State Key Laboratory of Coordination Chemistry, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, Jiangsu, China.
Langmuir. 2011 Oct 4;27(19):12058-68. doi: 10.1021/la202267p. Epub 2011 Sep 7.
Infrared spectroscopy was applied to investigate the well-known EDC/NHS (N-ethyl-N'-(3-(dimethylamino)propyl)carbodiimide/N-hydroxysuccinimide) activation details of poly(acrylic acid) (PAA) and poly(methacrylic acid) (PMAA) brushes grafted on porous silicon. Succinimidyl ester (NHS-ester) is generally believed to be the dominant intermediate product, conveniently used to immobilize biomolecules containing free primary amino groups via amide linkage. To our surprise, the infrared spectral details revealed that the EDC/NHS activation of PMAA generated anhydride (estimated at around 76% yield and 70% composition), but not NHS-ester (around 5% yield and 11% composition) under the well-documented reaction conditions, as the predominant intermediate product. In contrast, EDC/NHS activation of PAA still follows the general rule, i.e., the expected NHS-ester is the dominant intermediate product (around 45% yield and 57% composition), anhydride the side product (40% yield and 28% composition), under the optimum reaction conditions. The following amidation on PAA-based NHS-esters with a model amine-containing compound, L-leucine methyl ester, generated approximately 70% amides and 30% carboxylates. In contrast, amidation of PAA- or PMAA-based anhydrides with L-leucine methyl ester only produced less than 30% amides but more than 70% carboxylates. The above reaction yields and percentage compositions were estimated by fitting the carbonyl stretching region with 5 possible species, NHS-ester, anhydride, N-acylurea, unreacted acid, unhydrolyzed tert-butyl ester, and using the Beer-Lambert law. The different surface chemistry mechanisms will bring significant effects on the performance of surface chemistry-derived devices such as biochips, biosensors, and biomaterials.
近红外光谱被应用于研究在多孔硅上接枝的聚丙烯酸（PAA）和聚甲基丙烯酸（PMAA）刷的众所周知的 EDC/NHS（N-乙基-N'-(3-(二甲基氨基)丙基)碳二亚胺/N-羟基琥珀酰亚胺）的活化细节。琥珀酰亚胺酯（NHS-酯）通常被认为是主要的中间产物，方便地通过酰胺键固定含有游离伯氨基的生物分子。令我们惊讶的是，红外光谱细节表明，在有文献记载的反应条件下，EDC/NHS 对 PMAA 的活化产生了酐（估计产率约为 76%，组成约为 70%），而不是 NHS-酯（产率约为 5%，组成约为 11%），作为主要的中间产物。相比之下，在最佳反应条件下，EDC/NHS 对 PAA 的活化仍然遵循一般规律，即预期的 NHS-酯是主要的中间产物（产率约为 45%，组成约为 57%），酐是副产物（产率约为 40%，组成约为 28%）。随后，用含模型胺的化合物 L-亮氨酸甲酯对 PAA 基 NHS-酯进行酰胺化，生成了约 70%的酰胺和 30%的羧酸盐。相比之下，用 L-亮氨酸甲酯对 PAA 或 PMAA 基酐进行酰胺化，只生成了不到 30%的酰胺，但生成了超过 70%的羧酸盐。上述反应产率和百分比组成是通过用 5 种可能的物质（NHS-酯、酐、N-酰基脲、未反应的酸、未水解的叔丁酯）拟合羰基伸缩区域，并使用比尔-朗伯定律来估计的。不同的表面化学机制将对基于表面化学的器件（如生物芯片、生物传感器和生物材料）的性能产生重大影响。
