Green Forensic Toxicology: Optimizing Chlordiazepoxide Detection in Milk, Whiskey, and Whiskey Cream Matrices

Forensic toxicology plays a crucial role in detecting and quantifying drugs, toxins, and poisons in biological and non-biological samples to aid criminal and legal investigations. However, traditional approaches often involve significant resource use and pose environmental concerns. This study focuses on the identification and quantification of chlordiazepoxide, a benzodiazepine commonly associated with drug-facilitated crimes, in milk, whiskey, and whiskey cream matrices using Thin Layer Chromatography (TLC) followed by confirmatory UV-Vis spectroscopy. The methodology involved spiking the matrices with chlordiazepoxide, employing liquid-liquid extraction (LLE) with chloroform and water, and conducting preliminary tests such as the Marquis Test and TLC. The chloroform (90:10) solvent system emerged as the most effective for optimal separation, with an average retention factor (Rf) of 0.42 ± 0.03. UV-Vis spectroscopy provided quantitative confirmation, revealing absorption peaks at 246 nm, 262 nm, and 308 nm. Recovery rates varied across matrices, with milk exhibiting lower recovery (88.5%) due to protein interactions, while whiskey and whiskey cream showed higher recoveries (91.7% and 93.2%, respectively) following repeated extraction cycles. The study demonstrates the reliability and cost-effectiveness of combining TLC and UV-Vis spectroscopy for detecting psychoactive substances in complex food and beverage matrices. The findings emphasize the importance of a multi-tiered analysis strategy, aligning with forensic best practices, and highlight the need for confirmatory analysis to ensure legal admissibility. The successful application of these methods reinforces their relevance in high-throughput laboratory settings and their potential to support forensic investigations involving drug-facilitated crimes.