卵黄高磷蛋白及其磷酸肽的制备和研究
摘要
以鸡蛋黄中卵黄高磷蛋白(PV)为主要研究对象，在Losso．J．N和Naikai．S方法的基础上，通过实验提出了一种从鸡蛋黄中分离食品级卵黄高磷蛋白的新工艺。该工艺操作简便，且综合考虑了蛋黄中其它成分的利用。并研究了两种蛋白酶复合水解卵黄高磷蛋白的工艺条件，及其水解产物阻止Ca~(2+)形成沉淀的性能。
针对鸡蛋黄的成分，先用水去除卵黄高磷蛋白以外的可溶性蛋白质，再用有机溶剂去除脂类，最后用NaCl溶液提取卵黄高磷蛋白。实验研究结果表明：从10g鸡蛋黄中提取卵黄高磷蛋白时，在用水量为100mL；乙醇与正己烷混合液为20mL；乙醇：正己烷(V／V)为1∶1；NaCl溶液的浓度为1.75 mol／L的条件下卵黄高磷蛋白提取率最高。
探讨了碱性蛋白酶2709和中性蛋白酶AS1.398复合水解卵黄高磷蛋白的工艺条件。结果表明：用NaOH进行前处理15 min后，当样品在溶液温度保持50℃，底物浓度为2％，碱性蛋白酶2709浓度为4％，溶液pH值为11时的条件下水解150min；再在中性蛋白酶AS1.398浓度为6％，溶液pH值为7.5的条件下水解60min，卵黄高磷蛋白的相对水解度最好。
卵黄高磷蛋白水解产物为卵黄高磷蛋白磷酸肽(PPP)，研究了它阻止Ca~(2+)形成沉淀的性能，并与酪蛋白磷酸肽进行了比较。实验结果表明卵黄高磷蛋白磷酸肽可在一定程度上阻止Ca~(2+)形成沉淀，其效果较酪蛋白磷酸肽好。
The new technology was put forword to extract phosvitin (PV) from egg yolk based on the methods of Losso. J. N and Naikai. S. The technology was simple with no other accretion, and the other components of egg yolk were considered to all-round using. The process conditions of hydrolysis of phosvitin by two types of proteolytic enzyme were studied. The performance of the production to prevent Ca2 + from deposition was studied too.
For the components of egg yolk, water soluble proteins except phosvitin were removed with water firstly, then the lipids were eliminated with organic solvent, at last the phosvitin was isolated from the resulted granules with NaCl solution. Through experiments the optimal processing conditions were obtained as follows: the dosage of water and the organic solvent were 100 mL and 20 mL respectively, the ratio of ethonal and n-hexane was 1 : 1 (V/V) , the NaCl solution concentration was 1.75 mol/L.
The technical conditions for hydrolysis of phosvitin by 2709 alkalescence and AS 1.398 neutral proteolytic enzyme were studied. The result showed that the degree of hydrolysis was high when phosvitin was hydrolysised as follows: phosvitin was dealed with NaOH for 15 minute firstly, then hydrolyzed by alkalescence proteolytic enzyme 2709 for 150 minutes and hydrolyzed by neutral proteolytic enzyme AS 1.398 for 60 minutes at 50 centigrade when the substrate concentration was 2%. The optimum pH value and concentration for alkalescence proteolytic enzyme 2709 was pHll and 5%, for neutral proteolytic enzyme AS 1.398 was pH7.5 and 6% respectively.
The production of hydrolyzing phosvitin was phosvitin phosphopeptides (PPP). The performance of ppp to prevent Ca2+ from deposition was studied and compared with casein phosphopeptides (cpp). The result showed that PPP could prevent Ca2+ from deposition at a certain degree and the effect was better.
Liu Xiaoli (Chemical Biology engineering Directed by Professor Zhang Xiaoyan
针对鸡蛋黄的成分，先用水去除卵黄高磷蛋白以外的可溶性蛋白质，再用有机溶剂去除脂类，最后用NaCl溶液提取卵黄高磷蛋白。实验研究结果表明：从10g鸡蛋黄中提取卵黄高磷蛋白时，在用水量为100mL；乙醇与正己烷混合液为20mL；乙醇：正己烷(V／V)为1∶1；NaCl溶液的浓度为1.75 mol／L的条件下卵黄高磷蛋白提取率最高。
探讨了碱性蛋白酶2709和中性蛋白酶AS1.398复合水解卵黄高磷蛋白的工艺条件。结果表明：用NaOH进行前处理15 min后，当样品在溶液温度保持50℃，底物浓度为2％，碱性蛋白酶2709浓度为4％，溶液pH值为11时的条件下水解150min；再在中性蛋白酶AS1.398浓度为6％，溶液pH值为7.5的条件下水解60min，卵黄高磷蛋白的相对水解度最好。
卵黄高磷蛋白水解产物为卵黄高磷蛋白磷酸肽(PPP)，研究了它阻止Ca~(2+)形成沉淀的性能，并与酪蛋白磷酸肽进行了比较。实验结果表明卵黄高磷蛋白磷酸肽可在一定程度上阻止Ca~(2+)形成沉淀，其效果较酪蛋白磷酸肽好。
The new technology was put forword to extract phosvitin (PV) from egg yolk based on the methods of Losso. J. N and Naikai. S. The technology was simple with no other accretion, and the other components of egg yolk were considered to all-round using. The process conditions of hydrolysis of phosvitin by two types of proteolytic enzyme were studied. The performance of the production to prevent Ca2 + from deposition was studied too.
For the components of egg yolk, water soluble proteins except phosvitin were removed with water firstly, then the lipids were eliminated with organic solvent, at last the phosvitin was isolated from the resulted granules with NaCl solution. Through experiments the optimal processing conditions were obtained as follows: the dosage of water and the organic solvent were 100 mL and 20 mL respectively, the ratio of ethonal and n-hexane was 1 : 1 (V/V) , the NaCl solution concentration was 1.75 mol/L.
The technical conditions for hydrolysis of phosvitin by 2709 alkalescence and AS 1.398 neutral proteolytic enzyme were studied. The result showed that the degree of hydrolysis was high when phosvitin was hydrolysised as follows: phosvitin was dealed with NaOH for 15 minute firstly, then hydrolyzed by alkalescence proteolytic enzyme 2709 for 150 minutes and hydrolyzed by neutral proteolytic enzyme AS 1.398 for 60 minutes at 50 centigrade when the substrate concentration was 2%. The optimum pH value and concentration for alkalescence proteolytic enzyme 2709 was pHll and 5%, for neutral proteolytic enzyme AS 1.398 was pH7.5 and 6% respectively.
The production of hydrolyzing phosvitin was phosvitin phosphopeptides (PPP). The performance of ppp to prevent Ca2+ from deposition was studied and compared with casein phosphopeptides (cpp). The result showed that PPP could prevent Ca2+ from deposition at a certain degree and the effect was better.
Liu Xiaoli (Chemical Biology engineering Directed by Professor Zhang Xiaoyan
引文
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2 马美湖，葛长荣，罗欣等．动物性食品加工学，北京：中国轻工业出版社，2003
3 W.J.Stadelman．鸡蛋的新用途．国外畜牧科技，1999，26(6)：41-42
4 朱曜．禽蛋研究．北京：科学出版社，1985
5 郑建仙．功能性食品(第二卷)．北京：中国轻工业出版社，1999，235-236
6 林元藻．生化制药学．人民卫生出版社，1998
7 曹文军．用超临界CO_2流体萃取蛋黄粉中卵磷脂的研究．北京林业大学学报，1998，20(5)：1-6
8 龙中儿．卵黄免疫球蛋白IgY的研究及应用．中国生物制品学杂志，1996，9(2)：91-92
9 陈有亮，杨玉爱．功能性鸡蛋．广东微量元素科学，1999，6(8)：1-2
10 赵景联，宋克风．蛋壳制备乳酸钙的研究．现代化工，1997，1：31-33
11 郝素娥，宁明华．利用蛋壳制备丙酸钙的研究．化学研究及应用，1999，1：105-107
12 陈永红，陈欣，徐冬梅．用蛋壳制备柠檬酸钙的研究．化学世界，1999，7：366-369
13 罗有福，盛绍基．鸡蛋膜的最新研究进展．云南化工，2002，29(2)：21-23
14 何唯平，高速，王鸿翔．酪蛋白钙肽在食品中的应用．中国乳品工业，1998，26(6)：44-45
15 汤亚杰，吴思方．酪蛋白磷酸肽的研究进展．食品科学，1998，19(5)：3-6
16 葛可佑，闫怀成．90年代中国人群的膳食与营养状况．北京：人民卫生出版社，1996：89
17 Taborsky G, Mok C.Phosvitin: Homogeneity of molecular weight. J Biol Chem, 1967, 242: 1495-1501
18 Taborsky G. Phosvitin. Adv Inorg Biochem, 1983(5): 235-279
19 Morris E R, Greene F E. Utilization of the iron of egg yolk for hemoglobin
formation by the growing rat.J Nutr,1972,102:901-908
20倪莉，王璋，许时婴．鸡蛋中卵黄高磷蛋白的研究．食品工业，1999，3：20-24
21高真．蛋制品工艺学．北京：中国商业出版社，1992
22 Mechmdk,Olcotths.Phosvitin the principal phosphoprotein of egg yolk.J Am Chem Soc,1949,71:3670-3679
23 Goulas A,Triplett E L,Taborsky G.Oligophosphopeptides of varied structural complexity derived from the egg phosphoprotein,phosvitin.J.Protein Chem,1996,15:1-9
24 Takatoshi iton,Yutaka Abe,Susumu Adachi.Comparative studies on the a and 3 phosvitin from hen's egg yolk.Journal of Food Science,1983,48:1755-1757
25 M.Anton,M.Ledenmat,G.Gandemer.Thermostability of Hen Egg Yolk Granules:Contribution of Native Structure of Granules.Journal of Food Science,2000,65(40) :581-584
26 M.Le Denmat,M.Anton,and G.Gandemer.Protein Denaturation and Emulsifying Properties of Plasma and Granules of Egg Yolk as Related to Heat Treatment.Journal of Food Science,1999,64(2) :194-197
27 Nakamura S,Ogawa M,Nakai S,Kato.S.Antioxidant activity of a Maillard type phosvitin-galactomannan conjugate with emulsifying properties and heat stability.J.Agric.Food Chem.1998,46:3958-3963
28 A.Satter Khan,Elfadil E.Babiker,H.Azakami and A.Kato.Molecular mechanism of the excellent emulsifying properties of phosvitin-galactomannan conjugate.J.Agric.Food Chem,1999,47:2262-2266
29 S.Uma Maheswari,Candadai S.Ramadoss,P.R.Krishnaswamy.Inhibition of Fe(n)catalyzed linoleic acid oxidation and DNA damage by phosvitin.Molecular and Cellular Biochemistry,1997,177:47-51
30 Lu Chong-Ling,Robert C Barket.Effect of pH and food ingredients on stability of egg yolk phospholipids and the metal chelator antioxidant activity of phosvitin.Journal of Food Science,1987,52(3) :613-616
31 Joubert J,Cook W H.Preparation and characterization of phosvitin from
hen egg yolk. Can J Biochem Physiol. 1985, 36:399-408
32 Sundararajan A, Sampath Kumar K S V, Sarma P S. A simplified procedure for the preparation of phosvitin and vitellin. Biochim Biophys Acta.1960,38:360-362
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