An automatic detection method for a preparative chromatograph

Abstract

An automatic detection method for a preparative chromatograph, comprising the following steps: preparing a standard chromatogram; injecting a sample that matches the standard chromatogram, and the chromatograph outputs a chromatogram to be detected; setting the chromatographic peak list of the chromatogram to be detected to be displayed in descending order of peak area; selecting the first n chromatographic peaks of the chromatogram to be detected and the first n chromatographic peaks of the standard chromatogram, comparing whether the retention time of the first chromatographic peak of the chromatogram to be detected is close to the retention time of the first chromatographic peak of the standard chromatogram, and judging whether the retention time of this chromatographic peak of the chromatogram to be detected is within the collection range, obtaining a corresponding score, and the collection range is set in advance according to experience; continuing to complete the comparison of the other n-1 chromatographic peaks according to the same scoring rule; adding up the scores, and if the percentage obtained by dividing by the total score is within the set threshold, it is considered that the two chromatograms are similar and the separation can continue, otherwise it is prompted to stop the separation.

Description

Technical Field

[0001] The present invention belongs to the technical field of chromatographic analysis, and particularly relates to an automatic detection method for preparative chromatography. Background Art

[0002] For preparative separation chromatography, when the preparative chromatograph is working, the operator needs to keep an eye on the chromatograph, consuming a lot of energy. And the preparative chromatograph does not stop operating at night, but the operator cannot work without rest at night. At this time, how to ensure the stable operation of the preparative chromatograph, such as how to automatically check the separation effect and stop the machine in case of abnormality, just like the effect of an instrument operator checking at the side of the instrument at any time, which is a problem that has been urgently to be solved. Because for preparative separation chromatography, due to the large sample injection volume, once an abnormality occurs and the machine does not stop, the sample will be wasted, and this result is unacceptable. Especially for some precious samples, it is impossible to prepare them again in a short time after being wasted.

[0003] The operator judges whether the sample is separated normally by comparing chromatograms. The comparison of chromatograms is different from the comparison of two pictures. The key in the chromatogram is the subtle change of the curve. If the blank areas outside the curve are compared like two pictures, it may be concluded that the two pictures are very similar.

[0004] The comparison of chromatograms is also different from the comparison of spectrograms. For example, for two infrared spectrograms, as long as large peaks appear within the wavenumber range, and there are minor differences in specific details, the infrared spectrograms can be considered similar. This is because the influencing factors of spectrograms such as infrared spectrograms are relatively complex, and the peaks in the infrared spectrogram are generated by the stretching vibration of chemical bonds, etc. However, the core of the chromatogram is the chromatographic peak, and each chromatographic peak corresponds to a substance. Therefore, the comparison of chromatographic peaks is the key to the comparison of chromatograms.

[0005] In addition, there is no requirement for timeliness in spectral analysis comparison. The comparison of spectrograms is static, and the data can be collected and then compared one by one. While for preparative separation chromatography, it is necessary to make an immediate comparison, that is, to compare with the standard chromatogram after each injection, and the requirement for timeliness is very high.

[0006] An existing method for automatic recognition and analysis of chromatograms, such as the invention application with the application number 202111627579.0, records an automatic analysis method for chromatograms based on image recognition, including the following steps:

[0007] S1 For the spectral samples of target compounds in the standard spectral library, according to the properties of the target compounds, the samples are classified and labeled to obtain the classification sample set of the target compounds;

[0008] S2 Establishment of the YOLOv5 model:

[0009] S3 Recognition and analysis of the chromatographic image to be detected:

[0010] S31 performs multi-scale feature extraction on the chromatogram image to be detected to obtain the features of the chromatogram image to be detected;

[0011] S32 simultaneously inputs the features of the chromatogram image to be detected into one or more trained object detection models, and obtains the object identification frame, confidence level, and category information output by the detection model for the chromatogram image to be detected; according to the image area defined by the object identification frame, collect its recognition category, matching degree, and object identification frame coordinate information;

[0012] S33 compares the recognition matching degree, retention time deviation, and concentration of the chromatographic peaks in the image to be detected recognized by the YOLOv5 model with the preset conditions, and determines the chromatographic peaks that meet any of the following conditions as negative:

[0013] The recognition matching degree of the chromatographic peak is lower than the preset value;

[0014] The retention time deviation between the chromatographic peak and the standard substance chromatographic peak is greater than the preset value;

[0015] The quantitative concentration of the chromatographic peak is lower than the preset value;

[0016] S34 deletes the negative chromatographic peaks;

[0017] S35 performs quantitative analysis on the recognized chromatographic peaks:

[0018] Calculate the concentration value of the chromatographic peak; through the recognition result of the object detection model, calculate the retention time, matching degree, and identification frame coordinates of the recognized chromatographic peak;

[0019] S4 finally outputs the analysis results of the compound.

[0020] However, this automatic chromatogram analysis method is used for analyzing chromatograms. Since the result analysis of analyzing chromatograms first collects the data and then compares them one by one, it is not real-time. Even if the analysis instrument malfunctions during operation at night, the instrument can be repaired and restarted the next day. The purpose of preparative separation chromatography is not to obtain a good chromatogram, but to obtain a sufficient amount of target components, which is different from the purpose of analytical chromatography to obtain sample purity information. Therefore, the detection work of preparative separation chromatography has high requirements for timeliness. Summary of the Invention

[0021] The purpose of the present invention is to provide an automatic detection method for preparative chromatography to solve the above technical problems.

[0022] An automatic detection method for preparative chromatography includes the following steps:

[0023] Prepare a standard chromatogram;

[0024] Inject a sample that matches the standard spectrum, and the chromatograph outputs the spectrum to be detected;

[0025] Set the chromatographic peak list of the spectrum to be detected to be displayed in descending order according to the percentage peak area;

[0026] Select the first n chromatographic peaks of the spectrum to be detected and the first n chromatographic peaks of the standard spectrum, compare whether the retention time of the first chromatographic peak of the spectrum to be detected is close to the retention time of the first chromatographic peak of the standard spectrum, and determine whether the retention time of this chromatographic peak of the spectrum to be detected is within the collection range, and obtain the corresponding score. The collection range is set in advance according to experience;

[0027] Continue to complete the comparison of the other n - 1 chromatographic peaks according to the same scoring rules;

[0028] Accumulate the scores and divide by the full score. If the obtained percentage is within the set threshold, it is considered that the two chromatograms are similar and the separation can continue; otherwise, prompt to stop the separation.

[0029] Preferably, if the retention time of the i-th (i = 1, 2,..., n) chromatographic peak of the spectrum to be detected is within + / - m% of the retention time of the i-th chromatographic peak of the standard spectrum, and m <= 5, it is recorded as 1 point; otherwise, it is recorded as 0 point.

[0030] Preferably, if the retention time of the i-th chromatographic peak of the spectrum to be detected is within the collection range, it meets the requirements and is recorded as 10 points; otherwise, it is recorded as 0 point.

[0031] Preferably, also compare whether the half-peak width of the i-th chromatographic peak of the spectrum to be detected is within + / - p% of the half-peak width of the i-th chromatographic peak of the standard spectrum, and p <= 5, then it is recorded as 1 point; otherwise, it is recorded as 0 point.

[0032] The beneficial effects of the present invention are:

[0033] The present invention enables the chromatograph to automatically compare the spectrum of a just-completed sample with a manually specified standard spectrum. If an abnormality (deviation exceeds the limit) occurs, it will stop immediately, preventing waste of samples and mobile phases, saving resources, and reducing the manual workload.

[0034] The present invention compares the chromatographic peaks in reverse order of the maximum peak area, avoiding the problem of inaccurate detection caused by the mismatch of chromatographic peak numbers due to minor perturbations. Description of the Drawings

[0035] The drawings are used to provide a further understanding of the present invention and constitute a part of the specification. Together with the embodiments of the present invention, they are used to explain the present invention and do not constitute a limitation to the present invention. In the drawings:

[0036] Figure 1 is the standard spectrum of the comparative example;

[0037] Figure 2 is Figure 1 a partially enlarged view of;

[0038] Figure 3 is the spectrum to be detected of the comparative example;

[0039] Figure 4 is Figure 3 a partially enlarged view of;

[0040] Figure 5 is the spectrum to be detected of the embodiment of the present invention;

[0041] Figure 6 is Figure 5 a partially enlarged view of. Specific embodiments

[0042] Comparative example

[0043] As Figures 1 to 4 shown, please observe the peaks shown in gray within the collection range of these two chromatograms. Figure 2 In Figure 4 this peak is peak No. 8, and in

[0044] Example 1

[0045] As Figures 5 to 6 shown, an automatic detection method for preparative chromatography includes the following steps:

[0046] S1. Prepare a standard spectrum.

[0047] S2. Inject a sample that matches the standard spectrum, and the chromatograph outputs the spectrum to be detected.

[0048] S3. Set the chromatographic peak list of the spectrum to be detected to be sorted and displayed in real time from largest to smallest in terms of percentage peak area. This algorithm is an existing technology. For example, methods such as the bubble method are used for automatic sorting.

[0049] S4. Select the first n chromatographic peaks of the spectrum to be detected and the first n chromatographic peaks of the standard spectrum. The number of chromatographic peaks selected can be set by the user or default selected by the program. For example, select the first 5 peaks with the largest percentage peak area, or the first 10 peaks, or 50% of all peaks, etc. Please refer to Figure 6, in this embodiment, the first 5 chromatographic peaks are selected for comparison, and the peak areas of the chromatographic peaks are sorted from large to small. The meanings of the marks in the figure are as follows: for example, for the peak ranked 2nd by peak area: 24 - 11.130, where 24 is the original serial number of the chromatographic peak and 11.132 min is the retention time of the chromatographic peak; the two curves, one high and one low, in the figure represent the absorption values at two wavelengths of 220 nm and 254 nm.

[0050] S5. Determine the evaluation criteria according to the actual situation: for example, compare whether the retention time of the first chromatographic peak in the spectrum to be detected is close to the retention time of the first chromatographic peak in the standard spectrum, and judge whether the retention time of this chromatographic peak in the spectrum to be detected is within the collection range, and obtain the corresponding score. The collection range is set in advance according to experience. For example, in Figure 2 , Figure 4 , the first chromatographic peak is collected at 9 - 13 min. Considering the settings of the collection conditions for the target chromatographic peak by the user, for example, the user sets to collect the chromatographic peak at 11.718 min within the range of 9 - 13 min. Even if the change in the retention time of this chromatographic peak does not exceed 5%, but it exceeds the collection time range, the separation should be stopped.

[0051] S51. If the retention time of the i-th (i = 1, 2, ……, n) chromatographic peak in the spectrum to be detected is within + / - m% of the retention time of the i-th chromatographic peak in the standard spectrum, for example but not limited to m <= 5, then it is recorded as 1 point, otherwise it is recorded as 0 point.

[0052] S52. If the retention time of the i-th chromatographic peak in the spectrum to be detected is within the collection range, it meets the requirements and is recorded as 10 points (this is only an example here, and the full score of the scoring is not limited to 10 points), otherwise it is recorded as 0 point.

[0053] S6. Compare whether the half-peak width of the i-th chromatographic peak in the spectrum to be detected is within + / - p% of the half-peak width of the i-th chromatographic peak in the standard spectrum. For example but not limited to p <= 5, then it is recorded as 1 point, otherwise it is recorded as 0 point. Comparing the half-peak width is conducive to judging whether the peak shape of the chromatographic peak meets the requirements.

[0054] S7. Continue to complete the comparison of the other n - 1 chromatographic peaks according to the same scoring rules.

[0055] S8. Accumulate the scores, divide by the full score. If the obtained percentage is within the set threshold, for example, the obtained percentage is 95% and the set threshold is 90%, then it is considered that the two chromatograms are similar and the separation can continue, otherwise, prompt to stop the separation.

[0056] In short, compare the results after integrating the two chromatograms, and compare the chromatographic peak description parameters such as the retention time, collection range, and peak width of the main chromatographic peaks. If there are large variations, it should be considered that the curves of the two chromatograms are quite different and the separation should be stopped.

[0057] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or perform equivalent replacements for some of the technical features. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. An automatic detection method for a preparative chromatograph, characterized in that, Including the following steps: Prepare a standard chromatogram; Inject a sample that matches the standard chromatogram, and the chromatograph outputs the chromatogram to be detected; Set the chromatographic peak list of the chromatogram to be detected to be displayed in descending order according to the percentage peak area; Select the first n chromatographic peaks of the chromatogram to be detected and the first n chromatographic peaks of the standard chromatogram, compare whether the retention time of the first chromatographic peak of the chromatogram to be detected is close to the retention time of the first chromatographic peak of the standard chromatogram, and judge whether the retention time of this chromatographic peak of the chromatogram to be detected is within the collection range, and obtain the corresponding score; Continue to complete the comparison of the other n - 1 chromatographic peaks according to the same scoring rules; Accumulate the scores and divide by the full score. If the obtained percentage is within the set threshold, it is considered that the two chromatograms are similar and the separation can continue, otherwise, prompt to stop the separation.

2. The automatic detection method of the preparative chromatography according to claim 1, characterized in that, If the retention time of the i-th (i = 1, 2,..., n) chromatographic peak of the chromatogram to be detected is within + / -m% of the retention time of the i-th chromatographic peak of the standard chromatogram, where m <= 5, it is recorded as 1 point, otherwise it is recorded as 0 point.

3. The automatic detection method of the preparative chromatography according to claim 2, characterized in that, If the retention time of the i-th chromatographic peak of the chromatogram to be detected is within the collection range, it meets the requirements and is recorded as 10 points, otherwise it is recorded as 0 point.

4. The automatic detection method of the preparative chromatography according to claim 3, wherein Also compare whether the half-peak width of the i-th chromatographic peak of the chromatogram to be detected is within + / -p% of the half-peak width of the i-th chromatographic peak of the standard chromatogram, where p <= 5, it is recorded as 1 point, otherwise it is recorded as 0 point.

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