【J. Org. Chem.】最新報道，氨基酸不用保護直接縮合新方法，收率高達95%

傳統多肽合成方法已沿用數十年，但需多步保護-脫保護操作、化學計量量的縮合試劑與添加劑，且消耗大量溶劑，存在原子經濟性低、廢棄物產生量大等缺陷。利用非保護氨基酸直接構建肽鍵是解決上述問題的優選方案，但該策略面臨非保護氨基酸在有機溶劑中溶解度低、副反應與消旋化等難題。【】

2026年， Alex Boateng等人 開發了一種以廉價市售三甲氧基硅烷為介導的肽鍵直接合成方法。三甲氧基硅烷可增溶有機溶劑中的非保護氨基酸、瞬時保護氨基，并同步活化羧基用于縮合。該方法為一鍋法，在 70℃下反應 20 小時，可由非保護氨基酸與氨基酸叔丁酯制備N端游離二肽，收率中等至優異（最高 95%）；反應立體選擇性極高，非對映選擇性 dr>20:1，底物適用范圍廣泛，且可用于含生物活性序列的三肽合成。【J. Org. Chem., 2026, 91, 2530-2537； doi.org/10.1021/acs.joc.5c02942 】

實驗條件優化

以H-L-Phe-OH與H-L-Ala-Ot-Bu為模板，對反應條件進行篩選。

三甲氧基硅烷用量：2.0 當量收率 62%，3.0 當量收率提升至 68%，4.0 當量無進一步提升，確定3.0 當量為最優用量。

反應溫度：50℃收率降至 54%，20℃無產物，80℃收率 68%，100℃收率 56%，確定70℃為最優溫度。

催化劑驗證：加入 CuCl、CuCl?、B (C?F?)?、Ta (OEt)?、Ta (OMe)?、Zn (OTf)?等金屬 / 路易斯酸催化劑，產物收率下降或無產物，證明該體系無需催化劑

溶劑篩選：單一溶劑 CPME、1,4 - 二氧六環、THF、CHCl?收率均低于混合溶劑；CPME/1,4 - 二氧六環（1:1）分離收率 82%，CPME/MeCN（1:1）分離收率 75%，為最優混合溶劑。

最優條件確定：非保護氨基酸（1.0 當量）、氨基酸叔丁酯（2.0 當量）、三甲氧基硅烷（3.0 當量），CPME/1,4 - 二氧六環（1:1）為溶劑，70℃反應 20 小時。

底物適用范圍1. 非保護氨基酸（親電試劑）

以H-L-Ala-Ot-Bu 為親核試劑，烷基側鏈（丙氨酸、纈氨酸、亮氨酸等）、含硫醚（芐基半胱氨酸、甲硫氨酸）、醚（叔丁基絲氨酸）、酯（叔丁基谷氨酸）、酰胺、吲哚、芳環（取代苯丙氨酸、酪氨酸）、位阻型 / 易消旋苯甘氨酸等底物均適用；芐基半胱氨酸丙氨酸二肽克級合成（4.0 mmol）分離收率 92%，所有產物 dr>20:1。

2. 氨基酸叔丁酯（親核試劑）

以H-L-Phe-OH、H-L-Cys (Bzl)-OH為親電試劑，烷基、硫醚、醚、酯、雜環、胍基、胺基側鏈，以及非天然氨基酸酯、β- 氨基酸酯、易二聚 / 易消旋酯均兼容，產物收率45%–95%，立體選擇性保持 dr>20:1。

3. 三肽合成

采用兩步一鍋法：先按最優條件合成二肽，室溫下直接加入Fmoc 保護氨基酸酰氯，無需添加劑繼續反應 20 小時，成功合成含生物活性序列的三肽（如 Ala-Phe-Ala、鐵氧還蛋白序列、黃嘌呤氧化酶抑制劑序列、亮氨酸腦啡肽序列），收率良好且 dr>20:1。

反應機理

三甲氧基硅烷在加熱條件下雙重作用：

1、與非保護氨基酸的氨基形成瞬時保護結構，避免氨基副反應；

2、活化羧基，與氨基酸形成穩定五元環中間體，實現選擇性縮合；

3、無需額外催化劑，加熱即可完成活化與脫保護，全程一鍋完成。

實驗操作

General Procedure: Trimethoxysilane-Mediated Peptide Bond Formation

Unprotected amino acid 1 (0.5 mmol, 1 equiv), dry CPME (0.75mL), dry 1,4-dioxane (0.75 mL), and trimethoxysilane (0.191 mL, 1.5mmol, 3 equiv) were charged into a flame-dried 5 mL screw-cap vial equipped with a magnetic stirring bar inside a glovebox. Subsequently,amino acid tert-butyl ester 2 (1.0 mmol, 2 equiv) was added to the mixture via a syringe under glovebox conditions. The reaction vial was then sealed under argon and removed from the glovebox. The mixture was stirred at 70 °C in a preheated oil bath for 20 h. After completion, the reaction was diluted with chloroform (1.5 mL), and approximately 0.3 g of silica gel (SiO2) was added. The mixture was stirred vigorously at room temperature (rt) for 5 min, then filtered through a G4 sintered funnel containing a Celite pad. The Celite pad was washed thoroughly with chloroform (150 mL). Volatiles were removed under reduced pressure, and the crude reaction mixture was transferred onto a silica gel column using a pipet. Purification was carried out using methanol in chloroform (0.2?1% MeOH in CHCl3) by flash column chromatography to afford the desired product 3.

General Procedure: Tripeptide Synthesis

Unprotected amino acid 1 (0.5 mmol, 1 equiv), dry CPME (0.75mL), dry 1,4-dioxane (0.75 mL), and trimethoxysilane (0.191 mL, 1.5mmol, 3 equiv) were charged into a flame-dried 5 mL screw-cap vial equipped with a magnetic stirring bar inside a glovebox. Subsequently, amino acid tert-butyl ester 2 (1.0 mmol, 2 equiv) was added to the mixture via a syringe under glovebox conditions. The reaction vial was then sealed under argon and removed from the glovebox. The mixture was stirred at 70 °C in a preheated oil bath for 20 h. The reaction vial was removed from the oil bath and allowed to cool to ambient temperature. Fmoc-protected amino acid halide (Fmoc-AA3-Cl 4, 0.75 mmol, 1.5 equiv) (prepared from its respective Fmoc-AA3-OH according to the literature12) was then added, and the reaction was stirred at rt for another 20 h. After completion, the reaction was diluted with chloroform (1.5 mL), and approximately 0.3 g of silica gel (SiO2) was added. The mixture was stirred vigorously at room temperature for 5 min, then filtered through a G4 sintered funnel containing a Celite pad. The Celite pad was washed thoroughly with chloroform (150 mL). Volatiles were removed under reduced pressure, and the crude reaction mixture was transferred onto a silica gel column using a pipet. Purification was carried out using AcOEt in hexane (20?80% AcOEt in hexane) by flash column chromatography to afford the desired product 5.

本文建立了一種無需金屬催化劑、由非保護氨基酸與氨基酸叔丁酯合成N 端游離多肽的方法，核心試劑為廉價、市售易得的三甲氧基硅烷。該策略適用于含烷基、硫醚、醚、酚、酯、胺、酰胺、胍基、雜環等多樣側鏈的氨基酸，包括 β- 氨基酸、易消旋、易自二聚氨基酸，對應二肽收率45%–95%；通過兩步一鍋法可高效合成含生物活性基序的三肽，產物光學純度優異（dr>20:1）。

參考資料：Trimethoxysilane-Mediated Peptide Bond Formation from Unprotected Amino Acids and Amino Acid t?Butyl Esters； Alex Boateng,* Isai Ramakrishna, Tomohiro Hattori,* and Hisashi Yamamoto*；J. Org. Chem., 2026, 91, 2530-2537； doi.org/10.1021/acs.joc.5c02942