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The perchlorate anion is more effective than the trifluoroacetate anion as an ion-pairing reagent for reversed-phase chromatography of peptides

机译：高氯酸根阴离子比三氟乙酸根阴离子作为肽反相色谱的离子对试剂更有效

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摘要

The addition of salts, specifically sodium perchlorate (NaClO4), to mobile phases at acidic pH as ion-pairing reagents for reversed-phase high-performance liquid chromatography (RP-HPLC) has been generally overlooked. To demonstrate the potential of NaClO4 as an effective anionic ion-pairing reagent, we applied RP-HPLC in the presence of 0–100 mM sodium chloride (NaCl), sodium trifluoroacetate (NaTFA) or NaClO4 to two mixtures of synthetic 18-residue peptides: a mixture of peptides with the same net positive charge (+4) and a mixture of four peptides of +1, +2, +3 and +4 net charge. Interestingly, the effect of increasing NaClO4 concentration on increasing peptide retention times and selectivity changes was more dramatic than that of either NaCl or NaTFA, with the order of increasing anion effectiveness being Cl− ≪TFA− < ClO4−. Such effects were more marked when salt addition was applied to eluents containing 10 mM phosphoric acid (H3PO4) compared to 10 mM trifluoroacetic acid (TFA) due to the lesser starting anion hydrophobicity of the former mobile phase (containing the phosphate ion) compared to the latter (containing the TFA− ion).
机译：人们普遍忽略了在酸性pH下向流动相中添加盐，特别是高氯酸钠（NaClO4）作为反相高效液相色谱（RP-HPLC）的离子对试剂。为了证明NaClO4作为有效的阴离子配对试剂的潜力，我们在0–100 mM氯化钠（NaCl），三氟乙酸钠（NaTFA）或NaClO4的存在下，对两种合成的18残基肽混合物应用RP-HPLC ：具有相同净正电荷（+4）的肽的混合物以及具有+ 1，+ 2，+ 3和+4净电荷的四个肽的混合物。有趣的是，增加NaClO4浓度对增加肽保留时间和选择性变化的影响比NaCl或NaTFA更为显着，阴离子有效性增加的顺序为Cl - ≪TFA - -。当将盐添加到含有10 mM磷酸（H3PO4）而不是10 mM三氟乙酸（TFA）的洗脱液中时，由于前者流动相（含磷酸根离子）的起始负离子疏水性比苯甲酸酯低，因此这种效果更加明显。后者（包含TFA -离子）。

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