芐氧羰基(Cbz)保護氨基及脫保護方法

芐氧羰基（Cbz）是1932年Bergmann發現的一個很老的氨基保護基，但一直到今天還在應用。其優點在于：試劑的制備和保護基的導入都比較容易；N-芐氧羰基氨基酸和肽易于結晶而且比較穩定；芐氧羰基氨基酸在活化時不易消旋；能用多種溫和的方法選擇性地脫去。

芐氧羰基的導入，一般都是用Cbz-Cl。游離氨基在用NaOH 或NaHCO3 控制的堿性條件下可以很容易同Cbz-Cl反應得到N-芐氧羰基氨基化合物。α,β-二胺可用該試劑在pH= 3.5-4.5稍有選擇性地被保護，其選擇性隨碳鏈地增長而減弱，如H2N(CH2)nNH2, n=2時71%被單保護;n=7時29%被單保護[G.J. Atwell, W. A. Denny., Synthesis, 1984, 1032]。氨基酸酯同Cbz-Cl的反應則是在有機溶劑中進行，并用碳酸氫鹽或三乙胺來中和反應所產生的HCl。此外，Cbz-ONB（4-O2NC6H4OCOOBn）等芐氧羰基活化酯也可用來作為芐氧羰基的導入試劑，該試劑使伯胺比仲胺易被保護，但苯胺由于親核性不足，與該試劑不反應[D.R. Kelly, M. Gingell,Chem. Ind.(London),1991, 888]。

Cbz-Cl很容易用苯甲醇同光氣的反應來制備（見下式），在低溫下可以保存半年以上而不發生顯著的分解。

除Cbz-Leu為油狀物外，絕大多數氨基酸的芐氧羰基衍生物都可以得到結晶。有的N-芐氧羰基氨基酸能同它的鈉鹽按一定比例形成共晶，共晶產物的熔點較高，并難溶于有機溶劑。例如，苯丙氨酸經芐氧羰基化后再加酸析出Cbz-Phe時往往得到共晶產物（熔點144℃），此共晶產物用乙酸乙酯和1M HCl一道震搖時可完全轉化為Cbz-Phe而溶于乙酸乙酯中。因此。除Cbz-Gly以外，一般都是采用酸化后用有機溶劑提取的方法來得到純的N-芐氧羰基氨基酸。

反應實例

Cbz-Cl (18.5 μl,0.155 mmol) in diethyl ether (0.2 ml) was dropped to a solution of (R)-1(36.4 mg, 0.129 mmol) in 10% aqueous Na2CO3(1.8 ml) at 0°C,and stirred for 5 h. The reaction mixture was acidified with 10% citric acid,extracted with CHCl3(10mlX3). The organic layer was washed with water, driedover Na2SO4, evaporated to give light yellow gels, which werepurified by preparative TLC (CHCl3/MeOH=5:1) to afford (R)-6 (25.7 mg, 47.1%) as yellowamorphous solid.

【Konda-Yamada,Yaeko; Okada, Chiharu et al., Tetrahedrom; 2002, 58(39),7851-7865】

A 3-L, three-necked, Morton flask equipped with an efficient mechanical stirrer,thermometer, and a dropping funnel is charged with L-methionine methyl ester hydrochloride 1 ( 117.6 g , 0.56 mol), potassium bicarbonate ( 282.3 g , 2.82 mol, 5 eq.), water (750 mL), and ether(750 mL), and the solution is cooled to 0°C .Benzyl chloroformate ( 105 g , 88.6 mL, 0.62 mol, 1.1 eq.) is added dropwise over 1 hr, thecooling bath is removed, and the solution is stirred for 5 hr. Glycine ( 8.5 g , 0.11 mol, 0.2 eq.) is added (to scavenge excess chloroformate) and the solution is stirred for an additional 18 hr. The organic layer is separated, and the aqueous layer is extracted with ether (2 × 200 mL). The combined organic layers are washed with 0.01 M hydrochloric acid (2 × 500 mL), water (2 × 500 mL), and saturated brine (500 mL), and then dried (Na2SO4), filtered, and evaporated on arotary evaporator. The resulting oil is further dried in a Kugelrohr oven ( 50°C , 0.1 mm , 12 hr) to leave product 2 as a clear oil that solidifies upon cooling: 165– 166 g (98–99%), mp 42– 43°C .

【M. Carrasco, R. J. Jones, S. Kamel et a1., Org.Syn., 70, 29】

To a mixture oftoluene (3.85 L),water (3.85 L), andK2CO3(470 g, 3.40 mol) were successively added 1a (770 g, 2.72 mol) and CbzCl (488 g, 2.72 mol) with vigorous stirring at atemperature below 25 °C. After stirringat room temperature for 3 h, triethylamine (27.5 g, 270 mmol) and NaCl (578 g) were successively added, and the mixture wasstirred for a further 30 min. The organic layer was separated and concentratedto give the desired product as oil, which was used for the next reactionwithout purification. The analytical sample was prepared by columnchromatography。

【Inaba, Takashi; Yamada, Yasuki et al J. Org. Chem.,2000, 65(6), 1623-1628】

N-芐氧羰基（Cbz）的去保護

芐氧羰基的脫除主要有以下幾種方法：1). 催化氫解；2). 酸解裂解；3). Na/NH3（液）還原。一般而言目前實驗室常用簡潔的方法就是催化氫解，但當分子中存在對催化氫解敏感或鈍化的基團時，我們就必須采用化學方法如酸解裂解或Na/NH3（液）還原等。

催化氫解如下式所示。催化氫解的供氫體可以是H2、環己二烯、1，4-環己二烯、甲酸銨和甲酸等，以后面四個試劑為供氫體的反應又叫催化轉氫反應，通常這比催化氫化反應更迅速。

催化劑主要用5-10%的鈀-碳、10-20%的氫氧化鈀-碳或鈀-聚乙烯亞胺，鈀-聚乙烯亞胺/甲酸對于除去Cbz要比前兩者要好。當HBr/HOAc脫去Cbz保護基時，產物往往帶又一點顏色，而且分解產生的溴化芐會產生一些副反應并難以除盡，而催化氫解多數能得到無色得產物。由于硫能使催化劑中毒，因此，含有胱氨酸、半胱氨酸等含硫的肽等N-芐氧羰基氨基衍生物一般不用催化氫解法脫除。一般溶劑可以用甲醇，乙醇，乙酸乙酯，四氫呋喃等，在醇類質子溶劑中反應速度要快的多。

如果在Boc2O存在下用Pd/C進行氫化，則釋放出的胺直接轉變成Boc衍生物[ M. Sakaitani, K. Hori, Y. Ohfune., TetrahedronLett., 1988, 29, 2983]。而且這類反應往往要比不加Boc2O來的快，其主要由于氫解出來的胺往往會與貴金屬有一定的絡合，使催化劑的活性降低，和Boc2O反應為酰胺后則去除了這一效果。另外有時在氫解時加入適當的酸促進反應也是一樣的道理，避免了生成的胺降低反應的活性。

另外當分子中有鹵原子(Cl, Br, I)存在時，一般直接用Pd/C會造成脫鹵的發生，一般這種情況下，使用PdCl2為催化劑，以乙酸乙酯或二氯甲烷為溶劑可較好的避免脫鹵的發生。

用MeOH/DMF為溶劑時，在Cbz-賴氨酸衍生物氫化的過程中會生成N-甲基化的賴氨酸。使用氨為溶劑時，H2/Pd-C在-33℃下氫化，肽中的半胱氨酸或蛋氨酸單元不使催化劑毒化，此外，氨還會阻止BnO醚的還原，所以對Cbz可得到一些選擇性。

反應實例

A solution of (R)-8 (0.170 g, 0.52 mmol) in absolute methanol (3ml) was hydrogenated in the presence of 15% Pd/C (0.026 g)at room temperature for 12 h. The mixture was filtered (Celite) and washed with methanol. Then, perchloric acid (0.050ml, 0.83 mmol) was added and the mixture was stirred for 5 min. The solvent wasevaporated to afford (R)-7·HClO4, mp 233–235°C; [a]D23=?15.6 (c=0.68, methanol).

【C. Jaume; G. G.Santiago et al., Tetrahedron: Asymmetry, 2000, 11(22),4549-4458】

A solution of N-Cbz arylglycinol (17) (1.02 mmol) in MeOH (10 mL) wasstirred for 15 min in the presence of an excess of Pd(OH)2/C under a dihydrogenatmosphere. The solution was then filtered on a Celite pad and the solvent removedin vaccuo. Purification of the crude afforded the desired free 2-arylglycinols(S)-21 in 87% yield, white solid; [a]D20=+47.0 (c=0.78,CHCl3); mp 94-96°C(AcOEt)。

【B.Pierfrancesco; C. silvia et al., Tetrahedron, 1999, 55(10),3025】

576.6mg of compound 1 (1 mmol) was dissolved in 20 ml of methanol. Then 150 mg ofammonium formate (3 mmol) and 75 mg of 10% Pd-C was added and the reactionmixture was stirred at room temperature 10 min and then heated to reflux for 45min. The mixture was filtered through celite and the filtrate was evaporate todryness to give 430 mg of compound 2 (98%). This compound was used without further purification in the subsequentstep.

【Alargov, D. K; Naydenova, Z; Monatsh. Chem., 1997, 128(6-7),725-732】

Compound 1 (0.6 g, 0.8 mmol) was dissolved in 1:1 formicacid/methanol (60 mL) and added to a round-bottom flask (100 mL) containing 1equiv of palladium catalyst (10% Pd/C, 1.0 g, 0.9 mmol). The mixture was continuously stirred under refluxtemperature for 24 h. The catalyst was removed by filtration and washed with anadditional 10 mL of methanol. The combined solvents were removed by evaporationunder reduced pressure to give Compound 2 (0.34 g, 81%, a white solid, mp 96-98 °C). This compound was used withoutfurther purification in the subsequent step.

【Fyles, T. M.; Zeng, B.; J. Org. Chem., 1998, 63(23), 8337-8345】

10%Pd-C wasaddede to a solution of compound 1 (596 mg , 1.77 mmol) and (Boc)2O(773 mg, 3.54 mmol) in etnyl acetate (30 ml). The reation vessel was evacuatedand back-filled with nitrogen (three times), then back-filled with hydrogen (1atm). After 2 h, the mixture was filtered and concentrated. Purification bysilica gel chromatography (30% ethyl acetate/ hexanes - 50% ethyl acetate/hexanes) gave compound 2 (289 mg, 54%).

【WO2004092166】

To a solution ocompound 1 (900 mg) in methylene chloride (16.5 ml) was addede PdCl2 (30mg) and triethylamine (0.229 ml). Triethyl silane was added (2 x 0.395 ml) over2 h. The reaction mixture stirred 1 h and 2 ml of trifluoroacetic acid wasadded. After 30 min the reaction was basified with 2 N NaOH, extracted with methylene chloride, dried over MgSO4,filtered and concentrated. Chromatography was run on a biotage 40S column with3-5% MeOH/CH2Cl2 with 0.5% NH4OH to provide compound 2 as a oil (501mg, 74%).

【US20030144297】

酸解脫除

氨基甲酸芐酯在強酸性條件下容易去保護。HBr/HOAc 是酸解脫除芐氧羰基的最常用的試劑。脫除反應主要按下式進行。反應需要消耗2分子的HBr，Cbz的脫除速度隨HBr濃度的增大而增大，因此實際上都是采用高濃度的過量HBr/HOAc溶液（1.2M-3.3M）以保證反應的完全。

含有絲氨酸和蘇氨酸的肽或其它含羥基的氨基衍生物用HBr/HOAc脫除Cbz時會發生羥基的O-乙?；磻?。雖然O-乙?；苡脡A皂化或氨解脫去，但為了避免這個副反應，可以改用HBr/二氧六環或HBr/三氟乙酸來代替HBr/HOAc。由于HBr在三氟乙酸中的溶解度較小，因此不能預先制成HBr/三氟乙酸溶液，而只能將保護的肽或氨基衍生物溶于無水三氟乙酸中，先于0℃下通入干燥的HBr，待Cbz大部分脫除后，再室溫通短時間以求完全脫除變化基。Cbz被HBr分解產生的溴化芐能同肽中的某種氨基酸反應，也是需要加以注意的。如，甲硫氨酸的硫原子能同溴化芐反應生成S-芐基甲硫氨酸，防止的辦法是加入硫醚（CH3SC2H5）為捕捉劑。色氨酸被HBr/HOAc分解產生有色物質，防止的辦法是加入亞磷酸二乙酯。硝基精氨酸會發生硝基的部分脫落，改用液體HBr于-67℃處理可以避免。

用液體HF在0℃處理10-30分鐘即可將Cbz完全脫去。FSO3H、CH3SO3H、CF3SO3H和C6H5SCH3-TFA也是較好的試劑。Me3SiI在氯仿、乙腈中能于幾分鐘內選擇性脫去Cbz和Boc保護基。對于BBr3/CH2Cl2而言，較大分子的肽的Cbz衍生物可在TFA中去除，因為肽在酸中的溶解度比在CH2Cl2中大。從肽中脫去Cbz，可在TFA中添加0.5 M 4-（甲硫基）苯酚或使用HF/Me2S/對甲苯酚（25:65:10,v/v）來抑制Bn+對芳香氨基酸的加成。

此外，已經報道過的還有以下的一些不常用的方法。如HCl/CHCl3、HCl/HOAc、HBr/SO2、液體HBr、TosOH、HI/HOAc、碘化磷、Et3SiH、沸騰的TFA、8M HCl的乙醇液或6 M HCl回流1小時或濃鹽酸于25-75℃加熱處理1-1.5小時等。

A solution of the amine Cbz compund (208 mg, 0.44mmol) in 33 % hydrobromic acid in acetic acid (1 mL) and glacial acetic acid(0.6 mL) was stirred at rt for 3 h under an atmosphere of nitrogen. Thevolatiles were removed in vacuo to leave the free amine hydrobromide (168mg, 91 %) as a brown, highly hygroscopic powder; [α]D=-18.0° (c = 0.4, EtOH)

【B.Anna; P. Gerald., Heterocycles, 2002, 58, 521】

Me3SiI (0.73 ml,0.73 mmol) was added to a soluton of compound 1 (146 mg, 0.33 mmol) inacetonitrile (10 ml) at room temperature, and the resulting mixture was stirredat room temperature for 2 h. Et3N (0.12 ml) was added and themixture was stirred at room temperature for 15 min. The solvents were removedin vacuo, and the residue was extracted with ethyl acetate. The combinedorganics were washed with sodium bicarbonate and brine, dried over sodiumsulfate and filtered. Solvents were removed and the residue was used directlyin the next step.

【US20040204397】

2024年，Vinayagam等人報道了一種利用AlCl3/HFIP(1,1,1,3,3,3-六氟異丙醇）溫和脫Cbz的方法。該方法具有良好的官能團耐受性（包括其他可還原基團）、成本效益高、易于操作、安全性好、適合規?；a、高收率以及可在常溫下進行反應等優點。可以作為一種優秀的替代方案，避免了使用易燃的氫氣和金屬催化劑試劑，從而減少了嚴重的安全和環境問題。該方法最顯著的特點是在存在O-和N-芐基保護基的情況下，能夠選擇性脫保護N-Cbz基團，從而擴展了設計合成路線的范圍，適用于需要多種官能團轉化的目標化合物。

【 J. Org. Chem., 2024, 89, 5665. DOI: 10.1021/acs.joc.4c00177 】

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