Assessing Hair Decontamination Protocols for Diazepam, Heroin, Cocaine and Δ9-Tetrahydrocannabinol by Statistical Design of Experiments

Abstract

Prior to toxicological analysis, hair as a matrix requires pre-treatment measures including decontamination, homogenization, and extraction. Decontamination is performed to differentiate between drug present from superficial deposition and drug incorporated from systemic distribution following ingestion. There are many methods for decontamination of hair samples, mostly developed by empirically using a traditional “one factor at a time” (OFAT) approach, in which one independent variable at a time is changed to observe the effect on the dependent variable. The goal of the present work was to compare the efficacy of decontamination protocols using statistical “design of experiments” (DoE), which allows for analysis of multiple variables and interactions within a single experiment. Decontamination parameters included identity of aqueous and organic wash solutions, number of sequential aqueous and organic washes, order of aqueous and organic washes, and duration of each wash. DoE studies were completed to identify optimal decontamination conditions for four abused drugs with varying physiochemical properties. For this purpose, drug-free human hair was externally contaminated with diazepam (DZP), heroin (HER), cocaine (COC), or Δ9-tetrahydrocannabinol (THC). Each analyte was found to have a unique set of decontamination conditions that were most effective. These included three 30-min washes with methanol followed by three with 1% sodium dodecyl sulfide (SDS) for DZP, three 30 s washes with dichloromethane followed by one with water for HER, one 30-s wash with 1% SDS followed by three with dichloromethane for COC, and three 30 min washes with water followed by one with methanol for THC. The results provide proof-of-principle for a DoE approach to identify effective parameters for hair decontamination for a physicochemically diverse group of drugs. The major advantage of DoE is to elucidate combinations of parameters that result in effective removal of surface contamination, a goal that would be challenging to accomplish using an OFAT approach.

Evaluation of extraction parameters in authentic hair reference material using statistical design of experiments

“…This finding is intuitive and consistent with work done by Salomone et al, who found that extraction of EtG was significantly increased when pulverizing the hair as compared to cutting the hair into snippets [22]. These data align with the concept that when hair is pulverized into a powder, the cuticle, where most drugs bind, is more exposed to the extraction solvent (via increased surface area) than when the hair is cut into snippets [10,11]. For example, da Rosa Chagas et al determined that pulverization increased the amount of recovered cocaine and cocaethylene from authentic user hair [23].…”

“…Recently, we reported a 2 4 fractional factorial DoE to systematically evaluate decontamination procedures and extraction parameters for amphetamine, diazepam, heroin, cocaine, and Δ 9 -THC [10,11]. It was determined that DoE was particularly useful for this type of work, because the combinatorial effects of the factors were significant [10,11]. However, these studies used incorporated HRM, which does not account for how drugs incorporate into hair in vivo.…”

“…Recently, we reported a 2 4 fractional factorial DoE to systematically evaluate decontamination procedures and extraction parameters for amphetamine, diazepam, heroin, cocaine, and Δ 9 -THC [10,11]. It was determined that DoE was particularly useful for this type of work, because the combinatorial effects of the factors were significant [10,11].…”