Compartmentalization and quantitation of protein in human milk.
Compartmentalization and quantitation of protein in human milk.
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DOI：
10.1093/jn/117.8.1385
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发表时间：
1987-08
期刊：
影响因子：
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通讯作者：
B. Lönnerdal;Leslie R. Woodhouse;C. Glazier
中科院分区：
文献类型：
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作者：
B. Lönnerdal;Leslie R. Woodhouse;C. Glazier
用三种比色法和凯氏定氮法测定人乳蛋白。氮（N）和蛋白质的分布在不同的牛奶隔室。通过一系列离心和超离心分离后，用微量凯氏定氮法分析总氮、乳清、酪蛋白、非蛋白氮（NPN）、细胞氮和脂肪组分中的氮。将新鲜乳样品（初乳、过渡乳和成熟乳）以500 X g离心分离乳细胞，并以5000 X g离心脱脂乳。将脱脂乳和脱脂乳在189，000 X g下超浓缩以从乳清中分离脂肪和酪蛋白胶束。在三氯乙酸沉淀后分析NPN。用Lowry方法分析全脂奶、去角质奶、脱脂奶和乳清中的蛋白质，该方法针对含脂肪样品进行了修改，Bradford染料结合测定法（Bio-Rad）和Pierce二喹啉甲酸（BCA）测定法。成熟乳中的细胞氮有低于初乳的趋势。初乳中酪蛋白含量仅为6%，而成熟乳中酪蛋白含量为13%。脱脂脂肪的N含量低于超浓缩脂肪。三个泌乳期的平均NPN水平相似，占初乳N的10%和成熟乳N的25%。当与凯氏定氮值相关时，通过Bio-Rad方法测定的全脂牛奶样品中的蛋白质具有最低的变异性（平方根MSE）。所有三种测定法在分析乳清和脱脂乳时的变异性均低于分析全脂乳时的变异性。Lowry方法和Bio-Rad方法对乳清和脱脂乳样品的变异性较低，但Lowry方法产生的分析值最接近凯氏定氮法蛋白值。BCA方法始终高估凯氏定氮蛋白30%。
Human milk protein was determined by three colorimetric methods and by Kjeldahl analysis. The distribution of nitrogen (N) and protein was determined within various milk compartments. Total N, whey, casein, nonprotein nitrogen (NPN), cell N and N in the fat fraction were analyzed by micro-Kjeldahl analysis after a series of centrifugation and ultracentrifugation separations. Fresh milk samples (colostrum, transitional milk and mature milk) were centrifuged at 500 X g to separate milk cells and at 5000 X g to skim the milk. Decelled milk and skimmed milk were ultracentrifuged at 189,000 X g to separate fat and casein micelles from whey. NPN was analyzed after trichloroacetic acid precipitation. Whole milk, decelled milk, skimmed milk and whey were analyzed for protein with the Lowry method, modified for fat-containing samples, the Bradford dye-binding assay (Bio-Rad) and the Pierce bicinchoninic acid (BCA) assay. Cell nitrogen had a tendency to be lower in mature milk than in colostrum. Colostrum contained only 6% casein protein, whereas mature milk contained 13%. Fat from skimming was lower in N than fat from ultracentrifugation. Average NPN levels were similar for milk from all three lactation periods, and constituted 10% of colostrum N and 25% of mature milk N. Protein determined by the Bio-Rad method on whole milk samples had the lowest variability (square root MSE) when correlated to Kjeldahl values. All three assays had lower variability when analyzing whey and skimmed milk than when analyzing whole milk. The Lowry method and the Bio-Rad method had low variability for whey and skimmed milk samples, but the Lowry method yielded analytical values closest to Kjeldahl protein values. The BCA method consistently overestimated Kjeldahl protein by 30%.