In spite of the serious flaws in studies purporting to indicate a color bias in hair analysis, the suggestion of such a bias in hair testing continues to receive attention. Some of the limitations in such studies are:
• Failing to adequately wash the hair before extraction to remove sweat and contamination as the source of the measured drug (33-40);
• Extracting hair with sodium hydroxide (NaOH), a method that could never be used for workplace samples because it hydrolyzes 6-monoacetylmorphine and cocaine (33-35,38,39);
• Use of in vitro models that mimic soaking/contamination but are not valid models of in vivo incorporation into the growing hair fiber within the hair follicle (33,36,37);
• Use of animal models, which may not accurately reflect transport and biotransformation processes that occur in humans (33,38-40);
• Extremely small data sets with low statistical significances (41,42);
• Failure to use extraction methods that fully extract the drug from the hair matrix
• Failure to use a method that can exclude melanin from the extraction process itself (34-41).
As pointed out earlier, an efficient extraction method, along with washing of the hair to remove sweat or environmentally deposited drug, is a major component of valid quantitative testing of ingested drug. Any study performed without aggressive washing of the hair samples cannot be interpreted to represent ingestion, much less to assess the presence of a color effect. In this regard, sweat is a complex variable. It is known that individuals vary greatly in the amount of sweat produced, depending on gender, exertion, stress, climate and season, hormonal status, clothing, nutritional and hydration states, and many other factors.
Sweat is produced by two types of glands—eccrine (generally distributed over the entire body) and apocrine (located in the axilla and pubic regions) (3,52). Compounding the variations in sweat production, the kind and frequencies of shampoo and conditioner treatments used with different hairstyles also affect the amount of sweat residues left on hairs. In addition, the effects of an individual's sweat exposure on his/her own hair can vary greatly for different hair types. For example, porous hair may easily soak up hundreds of times more drug than nonporous hair, but such drug can also be removed with similar ease by effective washing procedures (30). With these considerations, studies that purport to show hair-color effects but use inadequate or invalidated decontamination and/or extraction methodologies must be weighted accordingly. Analysis of a large amount of data obtained with washed hair dissolved by enzymatic digestion followed by extraction of the melanin-free component have indeed shown no evidence of color bias (44-52).
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