Development and Validation of a Gradient‐HPLC‐PDAD Method for the Identification of Ballpoint Pen Ink Components: Study of Their Decomposition on Aging for Forensic Science Applications

Abstract

A reversed‐phase high performance liquid chromatographic (RP‐HPLC) method, using multi step gradient elution, is described for the simultaneous determination of ball point pen inks: blue violet, fuchsin, pararosaniline, crystal violet, and victoria blue. The analytical column, an Inertsil ODS‐3 5 m, 250 × 4 mm, was operated at ambient temperature. The elution solvents were classified as A: CH3COONH4 0.05 M, B: CH3CN and C: CH3OH. The samples were eluted according to the following ternary gradient: 50% in A, 30% in B and 20% in C as initial conditions, changing to 20% in A, 30% in B and 50% in C in 10 min, and ending to 50% in B and 50% in C after 10 min. A final isocratic step at the composition of 50% in B and 50% in C was incorporated for 5 min. Elution was performed at a flow rate of 0.8 mL/min and observed inlet pressure ranged from 105 to 270 kg/cm2. A diode‐array detector performed monitoring of ink components at 594 nm for the blue inks and 540 nm for red inks. Identification of examined compounds was made by comparing their retention time values and UV spectra in the range 390–596 nm with commercial standards stored in a data library. Detection limits of 0.1–2 ng were obtained per 20 L injection volume, while linearity held up to 50 ng L. The statistical evaluation of the method was examined performing intra‐day (n = 6) and inter‐day calibration (n = 6) and was found to be satisfactory, with high accuracy and precision results. Six commercial ballpoint pens were examined, among them 3 blue and 3 black pens. These samples were analyzed over a period of 20 weeks in order to establish a useful tool for dating entries on documents.