title: An evaluation of washing and extraction techniques in the analysis of ethyl glucuronide and fatty acid ethyl esters from hair samples.
abstract: Ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEEs) are alcohol metabolites measured in hair and are after a decade of research thought to be the best markers in hair to indicate alcoholism and abstinence Forensic Sci. Int. 218 (2012) 2. A great body of work concerning EtG and FAEEs detection in hair has been performed. However, no recent extensive comparison has been made concerning washing and extraction procedures. This work shows that the washing procedure of dichloromethane followed by a methanol rinse of the hair sample removes more than 16% of the FAEEs and 50% of the total EtG that is present in and on the hair. A review of ten washing protocols (where the removal is categorised: high, medium or low) showed that a relatively high percentage of FAEEs was removed and "medium" amount of EtG compared to the other washing protocols. This work shows promising results for the extraction of the FAEEs and the combined extraction of FAEEs and EtG by using 30min of sonication with methanol. More FAEEs were recovered from hair with methanol than with any other extraction solvent including the commonly used dimethyl sulfoxide/heptane mixture. When the sonication time was increased a higher percentage of transesterification of the FAEEs was observed, the extraction was "dirtier" as solids and a colour change was observed whereas the extraction efficiency did not increase. Therefore, washing the hair sample with dichloromethane and methanol followed by an addition of 1ml of methanol and sonication for 30min to extract the FAEEs and EtG from hair is recommended for FAEEs as well as for the combined analysis of EtG and FAEEs. A linear calibration curve (r(2)>0.99) was obtained for all analytes.
authors.affiliation: University of South Wales, ARW Building, Pontypridd CF374 AT, UK. Electronic address: lbossers@glam.ac.uk.
authors.forename: L C A M
authors.initials: LC
authors.lastname: Bossers
authors.affiliation: University of South Wales, ARW Building, Pontypridd CF374 AT, UK. Electronic address: richard.paul@southwales.ac.uk.
authors.forename: R
authors.initials: R
authors.lastname: Paul
authors.affiliation: University of South Wales, ARW Building, Pontypridd CF374 AT, UK. Electronic address: experts@lextox.co.uk.
authors.forename: A J
authors.initials: AJ
authors.lastname: Berry
authors.affiliation: Lextox, The Maltings East Tyndall Street, Cardiff CF24 5EA, UK. Electronic address: antony.berry@southwales.ac.uk.
authors.forename: R
authors.initials: R
authors.lastname: Kingston
authors.affiliation: University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands. Electronic address: carly.middendorp@gmail.com.
authors.forename: C
authors.initials: C
authors.lastname: Middendorp
authors.affiliation: University of South Wales, 4 Forest Grove, Pontypridd CF37 1DL, UK. Electronic address: alan.guwy@southwales.ac.uk.
authors.forename: A J
authors.initials: AJ
authors.lastname: Guwy
mesh_terms: D000437:Alcoholism / Q000378:metabolism; D005591:Chemical Fractionation / Q000379:methods*; D004951:Esterification; D005227:Fatty Acids / Q000032:analysis*; D008401:Gas Chromatography-Mass Spectrometry; D005965:Glucuronates / Q000032:analysis*; D006197:Hair / Q000737:chemistry*; D006801:Humans; D000432:Methanol; D008752:Methylene Chloride
chemical_list: D005227:Fatty Acids; D005965:Glucuronates; C093924:ethyl glucuronide; D008752:Methylene Chloride; D000432:Methanol
keywords: Alcohol; Ethyl glucuronide; Fatty acid ethyl esters; Sample preparation; Washing; extraction