磷酸化产物的抗原性最低。磷酸化反应后,产物呈现聚集状态,其二级结构变化较为明显,疏水性明显提高。磷酸化反应对蛋白结构的改变可能是影响PV抗原性降低的主要原因。     

Glycation and phosphorylation of α-lactalbumin by dry heating: Effect on protein structure and physiological functions

α-Lactalbumin (α-LA) was glycated with maltopentaose (MP) through the Maillard reaction (MP-α-LA) and subsequently phosphorylated by dry heating in the presence of pyrophosphate to investigate its structure and physiological functions. Glycation occurred effectively, and the sugar content of α-LA increased by approximately 22.3% through the Maillard reaction. The phosphorylation of MP-α-LA was enhanced with an increase in the dry-heating time from 1 to 5 d, and the phosphorous content of MP-α-LA increased by approximately 1.01% by dry heating at pH 4.0 and 85°C for 5 d in the presence of pyrophosphate. The electrophoretic mobility of α-LA increased with an increase in the phosphorylation level. The circular dichroism spectra showed that the change in the secondary structure of the α-LA molecule by glycation and subsequent phosphorylation was slight. However, the Trp fluorescence intensity was increased by phosphorylation after glycation. In addition, the differential scanning calorimetry thermograms of α-LA showed that the denaturation temperature of MP-α-LA was decreased by phosphorylation. These results indicated that molten (partially unfolded) conformations of α-LA were formed by dry heating in the presence of pyrophosphate after glycation. The anti-α-LA antibody response was significantly reduced by glycation and subsequent phosphorylation. The suppressive effect of α-LA on the production of proinflammatory cytokines such as IL-6 and tumor necrosis factor-α from THP-1 cells after stimulation with lipopolysaccharide was significantly enhanced by glycation with MP and was further enhanced by phosphorylation after glycation. The Ca phosphate-solubilizing ability of α-LA was enhanced by phosphorylation. The apoptotic activity of α-LA was reduced by glycation and subsequent phosphorylation. These results suggest that phosphorylation by dry heating in the presence of pyrophosphate after glycation with MP through the Maillard reaction is a useful method for improvement of the physiological functions of α-LA.     

虾蛄肌原纤维蛋白磷酸化工艺及其在虾蛄肉制品中的应用

研究多聚磷酸盐磷酸化虾蛄肌原纤维蛋白的最佳工艺条件,并探究复合磷酸盐在虾蛄肉制品中的应用。采用三聚磷酸钠(STP)、焦磷酸钠(SPP)、三偏磷酸钠(STMP)分别对虾蛄肌原纤维蛋白进行磷酸化改性,在单因素实验的基础上,利用响应面法优化虾蛄肌原纤维蛋白磷酸化条件,结果表明:STP最佳工艺条件为:反应pH8.0、STP添加量4%、反应时间2.0 h,所得磷酸化程度为102.87 mg/g;SPP最佳磷酸化条件为:反应pH7.5、SPP添加量2%、反应时间2.0 h,所得磷酸化程度为96.41 mg/g;STMP最佳磷酸化条件为:反应pH8.0、STMP添加量4%、反应时间2.5 h,所得磷酸化程度为76.53 mg/g。在此基础上,探究4组不同比例复合磷酸盐对新型果蔬虾饼品质的影响,结果表明:复合磷酸盐可显著降低虾饼的蒸煮损失和在加工过程中抗坏血酸含量的损失(p<0.05),并提高其质构特性;当STP:SPP:STMP比例为2:1:2时,能显著改善虾蛄肉制品的品质(p<0.05),证实了磷酸化改性的可行性。     

Improvement of functional properties of whey protein isolate through glycation and phosphorylation by dry heating

Whey protein isolate (WPI) was glycated with maltopentaose (MP) through the Maillard reaction, and the MP-conjugated WPI (MP-WPI) was then phosphorylated by dry heating in the presence of pyrophosphate. Glycation occurred efficiently, and the sugar content of WPI increased approximately 19.9% through the Maillard reaction. The phosphorylation of MP-WPI was enhanced with an increase in the dry-heating time from 1 to 5 d, and the phosphorus content of WPI increased approximately 1.05% by dry heating at pH 4.0 and 85°C for 5 d in the presence of pyrophosphate. The electrophoretic mobility of WPI increased with an increase in the phosphorylation level. The stability of WPI against heat-induced insolubility at pH 7.0 was improved by conjugation with MP alone, and further improved by phosphorylation. Although the emulsifying activity of WPI was barely affected by glycation and phosphorylation, the emulsifying stability of phosphorylated MP-WPI (5 d), was 2.2 times higher than that of MP-WPI. Gelling properties such as hardness, resiliency, and water-holding capacity of heat-induced WPI gel were markedly improved, and the gel was rendered transparent by phosphorylation. The calcium phosphate-solubilizing ability of WPI was enhanced by phosphorylation. These results suggested that phosphorylation by dry heating in the presence of pyrophosphate after conjugation with MP is a useful method for improving the functional properties of WPI.     

乳酸乳球菌胞外多糖磷酸化的工艺优化

采用单因素和正交试验,对乳酸乳球菌胞外多糖的磷酸化工艺进行研究,探讨磷酸盐用量、反应温度、反应时间、反应pH值对乳酸乳球菌胞外多糖最终PO43-接枝量的影响。所得的乳酸乳菌球菌胞外多糖磷酸化的工艺优化条件为：胞外多糖与磷酸盐质量比为6:1、温度90℃、时间4h、pH6.0,此条件下所得PO43-的接枝量为1.639mg/g。     

Improvement of functional properties of bovine serum albumin through phosphorylation by dry-heating in the presence of pyrophosphate

ABSTRACT:  Bovine serum albumin (BSA) was phosphorylated by 2 methods. One is dry-heating in the presence of pyrophosphate, and the other is conjugation with maltopentaose through the Maillard reaction and subsequent dry-heating in the presence of pyrophosphate. The phosphorus content of BSA was increased to approximately 0.45% by dry-heating at pH 4.0 and 85 °C for 5 d in the presence of pyrophosphate, and approximately 0.91% by glycation and subsequent phosphorylation. The circular dichroism spectra showed that the change of secondary structure in the BSA molecule by phosphorylation was mild. However, tryptophan fluorescence intensity of BSA decreased by phosphorylation. The differential scanning calorimetry thermograms of BSA showed a disappearing of the 1st peak and a lowering of the 2nd peak denaturation temperature by phosphorylation. These results indicated molten (partially unfolded) conformations of BSA formed by both phosphorylation methods. The functional properties of BSA such as heat stability and calcium phosphate solubilizing ability were improved by phosphorylation alone and further by phosphorylation after glycation. Transparent gels of BSA with relatively high water-holding capacity were obtained by phosphorylation alone, and the immunogenicity of BSA was reduced significantly by glycation and phosphorylation, respectively.     

酶法与非酶法磷酸化改性食品蛋白质的研究进展

磷酸化是一种重要的蛋白质修饰改性手段,可以有效改善食物蛋白质的功能性质,如乳化性、溶解性、凝胶性、热稳定性等。蛋白质磷酸化改性方法主要可分为酶法与非酶法两种,其中酶法磷酸化常使用蛋白激酶作为磷酸基团的供体,对蛋白质进行修饰。主要的蛋白激酶包括环磷酸腺苷依赖蛋白激酶(CAMPdPK)和酪蛋白激酶Ⅱ(CK-Ⅱ)。非酶法磷酸化常见的改性试剂主要包括三氯氧磷、焦磷酸钠、三聚磷酸钠和葡萄糖-6-磷酸等。本文对近年来蛋白质磷酸化的有关报道进行了总结归纳,并通过介绍磷酸化蛋白的磷酸键性质、磷酸化肽段及位点的鉴定,阐述了不同磷酸化试剂的反应机制及其对蛋白质构效关系的影响,为通过定向的磷酸化修饰而改善蛋白质特定的功能性质提供了理论依据与参考。     

磷酸化大豆多肽螯合钙的制备工艺优化及其对成骨细胞的活性影响

为了探究磷酸化大豆多肽螯合钙（PSPCC）的生理活性,提高磷酸化大豆多肽螯合钙的钙螯合量,本研究通过单因素实验和响应面试验相结合的方式优化磷酸化大豆多肽螯合钙的制备工艺,并通过体外实验评价了磷酸化大豆多肽螯合钙对成骨细胞的活性影响。结果表明,制备磷酸化大豆多肽螯合钙的最佳工艺条件为:三聚磷酸钠与大豆多肽的质量比1:2,磷酸化反应温度52 ℃,磷酸化反应pH7.0,磷酸化反应时间9.7 h,磷酸化大豆多肽与氯化钙的质量比2:1,螯合反应pH8.0,螯合反应温度50 ℃,螯合反应时间1.5 h,钙螯合量最大为107.25±0.10 mg/g;MTT法结果显示,磷酸化大豆多肽螯合钙组（D组）成骨细胞相对增殖率是大豆多肽组（A组）的1.6倍（第3 d）;ALP染色实验表明,D组染色阳性率是空白组的33.6倍。采用ALP试剂盒对各样品第7 d的ALP活性进行检测,结果显示D组ALP活力是空白组的6.2倍。通过茜素红染色实验发现,D组钙结节数量是空白组的94倍。本研究使用响应面法对磷酸化大豆多肽螯合钙的制备工艺优化合理,并初步证明了磷酸化大豆多肽螯合钙对成骨细胞具有显著的促增殖、分化作用（P<0.05）,且作用效果高于其它样品,为后续磷酸化大豆多肽螯合钙的进一步开发利用提供了一定的理论基础。     

Improvement of the Functionalities of Soy Protein Isolate through Chemical Phosphorylation

A method of chemical phosphorylation was developed to modify soy protein so as to improve its functional properties. The reaction was carried out by incubating soy protein isolate and cyclic sodium trimetaphosphate in an aqueous solution at pH 11.5 and 35°C for about 3 hours. The reactions ensued were the phosphoesterification of serine residues and the phosphoramidation of lysine residues in soy protein. The phosphorylated soy protein isolate prepared there-from exhibited much improved functional properties in terms of aqueous solubility, water-holding capacity, emulsifiability and whippability. The nutritive bioavailability of soy protein isolate was not impaired by phosphorylation.     

花生分离蛋白磷酸化改性研究

采用响应面优化法对花生分离蛋白进行磷酸化改性,以氮溶解指数(NSI)为指标得出花生分离蛋白磷酸化改性的最佳条件为三聚磷酸钠添加量7.77%、花生分离蛋白质量分数6.38%、反应温度44.85℃、反应体系pH8.24、反应时间5.68h。得到的花生改性蛋白NSI 最大值为77.74%。改性后,花生分离蛋白的吸油性、吸水性、持水性、乳化性、乳化稳定性、泡沫稳定性都有不同程度的提高。