羊乳和牛乳理化特性比較及其摻假檢測(cè)研究
發(fā)布時(shí)間：2019-07-10 09:44
【摘要】：乳制品的質(zhì)量與安全問(wèn)題的核心就是控制好乳源。本課題對(duì)羊乳和牛乳的蛋白質(zhì)含量及組分、酒精穩(wěn)定性和凝乳特性做了比較研究，并且以牛α-乳白蛋白和α-酪蛋白為目標(biāo)蛋白，建立了可以快速、靈敏、準(zhǔn)確地檢測(cè)羊乳中摻入牛乳含量的直接競(jìng)爭(zhēng)ELISA法。 羊乳和牛乳理化特性比較研究。以脫脂的羊乳和牛乳為樣品，通過(guò)凱氏定氮法和SDS-聚丙烯酰胺凝膠電泳測(cè)定羊乳和牛乳的蛋白質(zhì)含量及其蛋白質(zhì)組分。測(cè)得，羊乳的蛋白質(zhì)含量高于牛乳的，分別為（3.43±0.06）g/100mL和（3.19±0.07）g/100mL（置信度95%）；兩者的蛋白組分也存在一定的差異，通過(guò)圖譜可以看出，牛乳的酪蛋白中以α-酪蛋白含量最多，而羊乳中以β-酪蛋白含量最多。準(zhǔn)備原料乳、巴殺乳和冷凍復(fù)融乳乳樣，測(cè)得羊乳乳樣的酒精穩(wěn)定性值依次為46±1、42±2、40±1（%,V/V），凝乳時(shí)間依次為297±2、310±2、266±2（S）；牛乳乳樣的酒精穩(wěn)定性值依次為78±1、78±1、78±1（%,V/V）；凝乳時(shí)間依次為571±3、590±1、485±1（S）。實(shí)驗(yàn)證明，羊乳的膠體穩(wěn)定性比牛乳的差。 兔抗牛α-乳白蛋白和α-酪蛋白抗體的制備與特性研究。以牛α-乳白蛋白和α-酪蛋白為抗原，免疫新西蘭大白兔，采集高效價(jià)血清。依次采用飽和硫酸銨分級(jí)沉淀法和蛋白A親和色譜法純化血清抗體，得到純化兔抗牛α-LA-IgG和α-CN-IgG樣品。效價(jià)均可達(dá)到1:16000，蛋白質(zhì)含量分別為0.35mg/mL，0.43mg/mL，純度分別為88.31%、89.23%。硫氰酸銨洗脫法測(cè)得α-LA-IgG和α-CN-IgG對(duì)α-LA的親和力指數(shù)分別為2.6mol/L、1.1mol/L，對(duì)α-CN的親和力指數(shù)分別為1.6mol/L、2.5mol/L。anti-α-LA-IgG特異性很好，anti-α-CN-IgG與κ-CN和β-CN存在一定交叉反應(yīng)，兩種抗體均與羊乳存在一定的交叉反應(yīng)。經(jīng)抗體修飾技術(shù)處理后，這些交叉反應(yīng)均明顯降低。 牛乳α-乳白蛋白和α-酪蛋白的酶標(biāo)記。選用辣根過(guò)氧化物酶，通過(guò)過(guò)碘酸鈉氧化法分別標(biāo)記α-乳白蛋白和α-酪蛋白。結(jié)果顯示，當(dāng)HRP與α-LA的分子個(gè)數(shù)比為1.5:1時(shí)，標(biāo)記效果比較好（結(jié)合率25.4%，，標(biāo)記率85.7%）；α-CN的分子個(gè)數(shù)比也為1.5:1時(shí)，標(biāo)記效果比較好（結(jié)合率43.8%，標(biāo)記率84.49%）。經(jīng)ELISA檢測(cè)，酶標(biāo)抗原的效價(jià)分別可以達(dá)到1:320和1:640。 直接競(jìng)爭(zhēng)ELISA方法的建立。棋盤滴定法確定抗體最佳包被濃度（anti-α-LA-IgG和anti-α-CN-IgG分別為1:160,1:320）和酶標(biāo)抗原最佳稀釋濃度（都為1:20）。確定封閉液1%G的體積為200μL，封閉時(shí)間為1h。脫脂乳的最佳稀釋度為1:80。用anti-α-LA-IgG和anti-α-CN-IgG分別建立直接競(jìng)爭(zhēng)ELISA法檢測(cè)羊乳中摻入的牛乳含量。經(jīng)評(píng)價(jià)指標(biāo)分析，該方法的檢測(cè)線性范圍都為0%~50%（V/V），最低檢測(cè)限分別為2.5%和4%，變異系數(shù)小于5%。 本研究通過(guò)羊乳和牛乳的理化特性及其膠體穩(wěn)定性的比較，為科學(xué)合理地利用羊乳，生產(chǎn)具有獨(dú)特功能的羊乳制品提供了理論依據(jù)；建立的基于牛乳α-LA和α-CN的直接競(jìng)爭(zhēng)ELISA檢測(cè)法，特異性、重復(fù)性、敏感性都比較高，可以作為檢測(cè)羊乳及其制品中摻入牛乳檢測(cè)的技術(shù)手段。
文內(nèi)圖片：
圖片說(shuō)明：亞膠束模型中酪蛋白膠束的結(jié)構(gòu)
[Abstract]:The core of the quality and safety of dairy products is to control the milk source. The protein content and composition, alcohol stability and curd property of goat milk and milk were compared and studied. And the direct competition ELISA method for accurately detecting the milk content of the goat milk. Comparative Study on the Physical and Chemical Properties of Goat Milk and Milk The protein content of goat milk and milk and its protein group were determined by the method of Kjeldahl and SDS-polyacrylamide gel electrophoresis. The protein content of goat milk was higher than that of cow's milk (3.43-0.06) g/100 mL and (3.19-0.07) g/100 mL (95% confidence). in addition, in that goat milk, the content of the HCO3-casein is the most The alcohol stability of milk-like milk samples was 46,1,42,2,40,1 (%, V/ V), and the time of curd was 297, 2,310, 2,266 and 2 (S). The alcohol stability of milk-like milk was 78%,78%,78%,1 (%, V/ V). ); the time of the curd is 571-3,590-1,485-1 (S The experimental results show that the colloidal stability of the goat milk is higher than that of the milk. The preparation of rabbit anti-bovine serum-milk albumin and the anti-bovine-casein antibody and the preparation of the same Study on the sex of New Zealand white rabbits with bovine serum-milk albumin and calcium-casein as antigen, and high efficiency the serum antibody is purified by adopting a saturated sulfuric acid precipitation method and a protein A affinity chromatography in sequence, and the purified rabbit anti-bovine serum-LA-IgG and the antigen-CN-Ig are obtained The titer of G samples can reach 1:16000, the protein content is 0.35 mg/ mL, 0.43 mg/ mL, and the purity is 88.31%,89. The affinity index of L-LA-IgG and HCO3-CN-IgG to the antigen-LA was 2.6 mol/ L, 1.1 mol/ L, 1.6 mol/ L and 2.5 mol/ L respectively. The reaction of the two kinds of antibodies has a certain relation with the goat milk. The cross reaction is clear after treatment with the antibody modification technology. significantly reduced milk-milk-milk albumin and egg-casein White enzyme marker. Horseradish Peroxidase is used, and the sodium peroxide-milk albumin is respectively marked by the sodium periodate oxidation method. The results showed that when the molecular weight ratio of HRP to LA-LA was 1.5:1, the labeling effect was good (the binding rate was 25.4%, the labeling rate was 85.7%), and the number of the molecular number of the E-CN was also 1.5:1, the labeling effect was good (the binding rate was 43.8%, the labeling rate was 84). .49%). The titer of the enzyme-labeled antigen can be 1:320 and 1:320, respectively. 1:640. Direct competition ELI The best coating concentration of the antibody was determined by the checkerboard titration (1:160,1:320) and the optimal concentration of the enzyme-labeled antigen (1:160,1:320) and the best dilution of the enzyme-labeled antigen (1:160,1:320, respectively). the volume of the blocking solution 1% g was determined to be 200. m u.l, The closed time is 1 h. The best dilution of skim milk The release was 1:80. The direct competitive ELISA was used to detect the goat milk with anti-antigen-LA-IgG and anti-antigen-CN-IgG, respectively. The detection linear range of the method is 0% ~ 50% (V/ V) and the minimum detection limit is 2.5% and 4%, respectively. Through the comparison of the physical and chemical properties and the colloidal stability of the goat milk and the milk, this study provides a theoretical basis for the scientific and reasonable utilization of the goat milk and the production of the sheep milk products with unique functions; and the establishment of the direct competition ELI based on the bovine milk-LA and the L-CN The SA detection method is high in specificity, repeatability and sensitivity, and can be used as a detection goat milk and a product thereof.
【學(xué)位授予單位】：陜西科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R392
本文編號(hào)：2512521
文內(nèi)圖片：
圖片說(shuō)明：亞膠束模型中酪蛋白膠束的結(jié)構(gòu)
[Abstract]:The core of the quality and safety of dairy products is to control the milk source. The protein content and composition, alcohol stability and curd property of goat milk and milk were compared and studied. And the direct competition ELISA method for accurately detecting the milk content of the goat milk. Comparative Study on the Physical and Chemical Properties of Goat Milk and Milk The protein content of goat milk and milk and its protein group were determined by the method of Kjeldahl and SDS-polyacrylamide gel electrophoresis. The protein content of goat milk was higher than that of cow's milk (3.43-0.06) g/100 mL and (3.19-0.07) g/100 mL (95% confidence). in addition, in that goat milk, the content of the HCO3-casein is the most The alcohol stability of milk-like milk samples was 46,1,42,2,40,1 (%, V/ V), and the time of curd was 297, 2,310, 2,266 and 2 (S). The alcohol stability of milk-like milk was 78%,78%,78%,1 (%, V/ V). ); the time of the curd is 571-3,590-1,485-1 (S The experimental results show that the colloidal stability of the goat milk is higher than that of the milk. The preparation of rabbit anti-bovine serum-milk albumin and the anti-bovine-casein antibody and the preparation of the same Study on the sex of New Zealand white rabbits with bovine serum-milk albumin and calcium-casein as antigen, and high efficiency the serum antibody is purified by adopting a saturated sulfuric acid precipitation method and a protein A affinity chromatography in sequence, and the purified rabbit anti-bovine serum-LA-IgG and the antigen-CN-Ig are obtained The titer of G samples can reach 1:16000, the protein content is 0.35 mg/ mL, 0.43 mg/ mL, and the purity is 88.31%,89. The affinity index of L-LA-IgG and HCO3-CN-IgG to the antigen-LA was 2.6 mol/ L, 1.1 mol/ L, 1.6 mol/ L and 2.5 mol/ L respectively. The reaction of the two kinds of antibodies has a certain relation with the goat milk. The cross reaction is clear after treatment with the antibody modification technology. significantly reduced milk-milk-milk albumin and egg-casein White enzyme marker. Horseradish Peroxidase is used, and the sodium peroxide-milk albumin is respectively marked by the sodium periodate oxidation method. The results showed that when the molecular weight ratio of HRP to LA-LA was 1.5:1, the labeling effect was good (the binding rate was 25.4%, the labeling rate was 85.7%), and the number of the molecular number of the E-CN was also 1.5:1, the labeling effect was good (the binding rate was 43.8%, the labeling rate was 84). .49%). The titer of the enzyme-labeled antigen can be 1:320 and 1:320, respectively. 1:640. Direct competition ELI The best coating concentration of the antibody was determined by the checkerboard titration (1:160,1:320) and the optimal concentration of the enzyme-labeled antigen (1:160,1:320) and the best dilution of the enzyme-labeled antigen (1:160,1:320, respectively). the volume of the blocking solution 1% g was determined to be 200. m u.l, The closed time is 1 h. The best dilution of skim milk The release was 1:80. The direct competitive ELISA was used to detect the goat milk with anti-antigen-LA-IgG and anti-antigen-CN-IgG, respectively. The detection linear range of the method is 0% ~ 50% (V/ V) and the minimum detection limit is 2.5% and 4%, respectively. Through the comparison of the physical and chemical properties and the colloidal stability of the goat milk and the milk, this study provides a theoretical basis for the scientific and reasonable utilization of the goat milk and the production of the sheep milk products with unique functions; and the establishment of the direct competition ELI based on the bovine milk-LA and the L-CN The SA detection method is high in specificity, repeatability and sensitivity, and can be used as a detection goat milk and a product thereof.
【學(xué)位授予單位】：陜西科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R392
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