Rapid balance calibration method and system for gradient elution liquid chromatograph

Abstract

The invention relates to a rapid balance calibration method and system of a gradient elution liquid chromatograph, and relates to the technical field of chromatographic instrument sample separation, and the rapid balance calibration method comprises the following steps: collecting an initial temperature and an initial dead volume of a preset chromatographic column and an initial flow velocity of a preset mobile phase; calculating a quotient between the initial dead volume and a preset target delay time to generate a target flow rate; calculating according to the initial temperature, the initial flow velocity, the target flow velocity and a preset mobile phase volume expansion coefficient to generate a target temperature; the mobile phase viscosity at the target temperature is obtained; correcting the target temperature according to the mobile phase viscosity to generate an actual target temperature; and controlling a preset chromatographic column adjusting device to adjust the temperature of the chromatographic column according to the actual target temperature so as to promote the rapid balance of the gradient elution liquid chromatograph. The method has the effect of improving the sample analysis efficiency.

Description

Technical Field

[0001] This application relates to the technical field of chromatograph sample separation, and in particular to a rapid equilibration calibration method and system for a gradient elution liquid chromatograph. Background Technology

[0002] Gradient elution liquid chromatography refers to an analytical device that uses the instrument's solvent mixing system to continuously change the composition ratio or concentration of the mobile phase according to a preset program, thereby separating complex mixtures more efficiently.

[0003] In related technologies, the equilibrium calibration method for gradient elution liquid chromatography usually adopts a static waiting approach, that is, after setting the gradient elution parameters, waiting for the mobile phase to mix fully in the chromatographic column and the column temperature to stabilize, until the signal detected by the detector tends to be stable, thereby completing the equilibrium calibration.

[0004] Regarding the aforementioned technologies, when using the above-mentioned equilibrium calibration method for equilibrium calibration, since the dynamic parameter fluctuations during the elution process are not adjusted in real time, when in a multi-component mobile phase system or when frequently changing samples, on the one hand, the waiting process for equilibrium calibration is prolonged, thereby extending the analysis time of the gradient elution liquid chromatograph for the sample; on the other hand, it leads to lower accuracy of the analytical results, thereby increasing the number of times the gradient elution liquid chromatograph analyzes the same sample, resulting in low sample analysis efficiency, and there is still room for improvement. Summary of the Invention

[0005] To improve sample analysis efficiency, this application provides a rapid equilibration calibration method and system for a gradient elution liquid chromatograph.

[0006] In a first aspect, this application provides a rapid equilibrium calibration method for a gradient elution liquid chromatograph, employing the following technical solution: A rapid equilibrium calibration method for a gradient elution liquid chromatograph includes: The initial temperature, initial dead volume, and initial flow rate of the preset chromatographic column and the preset mobile phase are collected. Calculate the quotient between the initial dead volume and the preset target delay time to generate the target flow rate; The target temperature is generated by calculating based on the initial temperature, initial flow rate, target flow rate, and preset mobile phase volume expansion coefficient. Obtain the viscosity of the flowing phase at the target temperature; The target temperature is corrected based on the viscosity of the mobile phase to generate the actual target temperature; The column temperature is adjusted by the preset column adjustment device according to the actual target temperature to promote rapid equilibration of the gradient elution liquid chromatograph.

[0007] By adopting the above technical solution, the target flow rate is obtained by calculating the quotient between the initial dead volume and the target delay time. The target temperature is calculated based on the initial temperature, initial flow rate, target flow rate, and mobile phase volume expansion coefficient. The target temperature is then corrected based on the mobile phase viscosity to obtain the actual target temperature. The column adjustment device is then used to adjust the column temperature according to the actual target temperature to promote the rapid equilibration of the gradient elution liquid chromatograph, thereby improving the analytical efficiency of the gradient elution liquid chromatograph for samples.

[0008] Optionally, the column adjustment device includes a column temperature control component and a column volume adjustment component. The step of adjusting the column temperature according to the actual target temperature using the preset column adjustment device to promote rapid equilibration of the gradient elution liquid chromatograph includes: Determine whether the actual target temperature is greater than the preset safe temperature of the chromatographic column; If it is not greater than the actual target temperature, the column temperature control component will adjust the column temperature to the actual target temperature. If the temperature is greater than the safe temperature, the column temperature control component will adjust the column temperature to the safe temperature and the column volume adjustment component will adjust the column volume to compensate for the flow rate of the mobile phase.

[0009] By adopting the above technical solution, it is determined whether the actual target temperature is greater than the preset safe temperature of the chromatographic column. If it is not greater, the temperature of the chromatographic column is adjusted to the actual target temperature by the chromatographic column temperature control component, thereby enabling the gradient elution HPLC to quickly achieve equilibrium calibration. If it is greater, the temperature of the chromatographic column is adjusted to the safe temperature of the chromatographic column by the chromatographic column temperature control component, and the volume of the chromatographic column is adjusted by the chromatographic column volume adjustment component, thereby compensating for the flow rate of the mobile phase, and thus enabling the gradient elution HPLC to quickly achieve equilibrium calibration, shortening the equilibrium calibration time.

[0010] Optionally, the steps of adjusting the column temperature to the safe column temperature using the column temperature control component and adjusting the column volume using the column volume adjustment component to compensate for the mobile phase flow rate include: Obtain the target dead volume and the change in dead volume of the chromatographic column; Calculate the ratio of the target dead volume to the initial dead volume to generate the dead volume ratio; Determine whether the change in dead volume meets the preset requirement for increasing dead volume; If satisfied, the column volume adjustment component is used to increase the dead volume based on the dead volume ratio. If the condition is not met, the column volume adjustment component is used to reduce the dead volume based on the dead volume ratio.

[0011] By employing the above technical solution, the ratio of the target dead volume to the initial dead volume is calculated to obtain the dead volume ratio. It is then determined whether the change in dead volume meets the requirement for increasing the dead volume. If it does, the column volume adjustment component is controlled to increase the dead volume based on the dead volume ratio. If it does not, the column volume adjustment component is controlled to decrease the dead volume based on the dead volume ratio. This controls the dead volume to offset inaccurate sample analysis results caused by the temperature not reaching the target temperature, thereby reducing the number of analyses performed on the same sample by the gradient elution liquid chromatograph and improving sample analysis efficiency.

[0012] Optionally, the steps for obtaining the target dead volume and the change in dead volume of the chromatographic column include: The actual flow rate is calculated based on the initial flow rate, the mobile phase volume expansion coefficient, the initial temperature, and the column safety temperature. Calculate the product between the actual flow velocity and the target delay time to generate the target dead volume; Calculate the difference between the initial dead volume and the target dead volume to generate the change in the initial dead volume; Calculate the difference between the column's safe temperature and the initial temperature to generate the column temperature change. Calculate the product between the preset thermal expansion coefficient of the stationary phase and the change in column temperature to generate the dead volume effect of the stationary phase; The difference between the stationary phase dead volume effect and the initial dead volume change is calculated to generate the dead volume change.

[0013] By adopting the above technical solution, the actual flow rate is calculated based on the initial flow rate, the mobile phase temperature-volume influence coefficient, the initial temperature, and the column safety temperature. The target dead volume is obtained by multiplying the actual flow rate by the target delay time. The change in initial dead volume is obtained by calculating the difference between the initial dead volume and the target dead volume. The change in column temperature is obtained by calculating the difference between the column safety temperature and the initial temperature. The influence of stationary phase dead volume is obtained by multiplying the stationary phase thermal expansion coefficient by the change in column temperature. The change in dead volume is obtained by calculating the difference between the influence of stationary phase dead volume and the change in initial dead volume. It is then determined whether the change in dead volume meets the requirement for increasing dead volume, and the column volume adjustment component is controlled to increase the dead volume based on the dead volume ratio.

[0014] Optionally, the step of increasing the dead volume by controlling the column volume adjustment component based on the dead volume ratio includes: The initial column length of the preset stationary phase is collected; Calculate the product between the initial column length and the dead volume ratio to generate the target column length; Calculate the difference between the target column length and the initial column length to generate the column length adjustment amount; The actual position of the column volume adjustment component was recorded. Query the x-coordinate value of the actual position of the column volume adjustment component to generate the actual position length of the adjustment component; Calculate the difference between the actual position length of the adjustment component and the column length adjustment amount to generate the target displacement of the adjustment component; The column volume adjustment component adjusts the column length of the stationary phase to increase the dead volume, based on the target displacement of the adjustment component.

[0015] By adopting the above technical solution, the target column length is obtained by calculating the product between the initial column length and the dead volume ratio. The column length adjustment amount is obtained by calculating the difference between the target column length and the initial column length. The actual position length of the adjustment component is obtained by querying the value of the horizontal coordinate of the actual position of the column volume adjustment component. The target displacement of the adjustment component is obtained by calculating the difference between the actual position length of the adjustment component and the column length adjustment amount. Thus, the column length of the stationary phase is adjusted by the column volume adjustment component according to the target displacement of the adjustment component to increase the dead volume, thereby compensating for the flow rate of the mobile phase.

[0016] Optionally, the step of reducing dead volume by controlling the column volume adjustment component based on the dead volume ratio includes: The initial inner diameter of the chromatographic column was collected; Take the square root of the dead volume ratio and divide it by the initial inner diameter to generate the target inner diameter; The column volume adjustment component, based on the target inner diameter, adjusts the inner diameter of the column to reduce dead volume.

[0017] By adopting the above technical solution, the target inner diameter is obtained by taking the square root of the dead volume ratio and dividing it by the initial inner diameter. Based on the target inner diameter, the column volume adjustment component is controlled to adjust the inner diameter of the column to reduce the dead volume, thereby compensating for the flow rate of the mobile phase.

[0018] Optionally, the step of correcting the target temperature based on the viscosity of the mobile phase to generate the actual target temperature includes: The mobile phase viscosity, preset column parameters, and target flow rate are substituted into Poiseuille's law to calculate the predicted system pressure corresponding to the target flow rate. Determine whether the predicted system pressure is greater than the preset safe system pressure; If the target flow rate is greater than the target flow rate, the target flow rate is reduced in a stepwise manner, and the target temperature is corrected based on the van der Hoff equation to generate the actual target temperature. If it is less than, then continue to obtain the predicted system pressure and perform loop judgment.

[0019] By adopting the above technical solution, the mobile phase viscosity, chromatographic column parameters and target flow rate are substituted into Poiseuille's law to calculate the predicted system pressure corresponding to the target flow rate. It is then determined whether the predicted system pressure is greater than the safe system pressure. If it is, the target flow rate is reduced stepwise, and the target temperature is corrected based on the van der Hoff equation to generate the actual target temperature. This improves the analytical accuracy of the gradient elution liquid chromatograph and thus improves the sample analysis efficiency.

[0020] Secondly, this application provides a rapid equilibration calibration system for a gradient elution liquid chromatograph, employing the following technical solution: A rapid equilibrium calibration method for a gradient elution liquid chromatograph includes: The data acquisition module is used to acquire initial temperature, initial dead volume, and initial flow rate. A memory for storing a program for a rapid equilibrium calibration method for a gradient elution liquid chromatograph as described in any of the preceding claims; The processor and the program in the memory are capable of being loaded and executed by the processor to implement a rapid equilibrium calibration method for a gradient elution liquid chromatograph as described in any of the preceding claims.

[0021] By adopting the above technical solution, the processor loads and executes a program of a rapid equilibrium calibration method for a gradient elution liquid chromatograph stored in the memory, and controls the acquisition module to acquire a series of data related to the realization of rapid equilibrium calibration of the gradient elution liquid chromatograph, thereby...

[0022] In summary, this application includes at least one of the following beneficial technical effects: 1. The target flow rate is obtained by calculating the quotient between the initial dead volume and the target delay time. The target temperature is calculated based on the initial temperature, initial flow rate, target flow rate, and mobile phase volume expansion coefficient. The target temperature is then corrected based on the mobile phase viscosity to obtain the actual target temperature. The column adjustment device is then used to adjust the column temperature based on the actual target temperature to promote rapid equilibration of the gradient elution liquid chromatograph, thereby improving the analytical efficiency of the gradient elution liquid chromatograph for samples. 2. By determining whether the actual target temperature is greater than the preset safe temperature of the chromatographic column, if it is not greater, the column temperature control component is controlled to adjust the column temperature to the actual target temperature, thereby enabling the gradient elution HPLC to quickly achieve equilibrium calibration. If it is greater, the column temperature control component is controlled to adjust the column temperature to the safe temperature, and the column volume adjustment component is controlled to adjust the column volume, thereby compensating for the flow rate of the mobile phase, and thus enabling the gradient elution HPLC to quickly achieve equilibrium calibration, shortening the equilibrium calibration time. 3. By calculating the ratio of the target dead volume to the initial dead volume, the dead volume ratio is obtained. It is then determined whether the change in dead volume meets the requirement for increasing the dead volume. If it does, the column volume adjustment component is controlled to increase the dead volume based on the dead volume ratio. If it does not meet the requirement, the column volume adjustment component is controlled to decrease the dead volume based on the dead volume ratio. This controls the dead volume to offset the inaccuracy of sample analysis results caused by the temperature not reaching the target temperature, thereby reducing the number of analyses of the same sample by the gradient elution liquid chromatograph and improving sample analysis efficiency. Attached Figure Description

[0023] Figure 1 This is a flowchart of a rapid equilibrium calibration method for a gradient elution liquid chromatograph according to an embodiment of this application.

[0024] Figure 2 This is a flowchart of the steps in this application embodiment to adjust the column temperature according to the actual target temperature using a preset column adjustment device to promote rapid equilibration of the gradient elution liquid chromatograph.

[0025] Figure 3 This is a flowchart illustrating the steps in this application whereby the column temperature control component adjusts the column temperature to the safe temperature of the column according to the safe temperature of the column, and the column volume adjustment component adjusts the volume of the column to compensate for the flow rate of the mobile phase.

[0026] Figure 4 This is a flowchart of the steps for obtaining the target dead volume and the change in dead volume of the chromatographic column in the embodiments of this application.

[0027] Figure 5 This is a flowchart of the steps in this application embodiment to control the column volume adjustment component to increase the dead volume according to the dead volume ratio.

[0028] Figure 6 This is a flowchart of the steps in this application embodiment to control the column volume adjustment component to reduce dead volume based on the dead volume ratio.

[0029] Figure 7 This is a flowchart of the steps in this application embodiment to correct the target temperature based on the viscosity of the mobile phase to generate the actual target temperature. Detailed Implementation

[0030] To make the purpose, technical solution, and advantages of this application clearer, the following description is provided in conjunction with the appendix. Figures 1 to 7 The present application will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are for illustrative purposes only and are not intended to limit the scope of the application.

[0031] This application discloses a rapid equilibration calibration method for a gradient elution liquid chromatograph. This method primarily addresses the rapid equilibration calibration problem of gradient elution liquid chromatographs. Specifically, it discloses a gradient elution liquid chromatograph, a chromatographic column, a column status monitoring device, a processing terminal, and a column adjustment device. The processing terminal is communicatively connected to both the column status monitoring device and the column adjustment device to achieve data exchange and control. After the column status monitoring device sends column status data to the processing terminal, the processing terminal compares the column status data with normal column status data. When the column status data and normal column status data are inconsistent, the processing terminal controls the column adjustment device to adjust the column in the gradient elution liquid chromatograph based on the type of difference between the column status data and the normal column status data. This aims to rapidly and rationally control the column adjustment device to adjust the column, thereby improving the analytical efficiency of the gradient elution liquid chromatograph.

[0032] Reference Figure 1 This application discloses a rapid equilibration calibration method for a gradient elution liquid chromatograph, comprising the following steps: Step S100: Collect the initial temperature, initial dead volume of the preset chromatographic column, and initial flow rate of the preset mobile phase.

[0033] Among them, the chromatographic column refers to the cylindrical container in the chromatograph that uses the adsorption-desorption equilibrium difference of the stationary phase to separate the sample components one by one when the mobile phase gradient changes.

[0034] A chromatograph is a gradient elution liquid chromatograph, which is an analytical device that uses the instrument's solvent mixing system to continuously change the composition ratio or concentration of the mobile phase according to a preset program, thereby separating complex mixtures more efficiently.

[0035] The initial temperature refers to the internal temperature of the chromatographic column before equilibration calibration. The initial temperature can be obtained by measuring the temperature sensor in the column condition monitoring equipment.

[0036] The initial dead volume is the blank volume in the column that is not filled with stationary phase before equilibration calibration. In one embodiment, the operator can obtain the initial dead volume by looking up the column model in the column technical manual.

[0037] The stationary phase is a substance packed inside a chromatographic column that interacts with different components in the sample to cause differences in the retention time of each component within the column, thereby achieving separation.

[0038] The mobile phase is a liquid substance used to carry the sample through the chromatographic column to promote the separation of sample components on the stationary phase.

[0039] The initial flow rate refers to the volume of mobile phase flowing into the column per unit time before equilibration calibration. In one embodiment, the operator can obtain the actual flow rate by consulting the column technical manual according to the column model. When the column length is 50 mm, the initial flow rate ranges from 1.2 to 1.5 mL / min, and when the column length is 250 mm, the initial flow rate ranges from 0.6 to 0.8 mL / min.

[0040] A chromatographic column status monitoring device is a device consisting of a temperature sensor, a detector, a laser displacement sensor, and a communication module, used to monitor the working status of a chromatographic column in real time and acquire chromatographic column status data.

[0041] Step S101: Calculate the quotient between the initial dead volume and the preset target delay time to generate the target flow rate.

[0042] The target flow rate refers to the volume of the mobile phase flowing through the column from its entry into the column to the detector in the column status monitoring device within a unit time during the target delay time. The target flow rate is obtained by dividing the initial dead volume by the target delay time through the processing terminal. The target flow rate is a parameter that balances the residence time of the mobile phase in the column and the separation efficiency of the chromatograph. When the target flow rate is too high, the residence time of the mobile phase in the column is short, which will lead to incomplete separation of substances by the chromatograph and a decrease in resolution. When the target flow rate is too low, the residence time of the mobile phase in the column is long, which improves the resolution, but will cause the peak shape of the separated substances to be too broad, resulting in low accuracy of the detection results.

[0043] The target delay time refers to the ideal time required for the mobile phase to travel from entering the chromatographic column to reaching the detector in the column status monitoring device. In one embodiment, the target delay time can be obtained by the operator by searching for sample delay time literature according to the sample type. The target flow rate of the mobile phase can be obtained through the target delay time, so that the analyte is completely separated and the accuracy of the detection result is within the normal accuracy range.

[0044] Step S102: Calculate the target temperature based on the initial temperature, initial flow rate, target flow rate and preset flow phase volume expansion coefficient.

[0045] The target temperature refers to the temperature at which the mobile phase needs to be adjusted to achieve the target flow rate. By adjusting the mobile phase temperature to the target temperature, the mobile phase flow rate changes from the initial flow rate to the target flow rate, thereby maintaining a dynamic balance between the separation accuracy and analytical efficiency of the chromatograph. The processing terminal then uses the formula... The target temperature can be obtained by calculation, where, This refers to the target temperature. This refers to the initial temperature. This refers to the initial flow velocity. This refers to the target flow rate. The coefficient of volumetric expansion of the mobile phase is used to correlate temperature with the flow rate of the mobile phase by measuring the change in the volume of the mobile phase with temperature. This allows for the calculation of the target temperature. The ratio of the target flow rate to the initial flow rate is calculated to determine whether it is necessary to increase the temperature to expand the mobile phase and increase its flow rate. The required temperature is obtained by dividing the ratio of the flow rate change by the coefficient of volumetric expansion of the mobile phase. Finally, the required temperature is added to the initial temperature to obtain the target temperature.

[0046] The mobile phase volume expansion coefficient is a parameter used to describe how the volume of the mobile phase changes with temperature. It is calculated by the processing terminal using the formula... The result can be obtained by calculation, where, This refers to the volume expansion coefficient of the mobile phase. This refers to the volume of the mobile phase at a mobile phase temperature of 20°C or 25°C. In one embodiment, when the mobile phase is methanol and the mobile phase temperature is 20°C, =0.00119℃ -1 , This refers to the change in the volume of the mobile phase. In one embodiment, the operator measures the volume of the mobile phase at two different temperatures, and then subtracts the measured data to obtain the change in the volume of the mobile phase. This refers to the change in the temperature of the mobile phase. This formula correlates the flow rate of the mobile phase with the temperature of the mobile phase by quantifying the volume change when the temperature of different types of mobile phases changes. Thus, by adjusting the temperature of the mobile phase, the flow rate of the mobile phase can be adjusted to reach the target flow rate.

[0047] Step S103: Obtain the viscosity of the flowing phase at the target temperature.

[0048] Among them, the viscosity of the mobile phase refers to the physical property used to describe the magnitude of the internal resistance force during the flow of the mobile phase, which is determined by the formula at the processing terminal. The viscosity of the flowing phase can be obtained by calculation, where, This refers to the viscosity of the mobile phase. This refers to the viscosity of the mobile phase at a temperature of 20°C. Taking methanol as an example, this refers to the viscosity of the mobile phase. =0.59 mPa·s, This refers to the temperature coefficient of the mobile phase. Taking methanol as an example, =0.025℃ -1 , The actual mobile phase temperature is measured by a temperature sensor in the column condition monitoring device. Mobile phase viscosity affects the resistance the mobile phase experiences as it flows through the column. Higher viscosity results in greater resistance and thus greater pressure on the chromatography system, affecting the wear and tear on various parts of the instrument. Therefore, by calculating the viscosity of different mobile phases at the target temperature using the mobile phase viscosity calculation formula, the maximum pressure the chromatography system can withstand at that temperature is determined. This pressure value is then used to correct the target temperature, adjusting the mobile phase viscosity to keep the pressure on the chromatography system within a safe range and reduce wear and tear during operation.

[0049] Step S104: Correct the target temperature based on the viscosity of the mobile phase to generate the actual target temperature.

[0050] In this process, after the mobile phase viscosity at the target temperature is determined at the processing terminal, the target temperature is corrected based on the mobile phase viscosity. The specific method is described in [reference needed]. Figure 7 The steps involved generate the actual target temperature. By adjusting the mobile phase temperature to the actual target temperature, the chromatograph can operate safely and ensure the analytical accuracy of the chromatograph for the analyte.

[0051] Step S105: Adjust the column temperature using the preset column adjustment device according to the actual target temperature to promote rapid equilibration of the gradient elution liquid chromatograph.

[0052] After the processing terminal determines the actual target temperature, it controls the column adjustment device to adjust the column temperature based on the actual target temperature. The specific method is described in [reference needed]. Figure 2 This process facilitates rapid equilibration of the gradient elution liquid chromatograph.

[0053] A column adjustment device is a device used to adjust the volume of the stationary phase in a chromatographic column to improve sample analysis efficiency. It includes a column temperature control component and a column volume adjustment component. The column temperature control component is used to regulate and stabilize the column temperature. It consists of heating and cooling modules. When the temperature sensor in the column status monitoring device acquires the column temperature, it transmits the measured temperature data to the processing terminal. The processing terminal compares the measured column temperature with the required column temperature. If they are inconsistent, the processing terminal controls the heating and cooling modules to heat or cool the column to bring it to the required temperature.

[0054] A column volume adjustment assembly is a device used to adjust the available volume within a chromatographic column. It consists of a polyetheretherketone (PEEK) column located at the tail of the column, with a bottom diameter matching the column diameter and a length not exceeding the column length; a horizontal automatic telescopic rod; and multiple PEEK columns with a bottom diameter smaller than the column diameter and a length not exceeding the column length. When the processing terminal sends an adjustment command to the column volume adjustment assembly, the assembly moves the automatic telescopic rod to push, pull back, or switch columns according to the required adjustment volume, thereby changing the available volume of the column.

[0055] Reference Figure 2 The steps for adjusting the column temperature using a preset column adjustment device based on the actual target temperature to promote rapid equilibration of the gradient elution liquid chromatograph include: Step S200: Determine whether the actual target temperature is greater than the preset safe temperature of the chromatographic column.

[0056] The safe temperature of the chromatographic column refers to the maximum temperature to which the chromatographic column can be heated in order to operate normally. In one embodiment, the safe temperature range of the chromatographic column is 40°C-60°C.

[0057] The processing terminal determines whether the actual target temperature is greater than the safe temperature of the chromatographic column, thereby determining whether the chromatographic column is in a safe working state when the actual target temperature is reached.

[0058] Step S2001: If it is not greater than the actual target temperature, then adjust the temperature of the chromatographic column to the actual target temperature using the column temperature control component.

[0059] If the processing terminal determines that the actual target temperature is not greater than the safe temperature of the chromatographic column, it indicates that the chromatographic column is in a safe working state when the actual target temperature is reached. Therefore, the processing terminal controls the chromatographic column temperature control component to adjust the temperature of the chromatographic column to the actual target temperature, thereby enabling the chromatographic column to be quickly balanced and calibrated.

[0060] Step S2002: If the temperature is greater than the safe temperature, the column temperature control component adjusts the column temperature to the safe temperature and the column volume adjustment component adjusts the column volume to compensate for the flow rate of the mobile phase.

[0061] If the processing terminal determines that the actual target temperature is greater than the column's safe operating temperature, it indicates that the column is not in a safe operating state when the actual target temperature is reached. Therefore, the processing terminal controls the column temperature control component based on the column's safe operating temperature to adjust the column temperature to the safe operating temperature, and controls the column volume adjustment component to adjust the column volume. The specific method is described in [reference needed]. Figure 3This process involves steps to compensate for the flow rate of the mobile phase.

[0062] Reference Figure 3 The steps of adjusting the column temperature to the safe column temperature using the column temperature control component and adjusting the column volume using the column volume adjustment component to compensate for the mobile phase flow rate include: Step S300: Obtain the target dead volume and the change in dead volume of the chromatographic column.

[0063] The target dead volume refers to the target blank volume in the chromatographic column that is not filled by the stationary phase. For specific methods of obtaining this volume, please refer to [reference needed]. Figure 4 This process provides data support for obtaining the change in dead volume. The target dead volume can affect the retention time of the analyte and the mobile phase in the chromatographic column, thereby affecting the separation accuracy of the analyte. At the same time, by calculating the ratio between the target dead volume and the initial dead volume, the slope of the correlation function that is linearly related to the change in dead volume can be obtained, thus obtaining a reference value for adjusting the stationary phase volume. Based on this value, the column volume adjustment component is controlled to adjust the dead volume of the chromatographic column.

[0064] The change in dead volume refers to the difference between the target blank volume and the initial blank volume in the column that is not filled with stationary phase. By determining whether the change in dead volume is positive, the column volume control component is instructed to increase or decrease the dead volume. A positive change indicates that the column volume control component needs to increase the dead volume, while a negative change indicates that the dead volume needs to be decreased. For specific methods of obtaining this information, please refer to [link to relevant documentation]. Figure 4 This process provides data support for subsequent control of the column volume adjustment component to adjust the column volume in order to compensate for the flow rate of the mobile phase.

[0065] Step S301: Calculate the ratio of the target dead volume to the initial dead volume to generate the dead volume ratio.

[0066] The dead volume ratio is a parameter used to reflect the relative change in the dead volume of the chromatographic column. The dead volume ratio can be obtained by dividing the target dead volume by the initial dead volume through the processing terminal. Since the dead volume is directly proportional to both the stationary phase column length and the inner diameter of the chromatographic column, the required stationary phase length or inner diameter of the chromatographic column can be calculated based on the dead volume ratio. Thus, the column volume adjustment component can be controlled to adjust the dead volume of the chromatographic column based on the calculated target data.

[0067] Step S302: Determine whether the change in dead volume meets the preset requirement for increasing dead volume.

[0068] The requirement to increase the dead volume means that the column volume adjustment component can only be controlled to increase the dead volume of the column when the dead volume change is positive.

[0069] The terminal processing determines whether the change in dead volume meets the requirements for increasing the dead volume, thereby determining whether the dead volume of the chromatographic column needs to be increased or decreased.

[0070] Step S3021: If satisfied, the column volume adjustment component is controlled to increase the dead volume according to the dead volume ratio.

[0071] If the processing terminal determines that the change in dead volume meets the requirement for increasing the dead volume, it indicates that the dead volume of the column needs to be increased through the column volume adjustment component. Therefore, the processing terminal controls the column volume adjustment component to increase the dead volume based on the dead volume ratio. The specific method is described in [reference needed]. Figure 5 This process provides data support for subsequent compensation of the mobile phase flow rate.

[0072] Step S3022: If not satisfied, control the column volume adjustment component to reduce the dead volume according to the dead volume ratio.

[0073] If the processing terminal determines that the change in dead volume does not meet the requirement to increase the dead volume, it indicates that the dead volume of the column needs to be reduced through the column volume adjustment component. Therefore, the processing terminal controls the column volume adjustment component to reduce the dead volume based on the dead volume ratio. The specific method is described in [reference needed]. Figure 6 This process provides data support for subsequent compensation of the mobile phase flow rate.

[0074] Reference Figure 4 The steps for obtaining the target dead volume and the change in dead volume of the chromatographic column include: Step S400: Calculate the actual flow rate based on the initial flow rate, the mobile phase volume expansion coefficient, the initial temperature, and the column safety temperature.

[0075] The actual flow rate refers to the volume of mobile phase flowing through the column per unit time from its entry into the column to the detector in the column status monitoring device, as determined by the processing terminal according to the formula. The actual flow velocity is obtained by calculation, where, This refers to the actual flow velocity. This refers to the initial flow velocity. This refers to the volume expansion coefficient of the mobile phase. This refers to the safe temperature of the chromatographic column. This refers to the initial temperature. The logic of this formula is to quantify the impact of temperature changes on the actual flow rate by measuring the expansion of the mobile phase volume with temperature. Since temperature changes cause changes in the mobile phase volume, it is necessary to adjust the flow rate to compensate for the changes in the mobile phase volume, thereby ensuring the accuracy of the chromatograph's detection results for the analyte.

[0076] The initial flow rate and initial temperature in this step are the same as those in step S100 above. The volume expansion coefficient of the mobile phase in this step is the same as that in step S102 above. The safe column temperature in this step is the same as that in step S200 above, and will not be repeated here.

[0077] Step S401: Calculate the product between the actual flow velocity and the target delay time to generate the target dead volume.

[0078] The target dead volume in this step is the same as the target dead volume in step S300 above. The target dead volume can be obtained by multiplying the actual flow rate by the target delay time through the processing terminal.

[0079] Step S402: Calculate the difference between the initial dead volume and the target dead volume to generate the change in the initial dead volume.

[0080] The initial dead volume change refers to the difference between the target blank volume and the initial blank volume in the chromatographic column that are not filled by the stationary phase, without considering the influence of temperature on the volume of the stationary phase. The initial dead volume change can be obtained by subtracting the target dead volume from the initial dead volume through the processing terminal.

[0081] Step S403: Calculate the difference between the safe temperature of the chromatographic column and the initial temperature to generate the column temperature change.

[0082] The change in column temperature refers to the temperature difference between the initial temperature and the maximum achievable temperature within the column. This change in column temperature can be obtained by subtracting the initial temperature from the safe temperature of the column using a processing terminal.

[0083] Step S404: Calculate the product between the preset thermal expansion coefficient of the stationary phase and the change in column temperature to generate the dead volume effect of the stationary phase.

[0084] The dead volume effect of the stationary phase refers to the change in dead volume caused by the change in stationary phase volume due to temperature. The dead volume effect of the stationary phase can be obtained by multiplying the thermal expansion coefficient of the stationary phase by the change in column temperature. Since the temperature inside the column affects not only the volume of the mobile phase but also the volume of the stationary phase, and the column temperature is directly proportional to the volume of the stationary phase, the dead volume of the column is compressed. Therefore, when calculating the dead volume change, it is necessary to consider not only the influence of the mobile phase on the dead volume but also the influence of the change in stationary phase volume on the dead volume, so as to make the calculated dead volume change more accurate.

[0085] The coefficient of thermal expansion of the stationary phase refers to the proportionality of the change in volume of the stationary phase with temperature. In one embodiment, the operator can obtain the coefficient of thermal expansion of the stationary phase by consulting the technical manual based on the column model and the material of the stationary phase.

[0086] Step S405: Calculate the difference between the fixed phase dead volume influence and the initial dead volume change to generate the dead volume change.

[0087] The amount of dead volume change in this step is the same as the amount of dead volume change in step S300 above. The amount of dead volume change can be obtained by subtracting the amount of dead volume influence of the stationary phase from the amount of initial dead volume change through the processing terminal.

[0088] Reference Figure 5 The steps for increasing the dead volume using the column volume adjustment component based on the dead volume ratio include: Step S500: Collect the initial column length of the preset stationary phase.

[0089] The initial column length refers to the length of the stationary phase in the chromatographic column before equilibration calibration. In one embodiment, the operator can obtain the initial column length by consulting the chromatographic column technical manual according to the column model.

[0090] The stationary phase in this step is the same as the stationary phase in step S100 above, and will not be described again here.

[0091] Step S501: Calculate the product between the initial column length and the dead volume ratio to generate the target column length.

[0092] The target column length refers to the actual length that the stationary phase needs to reach in the chromatographic column. The target column length can be obtained by multiplying the initial column length by the dead volume ratio through the processing terminal. Under the premise that the thickness of the stationary phase attached to the chromatographic column remains unchanged, the volume of the stationary phase can be increased by increasing the column length, thereby increasing the dead volume of the chromatographic column and thus enhancing the separation ability of the chromatographic column.

[0093] Step S502: Calculate the difference between the target column length and the initial column length to generate the column length adjustment amount.

[0094] The column length adjustment amount refers to the length that needs to be adjusted when the stationary phase in the chromatographic column changes from the initial length to the target length. The column length adjustment amount can be obtained by subtracting the initial column length from the target column length through the processing terminal.

[0095] Step S503: Collect the actual position of the column volume adjustment component.

[0096] The actual position of the column volume adjustment component refers to the coordinates of the center of the column volume adjustment component within the column before the start of equilibration calibration. A two-dimensional coordinate system is mapped onto the inside of the column using a processing terminal, with the column center as the origin. A laser displacement sensor in the column status monitoring device emits a laser towards the column volume adjustment component, thus obtaining the distance between the laser displacement sensor and the component. The processing terminal then subtracts half the column length from the measured distance and adds half the length of the polyether ether ketone (PEEK) column. The inverse of this difference is calculated and used as the abscissa of the actual position of the column volume adjustment component. The ordinate of the actual position is set to 0. Finally, the processing terminal combines the abscissa and ordinate to obtain the actual position of the column volume adjustment component.

[0097] The column length refers to the distance between the center of the top surface of the column and the center of the bottom surface of the column. In one embodiment, the column length can be obtained by the operator by looking up the column length in the column technical manual.

[0098] Step S504: Query the value of the horizontal coordinate in the actual position of the column volume adjustment component to generate the actual position length of the adjustment component.

[0099] The actual position length of the adjustment component refers to the absolute value of the horizontal axis in the position coordinates of the column volume adjustment component. The horizontal axis value is obtained by querying the column volume adjustment component through the processing terminal, and then the absolute value of the horizontal axis value is obtained by the processing terminal.

[0100] Step S505: Calculate the difference between the actual position length of the adjustment component and the column length adjustment amount to generate the target displacement of the adjustment component.

[0101] The target displacement of the adjustment component refers to the set of lengths and directions that the column volume adjustment component needs to move in the column. The target displacement of the adjustment component can be obtained by subtracting the column length adjustment amount from the actual position length of the adjustment component through the processing terminal. The target displacement of the adjustment component is a data with direction and distance. By determining the target displacement of the adjustment component, the moving direction and moving length of the column volume adjustment component can be known, thereby increasing the column length of the stationary phase and thus increasing the dead volume of the column.

[0102] Step S506: The column length of the stationary phase is adjusted by the column volume adjustment component according to the target displacement of the adjustment component to increase the dead volume.

[0103] In this process, after the processing terminal determines the target displacement of the adjustment component, it determines whether the target displacement of the adjustment component is positive. If the processing terminal determines that the target displacement of the adjustment component is positive, it controls the column volume adjustment component to move the polyether ether ketone column to the right according to the column length adjustment amount, thereby increasing the column length of the stationary phase and thus increasing the dead volume.

[0104] Reference Figure 6 The steps for reducing dead volume by controlling the column volume adjustment component based on the dead volume ratio include: Step S600: Collect the initial inner diameter of the chromatographic column.

[0105] The initial inner diameter refers to the bottom diameter of the cylindrical channel in the chromatographic column before equilibration calibration, which is not occupied by the stationary phase. The laser displacement sensor in the chromatographic column status monitoring device emits a laser to the inner wall of the chromatographic column and receives the reflected laser to obtain the time from the laser emission to its return. The initial inner diameter is obtained by multiplying the above time and the speed of light by the processing terminal.

[0106] Step S601: Take the square root of the dead volume ratio and divide it by the initial inner diameter to generate the target inner diameter.

[0107] The target inner diameter refers to the required bottom diameter of the cylindrical channel not occupied by the stationary phase in the chromatographic column. It is obtained by calculating the square root of the dead volume ratio at the processing terminal and then dividing the initial inner diameter by this value. Under the premise of keeping the stationary phase column length unchanged, by increasing the required bottom diameter of the cylindrical channel not occupied by the stationary phase, the volume of the stationary phase is reduced, the dead volume of the chromatographic column is reduced, and thus the separation ability of the chromatographic column is weakened.

[0108] Step S602: Adjust the column inner diameter according to the target inner diameter using the column volume adjustment component to reduce the dead volume.

[0109] In this process, after the processing terminal determines the target inner diameter, it controls the column volume adjustment component to adjust the inner diameter of the column to reduce the dead volume. The processing terminal switches the polyether ether ketone column in the column volume adjustment component to a column with the same inner diameter as the target inner diameter, thereby controlling the column volume adjustment component to adjust the initial inner diameter to the target inner diameter, and thus reducing the dead volume.

[0110] Reference Figure 7 The steps for correcting the target temperature based on the viscosity of the mobile phase to generate the actual target temperature include: Step S700: Substitute the mobile phase viscosity, preset column parameters, and target flow rate into Poiseuille's law to calculate the predicted system pressure corresponding to the target flow rate.

[0111] The predicted system pressure refers to the system pressure value when the mobile phase flows through the chromatographic column at the target flow rate and mobile phase viscosity. This pressure is determined by processing the terminal according to Poiseuille's law. The predicted system pressure can be obtained by calculation, where, This refers to predicting system stress. This refers to the viscosity of the mobile phase. This refers to the length of the chromatographic column. This refers to the target flow rate. It refers to pi, in For example, =3.14, This refers to the target inner diameter. The logic of this formula is that pressure is directly proportional to flow resistance, which is directly proportional to the viscosity of the mobile phase, the length of the chromatographic column, and the flow rate of the mobile phase, and inversely proportional to the fourth power of the target inner diameter. By predicting the pressure that the chromatograph is subjected to under the current mobile phase conditions, it is determined whether the pressure resistance threshold of the chromatograph is exceeded. If it is exceeded, the pressure on the chromatograph is compensated by adjusting the column temperature to avoid hardware damage.

[0112] Column parameters refer to the data set that stores the target inner diameter and column length. The column parameters are obtained by summarizing the bottom diameter and column length of the column through the processing terminal.

[0113] The viscosity of the mobile phase in this step is the same as that in step S103 above. The column length in this step is the same as that in step S503 above. The target inner diameter in this step is the same as that in step S600 above. These details will not be repeated here.

[0114] Step S701: Determine whether the predicted system pressure is greater than the preset safe system pressure.

[0115] The safety system pressure refers to the maximum pressure value that the chromatograph can withstand under normal operating conditions. In one embodiment, an operator can find the maximum safe pressure value of the chromatograph and the upper limit of the column pressure in the chromatograph manufacturer's technical manual and the column manufacturer's instructions, respectively, based on the chromatograph model and the column model. The maximum safe pressure value of the chromatograph and the upper limit of the column pressure are then transmitted to the processing terminal. The processing terminal compares the maximum safe pressure value of the chromatograph and the upper limit of the column pressure, and the lower value is the safety system pressure.

[0116] After the processing terminal determines the prediction system pressure, it judges whether the prediction system pressure is greater than the safety system pressure to determine whether the chromatograph can work normally at the prediction system pressure corresponding to the target flow rate.

[0117] Step S7011: If the target flow rate is greater than the target flow rate, the target flow rate is reduced in a stepwise manner, and the target temperature is corrected based on the van der Hoff equation to generate the actual target temperature.

[0118] If the processing terminal determines that the predicted system pressure is greater than the safety system pressure, it indicates that the chromatograph may not be able to work properly at the predicted system pressure corresponding to the target flow rate. Therefore, the processing terminal reduces the target flow rate in a stepwise manner. The processing terminal multiplies the target flow rate by the flow rate reduction ratio to obtain the target flow rate reduction value. Subtracting the target flow rate reduction value from the target flow rate gives the adjusted target flow rate. The target temperature is then re-determined based on the adjusted target flow rate and is set as the actual target temperature.

[0119] The velocity reduction ratio refers to the percentage of the target velocity reduction value in the target velocity during the step-by-step reduction process. In one embodiment, the velocity reduction ratio is 10%.

[0120] The actual target temperature in this step is the same as the actual target temperature in step S104 above. The actual target temperature is calculated by the processing terminal using the van der Hoff equation. The actual target temperature can be obtained by calculation, where, This refers to the actual target temperature. This refers to the target temperature. This refers to the gas constant, in which... =8.314 J·mol -1 ·K -1 For example, This refers to the target delay time at the target temperature. This refers to the target delay time corresponding to the adjustment of the target flow rate. This refers to the enthalpy change of the mobile phase, in order to Taking -3.5 kJ·mol⁻¹ as an example, this formula combines the relationship between the column delay time and the mobile phase flow rate and the influence of the mobile phase energy change on temperature. By quantifying the relative change of the target delay time and combining it with the gas constant and the square of the initial temperature, the actual target temperature is finally obtained by correcting the enthalpy change of the mobile phase. Thus, by adjusting the temperature to compensate for the flow rate change, the influence of the flow rate change on the delay time is compensated, and the separation accuracy of the chromatograph for the analyte is further improved.

[0121] The target delay time corresponding to the adjusted target flow rate is processed by the terminal according to the formula. In the formula, This refers to the target flow rate. This refers to the adjusted target flow rate.

[0122] Step S7012: If it is less than, continue to obtain the predicted system pressure for cyclic judgment.

[0123] If the processing terminal determines that the predicted system pressure is less than the safety system pressure, then the predicted system pressure and the safety system pressure are continuously compared to continuously monitor the chromatograph pressure.

[0124] Based on the same inventive concept, embodiments of this application provide a rapid equilibration calibration method for a gradient elution liquid chromatograph, comprising: The data acquisition module is used to acquire initial temperature, initial dead volume, initial flow rate, initial column length, actual position of the column volume adjustment component, and initial inner diameter. A memory for storing a program for a rapid equilibrium calibration method for a gradient elution liquid chromatograph; The processor and the program in the memory can be loaded and executed by the processor to implement a rapid equilibrium calibration method for a gradient elution liquid chromatograph.

[0125] Those skilled in the art will clearly understand that, for the sake of convenience and brevity, the above-described division of functional modules is used as an example. In practical applications, the above functions can be assigned to different functional modules as needed, that is, the internal structure of the device can be divided into different functional modules to complete all or part of the functions described above. The specific working process of the system, device, and unit described above can be referred to the corresponding process in the foregoing method embodiments, and will not be repeated here.

[0126] This application provides a computer-readable storage medium storing a computer program that can be loaded by a processor and executed as a rapid equilibrium calibration method for a gradient elution liquid chromatograph.

[0127] Computer storage media include, for example, USB flash drives, portable hard drives, read-only memory (ROM), random access memory (RAM), magnetic disks, optical disks, and other media that can store program code.

[0128] Based on the same inventive concept, embodiments of this application provide a smart terminal, including a memory and a processor, wherein the memory stores a computer program that can be loaded by the processor and executed as a rapid equilibrium calibration method for a gradient elution liquid chromatograph.

[0129] Those skilled in the art will clearly understand that, for the sake of convenience and brevity, the above-described division of functional modules is used as an example. In practical applications, the above functions can be assigned to different functional modules as needed, that is, the internal structure of the device can be divided into different functional modules to complete all or part of the functions described above. The specific working process of the system, device, and unit described above can be referred to the corresponding process in the foregoing method embodiments, and will not be repeated here.

[0130] The above are all preferred embodiments of this application and are not intended to limit the scope of protection of this application. Any feature disclosed in this specification (including the abstract and drawings) may be replaced by other equivalent or similar features unless specifically stated otherwise. That is, unless specifically stated otherwise, each feature is only one example of a series of equivalent or similar features.

Claims

1. A rapid equilibrium calibration method for a gradient elution liquid chromatograph, characterized in that, include: The initial temperature, initial dead volume, and initial flow rate of the preset chromatographic column and the preset mobile phase are collected. Calculate the quotient between the initial dead volume and the preset target delay time to generate the target flow rate; The target temperature is generated by calculating based on the initial temperature, initial flow rate, target flow rate, and preset mobile phase volume expansion coefficient. Obtain the viscosity of the flowing phase at the target temperature; The target temperature is corrected based on the viscosity of the mobile phase to generate the actual target temperature; The column temperature is adjusted by the preset column adjustment device according to the actual target temperature to promote rapid equilibration of the gradient elution liquid chromatograph.

2. The rapid equilibration calibration method for a gradient elution liquid chromatograph according to claim 1, characterized in that, The column adjustment device includes a column temperature control component and a column volume adjustment component. The steps of adjusting the column temperature according to the actual target temperature using the preset column adjustment device to promote rapid equilibration of the gradient elution liquid chromatograph include: Determine whether the actual target temperature is greater than the preset safe temperature of the chromatographic column; If it is not greater than the actual target temperature, the column temperature control component will adjust the column temperature to the actual target temperature. If the temperature is greater than the safe temperature, the column temperature control component will adjust the column temperature to the safe temperature and the column volume adjustment component will adjust the column volume to compensate for the flow rate of the mobile phase.

3. The rapid equilibrium calibration method for a gradient elution liquid chromatograph according to claim 2, characterized in that, The steps of adjusting the column temperature to the safe column temperature using the column temperature control component and adjusting the column volume using the column volume adjustment component to compensate for the mobile phase flow rate include: Obtain the target dead volume and the change in dead volume of the chromatographic column; Calculate the ratio of the target dead volume to the initial dead volume to generate the dead volume ratio; Determine whether the change in dead volume meets the preset requirement for increasing dead volume; If satisfied, the column volume adjustment component is used to increase the dead volume based on the dead volume ratio. If the condition is not met, the column volume adjustment component is used to reduce the dead volume based on the dead volume ratio.

4. The rapid equilibrium calibration method for a gradient elution liquid chromatograph according to claim 3, characterized in that, The steps for obtaining the target dead volume and the change in dead volume of a chromatographic column include: The actual flow rate is calculated based on the initial flow rate, the mobile phase volume expansion coefficient, the initial temperature, and the column safety temperature. Calculate the product between the actual flow velocity and the target delay time to generate the target dead volume; Calculate the difference between the initial dead volume and the target dead volume to generate the change in the initial dead volume; Calculate the difference between the column's safe temperature and the initial temperature to generate the column temperature change. Calculate the product between the preset thermal expansion coefficient of the stationary phase and the change in column temperature to generate the dead volume effect of the stationary phase; The difference between the stationary phase dead volume effect and the initial dead volume change is calculated to generate the dead volume change.

5. The rapid equilibrium calibration method for a gradient elution liquid chromatograph according to claim 3, characterized in that, The steps for increasing the dead volume using the column volume adjustment component based on the dead volume ratio include: The initial column length of the preset stationary phase is collected; Calculate the product between the initial column length and the dead volume ratio to generate the target column length; Calculate the difference between the target column length and the initial column length to generate the column length adjustment amount; The actual position of the column volume adjustment component was recorded. Query the x-coordinate value of the actual position of the column volume adjustment component to generate the actual position length of the adjustment component; Calculate the difference between the actual position length of the adjustment component and the column length adjustment amount to generate the target displacement of the adjustment component; The column volume adjustment component adjusts the column length of the stationary phase to increase the dead volume, based on the target displacement of the adjustment component.

6. The rapid equilibrium calibration method for a gradient elution liquid chromatograph according to claim 3, characterized in that, The steps for reducing dead volume by controlling the column volume adjustment component based on the dead volume ratio include: The initial inner diameter of the chromatographic column was collected; Take the square root of the dead volume ratio and divide it by the initial inner diameter to generate the target inner diameter; The column volume adjustment component, based on the target inner diameter, adjusts the inner diameter of the column to reduce dead volume.

7. The rapid equilibrium calibration method for a gradient elution liquid chromatograph according to claim 1, characterized in that, The steps for correcting the target temperature based on the viscosity of the mobile phase to generate the actual target temperature include: The mobile phase viscosity, preset column parameters, and target flow rate are substituted into Poiseuille's law to calculate the predicted system pressure corresponding to the target flow rate. Determine whether the predicted system pressure is greater than the preset safe system pressure; If the target flow rate is greater than the target flow rate, the target flow rate is reduced in a stepwise manner, and the target temperature is corrected based on the van der Hoff equation to generate the actual target temperature. If it is less than, then continue to obtain the predicted system pressure and perform loop judgment.

8. A rapid equilibration calibration system for a gradient elution liquid chromatograph, characterized in that, include: The data acquisition module is used to acquire initial temperature, initial dead volume, and initial flow rate. A memory for storing a program for a rapid equilibration calibration method for a gradient elution liquid chromatograph as described in any one of claims 1 to 7; The processor and the program in the memory are capable of being loaded and executed by the processor to implement the rapid equilibrium calibration method for the gradient elution liquid chromatograph as described in any one of claims 1 to 7.

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