Method for monitoring separation and purification process of etimicin hydrolysate

Abstract

The invention discloses a method for monitoring separation and purification processes of etimicin hydrolysate, which comprises the following steps of sampling the etimicin hydrolysate and eluent purified on a column, detecting by an HPLC-post-column derivatization method, separating a sample to be detected by an HPLC chromatographic column, performing derivatization reaction after separation, and detecting the obtained derivative in a fluorescence detector, wherein the excitation wavelength of the fluorescence detector is 310-350 nm, and the emission wavelength of the fluorescence detector is 440-480 nm. By adopting the method provided by the invention, the purity of the etimicin hydrolysate and the column purification eluent and the content of each impurity can be detected on line by using a conventional detector and a high performance liquid chromatograph, the separation and purification process of the etimicin hydrolysate is monitored, and the method is simple, convenient, feasible, accurate, reliable and good in repeatability.

Description

technical field [0001] The invention relates to a method for monitoring the separation and purification process of etimicin hydrolyzate, which performs on-line rapid detection of the column-purified eluate of the etimicin hydrolyzate, so as to realize the monitoring of the separation and purification process. Background technique [0002] For etimicin hydrolyzate and its column purification eluent, since etimicin and its impurity components have no ultraviolet absorption, the finished product detection generally adopts the HPLC-ELSD detection method, but the ELSD detection method concentration and peak area The linear relationship is poor, the purity data cannot be obtained intuitively, and the calculation must be substituted by the accompanying curve method within a narrow concentration range, which is time-consuming and laborious. The concentration of the column purification eluent varies from small to large, and the range of variation is large. The ELSD method is used. Ge...

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Problems solved by technology

[0002] For etimicin hydrolyzate and its column purification eluent, since etimicin and its impurity components have no ultraviolet absorption, the finished product detection generally adopts the HPLC-ELSD detection method, but the ELSD detection method concentration and peak area The linear relationship is poor, the purity data cannot be obtained intuitively, and the calculation must be substituted by the accompanying curve method within a narrow concentration range, which is time-consuming and laborious. The concentration of the column purification eluent varies from small to large, and the range of variation is large. The ELSD method is used. Generally, accurate results cannot be obtained, so this method is generally not used for the detection of etimicin hydrolyzate on column purification eluent

[0003] At present, the monitoring of the purification process generally adopts the pre-column derivatization method, which can directly calculate the sample purity, but the disadvantage of this method is that the derivatization operation process is cumbersome, and it is easy to affect the quantitative accuracy; when a complex component sample is derivatized After the reaction, a variety of derivatized products may be produced, which brings difficulties in chromatographic separation and impurity characterization; the reproducibility is not good, and it is not easy to be continuous

That is, there is a big gap between the pre-column derivatization method and the finished product, and it cannot correspond to the impurities in the finished product one by one, and the product quality cannot be well controlled.

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