wherein, C₂Is the concentration of dobutamine hydrochloride in the sample solution, mu g/mL; c is the concentration of the reference solution, mu g/mL; a is the absorbance of the reference solution; a. the₁Is the absorbance of the sample solution.

To further ensure that the concentration and absorbance are linear, the concentration of the control solution and the concentration of the sample solution are preferably selected from 25-50. mu.g/mL, such as 30. mu.g/mL, 35. mu.g/mL, 40. mu.g/mL, 45. mu.g/mL, etc., independently. In the range, the correlation coefficient of the linear equation is more than 99.9%.

In order to improve the detection sensitivity and the detection accuracy, the invention preferably measures the absorbance of the dobutamine hydrochloride at the maximum absorption position, and the detection wavelength of the ultraviolet-visible spectrophotometer is 276-280nm, preferably 278 nm.

The solvents of the control solution and the sample solution are 0.8-1.0% hydrochloric acid solution, preferably 0.9% hydrochloric acid solution.

The concentration of solvent used in the sample solution and the control solution should be the same.

In order to make the result more accurate, the detection method further comprises the step (1') before the step (1): the uv-vis spectrophotometer is blank calibrated with 0.8-1.0% hydrochloric acid solution, preferably 0.9% hydrochloric acid solution.

The solvent concentration for blank correction according to the present invention should be the same as the solvent concentration used for the sample solution.

After the content of the dobutamine hydrochloride in the sample solution is obtained, the content of the dobutamine hydrochloride in the sample solution also needs to be calculated, and the detection method further comprises the following steps:

and when the sample is liquid, multiplying the content of the dobutamine hydrochloride in the sample solution by the dilution times of the sample solution prepared from the sample to obtain the content of the dobutamine hydrochloride in the sample.

In order to ensure the detection accuracy, the sample needs to be diluted to the concentration of about 30-50 mug/mL, so the concentration of the dobutamine hydrochloride in the sample can be obtained by multiplying the dilution factor after the concentration of the dobutamine hydrochloride in the sample solution is calculated.

And when the sample is solid, multiplying the content of the dobutamine hydrochloride in the sample solution by the volume of the solvent to obtain the content of the dobutamine hydrochloride in the sample.

When the sample is solid, the sample needs to be dissolved to obtain a sample solution, so the final calculation result needs to be multiplied by the volume of the solvent used for dissolving the sample to obtain the content of the dobutamine hydrochloride in the sample.

As a specific embodiment of the present invention, the detection method includes the steps of:

(1) dissolving a reference substance in 0.9% hydrochloric acid solution to obtain a reference substance solution with the concentration of 30-50 mug/mL;

(2) dissolving a sample containing dobutamine hydrochloride in a 0.9% hydrochloric acid solution to obtain a sample solution with the concentration of 25-50 mug/mL;

(3) adjusting the detection wavelength of an ultraviolet-visible spectrophotometer to 278nm, and performing blank correction by using a 0.9% hydrochloric acid solution;

(4) detecting the reference substance solution and the sample solution by using an ultraviolet-visible spectrophotometer with the detection wavelength of 278nm, and measuring the absorbance;

(5) calculating the content of dobutamine hydrochloride in the sample by using the following formula;

wherein, when the sample is liquid, the calculation formula is as follows:

C₂the content of dobutamine hydrochloride in the sample is [ mu ] g/mL; c is the concentration of the reference solution, mu g/mL; a is the absorbance of the reference solution; a. the₁Is the absorbance of the sample solution; b is₁Is the dilution factor when preparing the sample solution from the sample;

when the sample is solid, the calculation formula is:

w₁is the percentage content of dobutamine hydrochloride in the sample,%; c is the concentration of the reference solution, mu g/mL; a is the absorbance of the reference solution; a. the₁Is the absorbance of the sample solution; l is₁Volume of solvent used for sample solution, mL; m is the mass of the sample, μ g.

The detection method provided by the invention can be used for detecting any sample containing dobutamine hydrochloride, and preferably is applied to detecting raw materials, prepared intermediates or products and the like used in the preparation process of the dobutamine hydrochloride injection, wherein the sample comprises the dobutamine hydrochloride raw material, the dobutamine hydrochloride injection intermediate or the dobutamine hydrochloride injection.

The dobutamine hydrochloride raw material is used for preparing the dobutamine hydrochloride injection, and the adopted dobutamine hydrochloride raw material has good purity, but still has trace impurities, so that the dobutamine hydrochloride raw material also needs to be detected.

Compared with the prior art, the invention has the following beneficial effects:

the detection method provided by the invention can shorten the detection time, and has the advantages of high measurement accuracy, good stability, good repeatability and good durability.

Detailed Description

The following examples are intended to illustrate the invention but are not intended to limit the scope of the invention.

The technical solution of the present invention is further explained by the following embodiments. It should be understood by those skilled in the art that the examples are only for the understanding of the present invention and should not be construed as the specific limitations of the present invention.

Preparation example 1

This preparation provides a method for preparing a control solution.

The reference substance of dopamine-butylene hydrochloride (the source is China pharmaceutical biological products institute, lot number 101185-201202) is precisely weighed and dissolved in 0.9 percent hydrochloric acid solution to prepare the reference substance solution with the concentration of 40 mu g/mL.

Preparation examples 2 to 5

This preparation provides a method for preparing a control solution. The difference from preparation example 1 was that the concentrations of the control solutions were 25. mu.g/mL (preparation example 2), 50. mu.g/mL (preparation example 3), 20. mu.g/mL (preparation example 4), and 100. mu.g/mL (preparation example 5).

Example 1

The embodiment provides a method for detecting the content of dobutamine hydrochloride in a dobutamine hydrochloride injection intermediate.

(1) Precisely weighing the intermediate of the dobutamine hydrochloride injection, and dissolving the intermediate in a 0.9% hydrochloric acid solution to prepare a sample solution with the sample concentration of 40 mug/mL.

(2) Adjusting the wavelength of the ultraviolet-visible spectrophotometer to 278nm, performing blank correction by using a 0.9% hydrochloric acid solution, respectively placing the sample solution and the control solution prepared in the preparation example 1 into the ultraviolet-visible spectrophotometer to detect, measuring the absorbance, repeating the steps once, and calculating the average value of the absorbance;

(3) calculating according to the following formula, taking the intermediate of the dobutamine hydrochloride injection in the same batch to perform repeated experiments, and repeating three groups;

wherein, C₂The content of dobutamine hydrochloride in the sample is [ mu ] g/mL; c is the concentration of the reference solution, 40 mug/mL; a is the absorbance of the reference solution; a. the₁Is the absorbance of the sample solution; b is₁Is the dilution factor when preparing the sample solution from the sample;

the results are shown in Table 1

TABLE 1

The embodiment and the performance test show that the RSD measured three times is 0.6 percent, which shows that the detection method provided by the invention has good precision, stability and repeatability.

Comparative example 1

The comparative example provides a method for detecting the content of dobutamine hydrochloride in a dobutamine hydrochloride injection intermediate by using a high performance liquid chromatography.

(1) Adjusting the detection wavelength of a high performance liquid chromatograph to 280nm, using octadecylsilane chemically bonded silica as a filler in a chromatographic column, using a mobile phase A as a mobile phase, wherein the mobile phase B is 55:45, the mobile phase A is 2.6g of sodium octane sulfonate and is dissolved by adding 1000mL of water, adding 3mL of triethylamine and shaking up, adjusting the pH value to 2.5 by using phosphoric acid, and the mobile phase B is acetonitrile-methanol (18:82), and the sample injection amount is 20 mu L;

(2) preparing a sample: taking 1mL of the intermediate of the dobutamine hydrochloride injection in the same batch as the example 1, and adding a mobile phase to dilute the intermediate to 0.5 mg/mL;

preparing a reference substance: taking a reference substance of the dobutamine hydrochloride, and adding a mobile phase to dilute the reference substance to 0.5mg/mL for use;

system adaptation solution: taking appropriate amount of 4- (4-hydroxyphenyl) -2-butanone reference substance and dobutamine hydrochloride reference substance, and dissolving with mobile phase to obtain mixed solution containing 0.3mg and 0.5mg in each 1 mL;

(3) respectively sampling a system adaptive solution, a reference substance solution and a sample solution, and recording the separation degree of the system adaptive solution and the reference substance and the peak areas of the reference substance and the sample;

(4) after the separation degree is more than 1.5, the sample content is calculated by adopting the formula of the concentration of a reference substance, the peak area of the sample, the dilution factor of the sample and the peak area of the reference substance, wherein the peak area of the sample is 1236.2, the peak area of the reference substance is 1241.2, the concentration of the reference substance is 0.5mg/mL, and the result is 10.04 mg/mL.

As can be seen from the comparison between the embodiment 1 and the comparative example 1, the detection method provided by the invention can accurately detect the content of dobutamine hydrochloride in the sample, and the detection method of the comparative example 1 needs to be completed within 2 hours, while the preparation method of the invention can be completed within 30min from the preparation of the sample to the final detection, so that the detection time is short, the detection method is simple and rapid, the production period is shortened, the production yield is increased, and the production cost is reduced.

Examples 2 to 5

The embodiment provides a method for detecting the content of dobutamine hydrochloride in a dobutamine hydrochloride injection intermediate. The difference from example 1 is that the control solution provided in preparation example 1 was replaced with the control solutions provided in preparation examples 2 to 5, and the sample solutions prepared at the end of step (1) had concentrations of 25. mu.g/mL (example 2), 50. mu.g/mL (example 3), 20. mu.g/mL (example 4), and 100. mu.g/mL (example 5).

The test results are shown in table 2:

TABLE 2

As can be seen from the examples and performance tests, the concentration of the control and sample is preferably in the range of 25-50 μ g/mL, and the concentration is too high or too low, which may cause the test result to have a deviation in a small range.

Example 6

The embodiment provides a method for detecting the content of dobutamine hydrochloride in a dobutamine hydrochloride injection. The difference from example 1 is that the sample in step (1) is dobutamine hydrochloride injection.

The test results are shown in table 3:

TABLE 3

The embodiment and the performance test show that the detection method provided by the invention is also suitable for the dobutamine hydrochloride injection, and has good test stability and good repeatability.

Comparative example 2

The comparative example provides a method for detecting the content of dobutamine hydrochloride in a dobutamine hydrochloride injection by using a high performance liquid chromatography. The difference from comparative example 1 is that the sample in step (2) was replaced with dobutamine hydrochloride injection of the same batch as in example 6.

The assay result was 9.91. mu.g/mL.

As can be seen from the comparison between example 6 and comparative example 2, the detection method provided by the present invention has high accuracy and can be used in place of high performance liquid chromatography.

Example 7

The embodiment provides a method for detecting a dobutamine hydrochloride raw material. The difference from the example 1 is that the sample in the step (1) is dobutamine hydrochloride raw material, and the calculation method in the step (3) is as follows:

w₁is the percentage content of dobutamine hydrochloride in the sample,%; c is the concentration of the reference solution, 40 mug/mL; a is the absorbance of the reference solution; a. the₁Is the absorbance of the sample solution; l is₁Volume of solvent used for sample solution, mL; m is the mass of the sample, 10. mu.g.

The test results are shown in table 4:

TABLE 4

The embodiment and the performance test show that the detection method provided by the invention is also suitable for the dopol butamine hydrochloride raw material, and has good test stability and good repeatability.

Examples 8 to 9

The embodiment provides a method for detecting the content of dobutamine hydrochloride in a dobutamine hydrochloride injection intermediate. The difference from example 1 is that the inspection wavelengths were adjusted to 270nm (example 8) and 290nm (example 9).

The test results are shown in table 5:

TABLE 5

As can be seen from the examples and performance tests, the wavelength of the present invention is preferably in the range of 276-280nm, and deviation of the test results may possibly occur if the wavelength is not in the range.

Although the invention has been described in detail hereinabove by way of general description, specific embodiments and experiments, it will be apparent to those skilled in the art that many modifications and improvements can be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.

Claims

1. The method for detecting the content of dobutamine hydrochloride is characterized by adopting an ultraviolet-visible spectrophotometry.

2. The detection method according to claim 1, characterized in that the detection method is: and comparing the absorbance of the reference substance with known concentration in a specific wavelength range with the absorbance of the sample, and calculating to obtain the content of the dobutamine hydrochloride in the sample.

3. The detection method according to claim 2, characterized in that it comprises the steps of:

(1) respectively detecting the sample solution and a reference solution with known concentration by using an ultraviolet-visible spectrophotometer, and determining absorbance;

4. The assay of claim 3, wherein the concentration of the control solution and the sample solution is independently selected from 25-50 μ g/mL.

5. The detection method according to claim 3 or 4, wherein the detection wavelength of the UV-Vis spectrophotometer is 276 nm and 280nm, preferably 278 nm.

6. The test method according to any one of claims 3 to 5, wherein the solvent of the control solution and the sample solution is 0.8 to 1.0% hydrochloric acid solution, preferably 0.9% hydrochloric acid solution.

7. The detection method according to claim 6, further comprising, before performing step (1), performing step (1'): the uv-vis spectrophotometer is blank calibrated with 0.8-1.0% hydrochloric acid solution, preferably 0.9% hydrochloric acid solution.

8. The detection method according to any one of claims 3-7, further comprising:

when the sample is liquid, multiplying the content of the dobutamine hydrochloride in the sample solution by the dilution times of the sample solution prepared from the sample to obtain the content of the dobutamine hydrochloride in the sample;

9. The detection method according to any one of claims 3 to 8, characterized in that it comprises the steps of:

wherein, when the sample is liquid, the calculation formula is as follows:

C₁is the concentration of dobutamine hydrochloride in the sample, mu g/mL; c is the concentration of the reference solution, mu g/mL; a isAbsorbance of the control solution; a. the₁Is the absorbance of the sample solution; b is₁Is the dilution factor when preparing the sample solution from the sample;

when the sample is solid, the calculation formula is:

10. The assay of any one of claims 2-9, wherein the sample is selected from a dopamine hydrochloride starting material, a dopamine hydrochloride injection intermediate, or a dopamine hydrochloride injection.