Method for preparing carbetocin through solid phase and liquid combining method

Abstract

The invention belongs to the field of polypeptide synthesis, and particularly relates to a method for preparing carbetocin through a solid phase and liquid combining method. A solid-phase synthesis method is firstly adopted for preparing an X-(CH2)3-CO-Tyr(Me)-Ile-Gln-Asn-Cys-OH fragment which is subjected to a coupled reaction with an H-Pro-Leu-Gly-NH2 fragment prepared through a liquid-phase method, carbetocin linear peptide is obtained, and then a cyclization reaction of linear peptide is carried out in the liquid phase to prepare carbetocin. According to the method, the operation process is easy and convenient, production cost is low, the situation that glycine is in lack of peptide impurities can be avoided, the subsequent purifying process is facilitated, and the method is suitable for large-scale production.

Description

technical field [0001] The invention relates to the field of polypeptide synthesis, in particular to a method for preparing carbetocin by a solid-liquid phase combination method. Background technique [0002] Carbetocin, whose English name is Carbetocin, is a long-acting oxytocin nonapeptide analog with agonist properties developed and synthesized by Ferring Inc. of Canada. It was launched in China in 2009. Its structure is as follows: [0003] [0004] Carbetocin is used for the prevention of uterine atony and postpartum hemorrhage after elective epidural or spinal anesthesia after caesarean section. Like oxytocin, carbetocin binds to the oxytocin receptor of uterine smooth muscle, causing rhythmic contraction of the uterus, increasing its frequency and increasing uterine tension on the basis of the original contraction; in the non-pregnant state, the uterine Oxytocin receptor levels are low, increase during pregnancy, and peak at parturition; carbetocin therefore has ...

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Problems solved by technology

[0005] At present, the preparation method of carbetocin has been widely reported at home and abroad: Spanish patent ES2115543 discloses a preparation method of carbetocin: using Fmoc-Cys(Trt)-OH and chlorobutyric acid as raw materials to synthesize carbetocin Betocin linear peptide resin, after cleavage to get linear peptide, then dissolve it in the solution, through NaOH / LiOH / DIEA / DMAP / NaHCO 3 Carbetocin is obtained by cyclization with alkaline reagents; this method uses pure solid-phase synthesis of carbetocin linear peptides, the cost is high, and the purity of crude peptide samples is low

[0006] In addition, Czech Patent CS8605461 discloses a method for synthesizing Z-Ile-Gln-Asn-Cys(Bzl)-Pro-Leu-Gly-NH by solid-phase method 2 Linear peptide, hydrogenated to give Ile-Gln-Asn-Cys-Pro-Leu-Gly-NH 2 Then react with bromobutyric acid, and then cyclize to obtain carbetocin. Due to the existence of hydrogenation operation in this method, the safety risk is large, and it is not suitable for industrial production

[0007] In China, patents CN104262464A, CN102796178A, CN104086631A, CN101555272B and CN102260326A, CN103467573A respectively disclose a kind of solid-phase synthesis of carbetocin linear peptide resin, and then carry out cyclization reaction on the solid phase. Glycine-deficient peptide impurities are easily produced during the synthesis process, and amino resins are generally used, which has high production costs and is not suitable for large-scale production

[0008] In Chinese patent CN103833831A, a method for solid-phase synthesis of carbetocin by fragment method, the method adopted is to first synthesize fragment 1 peptide resin Br(CH2)3-CO-Tyr(Me)-Ile-Gln(Trt)- Cys-Wangresin and fragment 2 peptide resin H-Pro-Leu-Gly-Rinkamide-AMresin, and then fragment 1 peptide resin is cyclized and cleaved and then connected to fragment 2, and carbetocin can be obtained after cleavage again. This method needs to use two kinds of solid-phase supports, and after a two-step cracking process, the production cost is high, which is not conducive to scale-up production

[0009] Chinese patent CN102477082A discloses a method for synthesizing carbetocin in pure liquid phase: firstly synthesize three carbetocin intermediate fragments in the liquid phase, and then connect the three fragments to form carbetocin Carbetocin is prepared by cyclization reaction to prime linear peptide. This method involves a variety of intermediates. The steps are cumbersome and inconvenient to operate, which is not conducive to large-scale production.

[0010] To sum up, in the currently reported synthetic methods of carbetocin, pure solid-phase synthesis method is easy to produce glycine-deficient peptide impurities, sample purification is difficult, and it is necessary to use expensive amino resins as raw materials for the synthesis of peptide chains or fragments. , the production cost is high, and it is not suitable for large-scale production; and there are too many intermediates in the method for synthesizing carbetocin by the pure liquid phase method, the post-reaction treatment work is cumbersome, and the operation is not convenient, which is also not conducive to carrying out Zooming in on production issues

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