Method for separation purification of polymyxin B1 from polymyxin B mixed component

Abstract

The present invention provides a method for separation purification of polymyxin B1 from a polymyxin B mixed component. The method comprises: (a) loading a chromatography medium of a macroporous absorption resin to obtain a chromatography column; and (b) adding a polymyxin B solution, and adopting an organic solvent-containing acid solution as an eluent to sequentially elute various components of the polymyxin B so as to obtain the polymyxin B1 solution after elution. According to the method, the operation is simple, the adopted separation medium has characteristics of stable chemical characteristic and high re-use rate, industrial scale-up is easily achieved, and the peak purity of the obtained polymyxin B1 can be more than 98%.

Description

technical field [0001] The invention belongs to the technical field of medicine, and in particular relates to a method for separating and purifying single-component polymyxin B1 from mixed components of polymyxin B. Background technique [0002] Polymyxin B (polymyxin B, whose general structural formula is shown in the following formula I) is a basic cyclic polypeptide antibiotic composed of various amino acids and fatty acids produced by Bacillus polymyxa. The medicinal form of polymyxin B is polymyxin B sulfate and its compound preparation with other drugs. It is mainly used as an anti-infective drug clinically to treat infections caused by Gram-negative bacteria such as Pseudomonas aeruginosa. Infections in skin wounds, urinary system, eyes, ears, trachea, etc. can also be used for sepsis, peritonitis, etc. Polymyxin B is a multi-component mixture with similar structure. Seven components have been found, namely B1, B1-I, B2, B3, B4, B5 and B6 (see formula 1 for the gener...

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Problems solved by technology

[0010] Polymyxin B1 is the most important component of polymyxin B. The literature reports on the separation and purification method of polymyxin B1 are as follows: W. Hausmann et al. components, but the separation effect is relatively poor (see W.Hausmann et al., Polymyxin B1.1Fractionation, Molecular Weight Determination, AminoAcid and Fatty Acid Composition.Journal of the American Chemicalsociety, 1954, 76(19):4892-4896); H. Kalasz et al. used displacement chromatography, using the analytical chromatographic column LiChrosorb RP-18 (5 μm, 250 × 4.6mm I.D.) as the stationary phase to separate B1 and B2, but only trace amounts of polymyxin B1 could be prepared (see H.Kalasz et al., Prepare-scale separation of polymyxins with an analytical high-performance liquid chromatography system by using displacement chromatography. Journal of Chromatography A, 1981, 251: 295-302); Y.Kimura et al. used Hitachi gel 3010 (granule 25 μm in diameter) and Amberlite XAD-2 (100-200 mesh) as filler, and linear gradient elution was used to effectively separate B1 and B2 (see Y.Kimura et al., Analytical and preparative methods for polymyxinantibiotics using high-performance liquid chromatography with a porous styrene-divinyllbenzene copolymer packing.Journal of ChromatographyA, 1981, 206: 563-572); I.Elverdam et al. used a reverse silica gel column LiChrosorb Si100ODS (10μm, 250×40mm I.D.), with isocratic elution method, for polyviscosity The single component of mycocin B was isolated (see I.Elverdam et al., Isolation and characterization of three newpolymyxins in polymyxins B and E by high-performance liquid chromatography.Journal of Chromatography A, 1981, 218: 653-661); He Fengyun in China also used reverse phase silica gel YMC C18-ODS-A (C18 bonded silica gel, 5μm, 150mm ×30mm) preparative column, with acetonitrile / water (21 / 79, V / V) containing 0.1% trifluoroacetic acid as mobile phase, to prepare polymyxin B1, but its single batch preparation amount is also low, the average per The sample load per milliliter of filler is 1.1 mg (see He Fengyun et al. Research on the preparation method of polymyxin B1 in polymyxin B. Chemical Times, 2011, 25(8): 20-22)

[0011] The separation and purification of polymyxin B1 by the above method has the following disadvantages: the separation effect is poor when using the countercurrent distribution method; High equipment requirements, poor stability of the separation medium, less repeated use times, and high cost

In short, the above methods are not easy to scale up

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