Analytical grade hydralazine hydrochloride (HzHCl) and bromophenol blue indicator (BpB) were supplied from Sigma. Apresoline tablets containing hydralazine hydrochloride (25 mg HzHCl tablet) were purchased from Novartis Pharma AG, Basel, Switzerland.

A UV-visible double beam spectrophotometer with matched quartz cell (1 cm) (model no. Evolution 201) by Thermo Scientific, 81 Wyman Street Waltham, Massachusetts, US was used to record the absorption spectra.

An aqueous mixture of 10 mL 1 × 10⁻³ M of BpB and 3 mL of pH 3 citrate buffer was mixed with 5, 10, 15, 20, and 25 mL of HzHCl of 1 × 10⁻³ M in 100 mL volumetric flasks.

To assay Apresoline tablets containing HzHCl (25 mg/tablet), ten tablets of Apresoline were grounded and weighed (1.0068 g). Apresoline solution of 1 × 10⁻³ M concentration was prepared by dissolving 0.0792 g of the powder in the double distilled water, 10 mL of 1 × 10⁻³ M BpB, and 3 mL pH 3 citrate buffer and 10 mL of 1 × 10⁻³ M Apresoline solution was diluted in a 100 mL volumetric flask to the mark.

The spectrophotometric method was developed and optimized for present purposes. Different experimental trials of buffer concentration, diluent composition, and other chromophoric conditions were performed to choose the optimum condition for the developed method, which resulted in the optical characteristics and statistical analysis for Hz-BpB ion-pair determination shown in Table 1 .

*y = bx + a, where x is the concentration of analyte in μg/mL and y is the absorbance unit.

The method was then validated to confirm that the analytical procedure is suitable for HzHCl quantification in pure and pharmaceutical formulation. The validation characteristics of ICH under consideration in this work are: specificity, linearity, range, limit of quantification (LOQ), limit of detection (LOD), precision, accuracy, solution stability, and robustness.

Preliminary trials of different compositions and concentrations of the constituents were made to choose the optimum conditions suitable for the method of analysis. According to the findings, specific analytical conditions were derived to prepare the standard and pharmaceutical solutions mentioned in section 2.3 and 2.4.

The selection of wavelength used to measure the absorbance of standard and pharmaceutical solutions implies a new band for the Hz-BpB ion pair. Both HzHCl and BpB were scanned separately in Figure 1 and Figure 2 showing a λmax at 328 nm and 590 nm, respectively. Figure 3 showed a new band at λmax = 416 nm generated. This proves that an ion-pair of Hz-BpB is formed.

The peaks of both the standard solution and the pharmaceutical solution were checked for resolution from the nearest peak by scanning each solution separately ( Figure 3 ).

A calibration curve of 5 points was created. The curve showed a linear response of a linear equation y = 0.0518x + 0.022 and a correlation coefficient R² = 0.9937 ( Table 2 , Figure 4 ).

Five samples of Apresoline were analyzed as per the procedure for normal weight taken for analysis. Mean, standard deviation, and percentage relative standard deviation (%RSD) were calculated. The method was found to be precise since the %RSD were ˂ 0.5816% and ˂ 1.8568% for the repeatability and intermediate precision shown in Table 3 . Analytical application of F-test in case of pure solution with pharmaceutical solution was applied to test the effect of the matrix on the method. The results showed F_calc = 11.36, less than that of F_Theor = 19.00, at degree of freedom (v1 and v2) equal to 2 in both solutions. This indicates that the matrix

of the pharmaceutical solution has a negligible effect on the magnitude of the random error associated with the proposed method. This means that the proposed method can be precise and repeatable for both pure and pharmaceutical solutions.

The standard addition method was used to test the accuracy of the method. Three levels of standard quantity equivalent to 50%, 100%, and 125% were added to the sample. The highest recoveries were (96.90 - 99.77), indicating the high accuracy of the method, as shown in Table 4 .

The stability study of Apresoline solution was applied. Test preparation Apresoline solution absorbance was measured at different time intervals; results are shown in Table 5 and Table 6 . The test preparation Apresoline solution was found to be stable up to 8 h at room temperature. The recoveries did not drop below the minimum as the time increased.

*Limit can be calculated by the following formula: $\text{Limit}=\frac{Abs.oftandardinitial-Abs.ofstandardatdifferenttime}{Abs.ofstandardinitial}\times 100$.

The evaluation of robustness showed the reliability of an analysis with respect to deliberate variations in method parameters. Otherwise, if the measurements are varied in analytical method, a precautionary statement should be included in the procedure. The results in Table 7 showed that, within all variance conditions, the assay of test preparation solution was not affected. Therefore, the analytical method can be considered robust.