Development and validation of ICPMS methods for simultaneous determination of elemental impurities in topical cream containing ximenynic acid

ExperimentalChemical and laboratory reagents

The Pharma internal standard 1, ultrapure grade hydrogen peroxide (supplied by JT Baker), ICH/USP target element standard A (supplied by Merck), and ICH/USP target element standard B (supplied by Merck) are all products that may be purchased from trustworthy vendors (Merck). The guidelines found on the supplier's certificate of analysis about the proper storage of the test materials were followed. The temperatures at which the test solutions were kept were the same as those employed during the testing. In each and every one of the volumetric flasks that were used for the procedures, polymethyl pentene (PMP) and polypropylene (PP) were both used.

Instrumentation

An Agilent 7800 ICP-MS was used in the gathering of each and every reading. The typical method of sample introduction was replicated with this apparatus by using a Micromist concentric nebulizer, a quartz spray chamber, and a quartz flame that was outfitted with a 2.5 mm id injector. All of these components were included into its construction. Ni fashioned them by hand all by himself (with a Ni-plated Cu-core sampler). In order to stop the build-up of salt on the nebulizer, an Agilent argon gas humidifier was attached to the gas that was being carried by the carrier. The helium mode demonstrates the degree to which the elements are sensitive at the USP levels. During the validation process, both the conventional and the cutting-edge high-energy helium (He) mode were used.

Method development [22, 23]

When determining which digestion technique to use, many factors, including the sample, the analyte, the availability of reagents, and the capabilities of the equipment, all play a part. This is due to the fact that they direct the selection of settings that are ideal for extracting the greatest amount of metals from the sample. The circumstances of the ICPMS, the diluent used in the preparation of the sample, the programmed of the microwave digester, and the settings of the instrument are all examples of aspects that may be optimized.

Selection of ICPMS conditions

On the basis of the most recent and accurate information and the accumulated knowledge at the time, the following criteria were set for each metal: (Table 1).

Selection of diluents for digestion of sample

The sample matrix and the metals of interest are taken into consideration while selecting the digestion reagents to use. Because of their strong oxidizing capabilities, HCl, H2SO4, HF, and HNO3 are the most effective acids for breaking down inorganic and organic compounds, respectively. This is the case for both inorganic and organic substances. In addition to hydrochloric acid and nitric acid, other combinations of concentrated acids with either thirty percent or fifty percent hydrogen peroxide were put through their paces throughout the testing process. It is not typical practice to make use of certain acids since it is difficult to reclaim volatile substances once digestion has taken place. In three different trials, we combined 1 mL of hydrogen peroxide with 5 mL of concentrated nitric acid, 4 mL of concentrated hydrochloric acid with 4 mL of water, and 5 mL of concentrated nitric acid with 1 mL of hydrochloric acid.

Instrument parameters

In order to get started on the job, the following ICPMS parameters were used, which were chosen based on previous knowledge and experience (Table 2).

Table 2 Instrument parametersSelection of microwave digestion system programme

During the digesting process, the length of time that the matrix is exposed to the oxidizing acid is also an extremely important factor. Prolonged exposure has the potential to hasten the exothermic reactions that occur, which in turn makes it easier for the metal of interest to dissolve from the biological matrix and then be released into the air. The following procedure for digesting the samples was devised after the preliminary testing according to the findings of the following tests (Table 3).

Table 3 Digestion programmeFinal methodologySample and standard preparationPreparation of 2%V/V nitric acid and 1% v/v hydrochloric acid (diluent)

Putting 50 mL of water to a volumetric flask with a 100 mL capacity. Add 2.0 mL concentrated nitric acid and 1.0 mL hydrochloric acid after that. Then, until the volume reached a total of 100 mL for the experiment, add some water to it.

Preparation of standard stock solution

Standard A: Transferred 1 mL of the target element (As 15 ppm, Cd 5 ppm, Hg 30 ppm, Pb 5 ppm), and Standard B (Ag 150 ppm, Co 50 ppm, Ni 200 ppm, Se 150 ppm, Tl 8 ppm, V 100 ppm) from the ICH/USP target element to each individual 50 mL volumetric flask. When the diluent was used, the volume was at a level that was satisfactory.

Internal standard preparation

Pharma internal standard-1 solution, 1 ml, was transferred to a 50 mL volumetric flask, and the volume was made up with diluent.

Sample diluent

1 mL of concentrated hydrochloric acid and 5 mL of concentrated nitric acid were combined.

Preparation of sample blank

This digesting tank was supplied with 5 mL of concentrated nitric acid as well as 1 mL of concentrated hydrochloric acid. The timetable for the digestion was adhered to. Following the chilling process, the sample was transferred into a volumetric flask of 50 mL, which was then filled with water.

Preparation of sample solution

The Ximenynic acid Cream was then administered to the digestion vessel once the appropriate quantity had been determined using the measurement device. This digestion vessel was predigested for fifteen minutes with five mL of concentrated nitric acid and one mL of concentrated hydrochloric acid. The digestion process has been finished without any problems. After the liquid had cooled, the sample was transferred to a volumetric flask of 50 mL and the remainder of the space was filled with water.

Preparation of sample solution at 30% level of specification limit (0.25 J)

The Ximenynic acid Cream was then administered to the stomach once the appropriate quantity had been determined using the measurement device. This digestion vessel was predigested for fifteen minutes with five mL of concentrated nitric acid and 1 mL of concentrated hydrochloric acid. Following that, 0.25 mL of the stock solution that served as the gold standard was added. The digestion process has been finished without any problems. After the liquid had cooled, the sample was transferred to a volumetric flask of 50 mL and the remainder of the space was filled with water.

Preparation of sample solution at 100% level of specification limit

The Ximenynic acid Cream was then administered to the stomach once the appropriate quantity had been determined using the measurement device. This digestion vessel was predigested for fifteen minutes with five mL of concentrated nitric acid and one mL of concentrated hydrochloric acid. After that, one mL of the stock solution was put in. The digestion process has been finished without any problems. After the liquid had cooled, the sample was transferred to a volumetric flask of 50 mL and the remainder of the space was filled with water.

Method validations [10, 11, 23]

In line with the recommendations that came out of the International Conference on Harmonization (ICH), the method that has been determined to be the most effective for concurrently measuring seven different elements is ICPMS (R1). We investigated the suitability of the system, linearity, recovery, optimization of LOD and LOQ, method precision, and intermediate precision.

Linearity and range

The ability to get results that are proportionate to the quantity of analyte that was employed in the test is what is meant by the term “linearity”. In order to evaluate the linearity, seven distinct concentrations ranging from the lower limit of quantification (LOQ) to 250% of the specification level were aspirated. The correlation between the analyte's reaction and the ISTD's response was displayed versus the concentration. The calibration curve was used as the basis for the calculation of the slope, intercept, and correlation. If the correlation coefficient (‘R’) value is more than 0.99, this indicates that the data reasonably well match the regression line.

Limit of detection and limit of quantitation

The limit of detection (LOD) refers to the lowest possible concentration of an analyte that may be determined from a sample under the conditions of the experiment. The lowest acceptable concentration (LOQ) of an analyte is specified as the lowest concentration of the analyte that still provides a credible measurement under the conditions of the experiment. A blank sample was made and aspirated ten times before being evaluated. We used the linear regression equation to determine the LOD and LOQ for each element. This equation requires two inputs the standard deviation of the intercept, and the slope in order to function properly.

$$} = 3.3*\sigma/},\;} = 10*\sigma/}$$

where σ = standard deviation of Ratio of analyte response to ISTD response of sample blank, s = slope of calibration curve.

Precision at limit of quantitation

In order to guarantee that the LOQ answer is accurate, six duplicates of it were created (0.25 J level). For the purpose of illustrating LOQ accuracy, relative standard deviation (%RSD) was used; at the LOQ level, the RSD value should be either less than or equal to 20.0.

Specificity

The specificity of the ICPMS technique was discovered via the study that investigated how the calibration blank influenced the recovery research at the LOQ level. These steps were taken to remove any possible impact that might have been caused by the excipients in the formulations. In order to assess the degree of accuracy, a calibration blank was made and then sucked a total of ten times. The mean percentage recovery at the LOQ level should be between 50.0 and 150%, and the average significant interference of the calibration blanks should be lower than that of the calibration standard solution1.

Accuracy (recovery)

The accuracy of an analytical technique may be evaluated by contrasting the expected value with the actual value that was achieved. This number may be obtained by calculating the proportion of recovered elements, denoted as R%. The tried-and-true ICPMS method was put to the test in this circumstance by repeatedly (n = 3) analysing spiked samples at four different concentrations (LOQ, 50%, 100%, and 150% of the LOD). Using the formula [% Recovery = (Recovered content of analyte/Actual spike content of analyte) 100], the experimental data were subjected to a statistical analysis in order to establish the level of recovery achieved by the method as well as its overall validity. The average recovery at the LOQ level of any analyte must be between 50 and 150% for it to be considered acceptable, and the recovery at all other levels must likewise be between 70 and 150% for it to be considered acceptable (Table 4).

Table 4 Preparation of samples for accuracyPrecision

The degree to which distinct studies performed on numerous duplicates provide the same findings may be used to characterize the accuracy of a measurement technique. The system precision, the method precision, and the intermediate precision have all been investigated throughout the studies of precision. Testing the accuracy of the system assured that the results of repeated measurements would be reliable. On the System Precision Solution, the Calibration Blank, the Calibration Standards, and the Standard Check Solution, extractions and analyses were carried out. It is recommended that the correlation coefficient (‘R’) value of the calibration curve be less than 0.99 for each analyte. In order to evaluate the accuracy of the procedure, the Specification Level produced three samples that were left "as is" and six samples that were “spiked” (1 J). The preparation and aspiration of three as-is samples and six individual spike samples at the specification level required a large number of analysts who worked on various days. This was done in order to achieve intermediate accuracy. The relative standard deviation (%RSD) was the metric that was used to evaluate how well the method performed in general. The relative standard deviation (RSD) over all six Spike at Specification level solution preparations should not exceed 15%. In order to prevent RSD readings from being more than 15%, it is necessary to have a high level of both method accuracy and intermediate precision for each analyte.

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