Development and validation of methods for the spectrophotometric determination of prospidium chloride for identification and assay analysis

Objective. The work is devoted to the development and validation of UV-spectrophotometric analysis of propidium chloride, an antitumour drug of the dyspyropiperazine group. The relevance of the study is due to the need to introduce into the practice of quality control of prospidium chloride analysis method characterised by high sensitivity, accuracy, rapidity and availability.

Material and methods. The object of the study was a medicinal substance-powder Prospidin (Prospidium chloride, series 271222, expiry date until 12.2027g, produced by «Unitehprom BSU», Minsk). Solutions with working concentration range from 0.05 mol/l to 0.2 mol/l for spectrophotometric analysis were prepared using purified water. Absorption spectra with determination of analytical wavelength (λ_max, nm) and value of molar absorption coefficient ε, (l mol^-1cm^-1) in the range from 200 nm to 400 nm were obtained and analysed on Agilent Cary 60 equipment. Validation parameters were determined in accordance with the regulations of the General Pharmacopoeial Article 1.1.0012.15.

Results. Validation evaluation of the developed analytical techniques for the determination of authenticity and active ingredient content by UV spectrophotometry included tests for specificity (in the presence of NaCl 0.9% solution), analytical range (0.05−0.2 mol/L), linearity (r=0.9997), accuracy, precision in terms of repeatability 3.89±0.24 (±SD) and intra-laboratory reproducibility (ε=2.1%) and stability.

Conclusion. The analysis of electronic spectra of aqueous solutions of propidium chloride demonstrated the presence of R-band absorption (German radikalartig − radical, for systems with unshared electron pairs) with a characteristic wavelength in the far-UV region at λ_max=282±0.4 nm, and λ_min=255±0.4 nm. It was found that the absorption at λ_max in the concentration range from 0.05 mol/L to 0.2 mol/L of a solution of propidine chloride containing nitrogen heteroatoms with unshared electron pairs is due to the implementation of the so-called formally forbidden n→σ* electronic transition involving molecular orbitals. The characteristic appearance of the electronic spectrum with λ_max and λ_min, compliance with the Beer–Lambert law in the given concentration range, the calculated value of the molar extinction coefficient ε (l mol^-1cm^-1) − allow to use the above described spectral characteristics to determine the authenticity and content of the active substance in quality control of prospidium chloride.