Mant C T, Burke T W, Black J A, Hodges R S
Department of Biochemistry, University of Alberta, Edmonton, Canada.
J Chromatogr. 1988 Dec 23;458:193-205. doi: 10.1016/s0021-9673(00)90564-8.
The use of amino acid retention or hydrophobicity coefficients for the prediction of peptide retention time and/or the elution order on hydrophobic stationary phases is based on the premise that amino acid composition is the major factor affecting peptide retention in reversed-phase chromatography. Although this assumption generally agrees well for small peptides (up to ca. 15 residues), the retention times of increasingly larger peptides are less than expected from a simple summation of retention coefficients. In the present study, we report the synthesis of four series of peptide polymers which vary significantly in overall hydrophobicity and polypeptide chain length (5-50 amino acid residues, Ac = acetyl): Ac-(G-L-G-A-K-G-A-G-V-G)n-amide (n = 1-5), Ac-(G-K-G-L-G)n-amide (n = 1, 2, 4, 6, 8, 10), Ac-(L-G-L-K-A)n-amide (n = 1, 2, 4, 6, 8, 10) and Ac-(L-G-L-K-L)n-amide (n = 1, 2, 4). From the retention behaviour of these peptide polymers on C4, C8 and C18 stationary phases under gradient elution conditions, we have clearly established the effect of polypeptide chain length and hydrophobicity on peptide retention. This, in turn, has enabled us to extend the utility of retention time prediction for peptides containing up to 50 residues by introducing a peptide chain-length correction.
利用氨基酸保留系数或疏水性系数预测肽在疏水固定相上的保留时间和/或洗脱顺序，其前提是氨基酸组成是反相色谱中影响肽保留的主要因素。尽管这一假设对于小肽（最多约15个残基）通常非常适用，但越来越大的肽的保留时间却比保留系数简单相加所预期的要短。在本研究中，我们报道了四类肽聚合物的合成，它们在整体疏水性和多肽链长度上有显著差异（5 - 50个氨基酸残基，Ac = 乙酰基）：Ac-(G-L-G-A-K-G-A-G-V-G)n-酰胺（n = 1 - 5）、Ac-(G-K-G-L-G)n-酰胺（n = 1、2、4、6、8、10）、Ac-(L-G-L-K-A)n-酰胺（n = 1、2、4、6、8、10）和Ac-(L-G-L-K-L)n-酰胺（n = 1、2、4）。通过这些肽聚合物在梯度洗脱条件下在C4、C8和C18固定相上的保留行为，我们明确确定了多肽链长度和疏水性对肽保留的影响。反过来，这使我们能够通过引入肽链长度校正来扩展对含多达50个残基的肽的保留时间预测的实用性。
