Background and overview[1]
In the process of peptide-to-peptide coupling activation, the condensation reagent determines the degree of isomerization and yield of the polypeptide, benzotriazole-N, N, N’, N’-tetramethylurea hexafluorophosphate Salt (HBTU) is a typical urea cationic polypeptide condensation reagent. In peptide synthesis, benzotriazole-N, N, N’, N’-tetramethylurea hexafluorophosphate is used as a coupling reagent to synthesize peptides. Compared with other coupling reagents, it has the advantage of synthetic peptide elimination. It has the advantages of low rotation amount, simple coupling conditions, short reaction time, and high yield. HBTU has the characteristics of simple and convenient synthesis process and long shelf life.
The traditional method for preparing HBTU is to use tetramethylurea and phosgene as raw materials to synthesize the intermediate N, N, N’, N’-tetramethyl chloroformamidine hexafluorophosphate (TCFH). The intermediate TCFH is mixed with 1 -The potassium salt of hydroxybenzotriazole is reacted to synthesize benzotriazole-N, N, N’, N’-tetramethylurea hexafluorophosphate (HBTU). This synthesis method is simple and low-cost, but phosgene It is a serious asphyxiating gas and can easily cause safety accidents. Therefore, it is extremely important to find an effective substitute for phosgene. In 1989, research showed that dangerous phosgene was replaced by oxalyl chloride, and it was proposed that tetrafluoroborate could be used as a non-nucleophilic counterion, and a one-pot method in anhydrous organic solvents was established. In 1998, in the process of synthesizing tetramethylfluoroformamidine hexafluorophosphate (TFFH), a process was developed to replace phosgene with phosphorus oxychloride and expensive ammonium hexafluorophosphate with HPF6. The advantages of this method are The disadvantage of using phosphorus oxychloride instead of dangerous phosgene is that the hexafluorophosphoric acid used is severely corrosive to equipment and toxic to the human body.
Preparation[1]
Benzotriazole-N, N, N’, N’-tetramethylurea hexafluorophosphate is prepared as follows: using phosphorus oxychloride and tetramethylurea as raw materials, and potassium hexafluorophosphate as ion exchange agent, synthesize the intermediate N, N, N’, N’-tetramethylchloroformamidine hexafluorophosphate (TCFH), react the synthesized TCFH with the potassium salt of 1-hydroxybenzotriazole to synthesize benzotriazole Azole-N, N, N’, N’-tetramethylurea hexafluorophosphate (HBTU). The results show that when potassium hexafluorophosphate is used as the ion converting agent, the volume ratio of dichloromethane to phosphorus oxychloride is 15.6, the dropping time is 90 minutes, and the dropping temperature is 25°C, the yield of the intermediate TCFH reaches more than 60%. , reaction time and rotation speed have a certain impact on HBTU, and the total yield of HBTU can reach 95%.
1) Synthesis of the intermediate N, N, N’, N’-tetramethyl chloroformamidine hexafluorophosphate (TCFH) (1): Synthesis of tetramethyl chloroformamidine dichlorophosphate: Add 0.088 mol of tetramethylurea into the reaction kettle. When the system reaches a certain temperature, add 0.098 mol of POCl3 dropwise. After the addition is completed, move the reaction kettle to the oil bath, raise the temperature to 100°C, react for 60 minutes, and cool to A clear yellow liquid was obtained at room temperature.
2) Synthesis of N, N, N’, N’-tetramethylchloroformamidine hexafluorophosphate: Add methylene chloride to the reaction solution obtained in step (1), and drop it under ice bath conditions. Add 0.088 mol of ion exchanger potassium hexafluorophosphate aqueous solution. After the dropwise addition, continue stirring under the ice bath, separate the liquids, wash the organic phase with deionized water, separate the liquids again, dry the organic phase with magnesium sulfate, and spin at 50°C. The organic phase was evaporated, crystallized and dried to obtain a white solid.
3) Synthesis of benzotriazole-N, N, N’, N’-tetramethylurea hexafluorophosphate (HBTU). Weigh 0.065 mol of the potassium salt of 1-hydroxybenzotriazole ( KOBt), dissolve in 30 mL of water, weigh 0.065 mol of TCFH and dissolve in 200 mL of methylene chloride. At room temperature, pour the KOBt solution into the TCFH solution and stir. After the reaction is completed, add anhydrous magnesium sulfate to the reaction solution, filter it with suction, filter out the anhydrous magnesium sulfate and N-HBTU polyurethane coating, and rotary evaporate the filtrate to obtain The solid was recrystallized from diethyl ether to obtain O-HBTU. Dissolve the solid mixture of anhydrous magnesium sulfate and N-HBTU in acetonitrile, wash it 2 to 3 times, filter out the insoluble matter, and rotary evaporate to obtain white solid N-HBTU. Recrystallize it with diethyl ether to obtain N-HBTU.
