AimsInternational proficiency tests are oneof the main objectives of the Society ofHair Testing. The 2012 proficiencytesting of the alcohol consumptionmarkers EtG and FAEE was conductedon behalf of the SoHT. Beyond that, thisinter‐laboratory comparison aimed atestablishing the current degree ofequivalence of measurement resultsfrom different laboratories bymeasuring similar test samples.

Providing hair samples with differentcharacteristics as well as an overview ofthe diversity of analytical methodscurrently applied, could be helpful toolsfor better understanding ofmethodological distinctions and maylead to a better interpretation ofproficiency test results.

The 2012 Ethyl Glucuronide (EtG) and Fatty Acid Ethyl Esters (FAEE)Proficiency Testing on behalf of the Society of Hair Testing (SoHT)

1 MEDICHEM Diagnostica GmbH & Co. KG, Steinenbronn, Germany2 BAM Federal Institute for Materials Research and Testing, Berlin, Germany3 University Centre of Legal Medicine, Geneva‐Lausanne, Switzerland

Methods Measurement of hair samples 

prepared in three different ways (authentic, soaked and spiked) within the bounds of the 2012 EtG and FAEE proficiency testing 

Requirement of specific conditions for the pre‐analytic treatment (e.g. with and without washing of hair samples)

Requesting additional calibrations by using the provided solutions

Detailed questionnaire on the procedures applied 

Proficiency assessment of laboratories according to ISO 5725‐5 (based on the robust consensus values derived from the participants` results) 

Processing of results using the certified software PROLabTM Plus (Quodata GmbH, Germany)

ConclusionPulverization and extractionsignificantly contribute to the overallscatter of EtG results betweenlaboratories. The harmonization ofstandard operating procedures isconsidered as a major step towardsimproving the degree of equivalence ofresults from different laboratories.Non‐pulverized homogenous hairsamples containing authentic,incorporated or spiked EtG,respectively, are versatile tools tocompare analytical protocols on theinter‐laboratory level.

References[1] S.C. Turfus, J. Beyer, D. Gerostamoulos, O.H.

Drummer. A comparison of the performance ofquality controls prepared from spiked, fortifiedand authentic hair for ethyl glucuronide analysis.Forensic Sci. Int. 232 (2013) 60‐66 .

[2] M.E.Albermann, F. Musshoff, L. Aengenheister, B.Madea. Investigations on the influence ofdifferent grinding procedures on measured ethylglucuronide concentrations in hair determinedwith an optimized and validated LC‐MS/MSmethod, Anal. Bioanal. Chem. 403 (2012) 769‐776.

[3] B. Mönch, R. Becker, I. Nehls. Quantification ofEthyl Glucuronide in Hair: Effect of Milling onExtraction Efficiency. Alcohol and Alcoholism(2013) doi: 10.1093/alcalc/agt059 First publishedonline: June 28, 2013.

AcknowledgmentsWe would like to thank all participants for supporting this inter‐laboratory comparison ‒ with special thanks to Dr. Hans Sachs, Forensic Toxicological Centre FTC Munich forperforming almost all prior testings of hair strands; Mr. Dominic Ammann, BAM Federal Institute for Materials Research and Testing, Berlin; Dr. Clementine Klemm, Instituteof Legal Medicine, St. Gallen and Mr. Martin Hastedt, Toxicological Institute, Charité Berlin for performing the assessments of homogeneity; for additional quality controls andassessments of homogeneity: the laboratory team of the Division of Toxicology and Forensic Chemistry, CHUV Lausanne; Dr. Klaus Singer, MVZ Ettlingen; Mr. Josef Ippisch,synlab Weiden and Dr. Silke Süße, Drogencheck Ulm.

Mr. Insa LôMEDICHEM Diagnostica GmbH & Co. KGKringstraße. 3‐5, DE‐71144 Steinenbronnwww.medichem.deinsa.lo@medichem.de

Dr. Roland BeckerBAM Federal Institute for Materials Research and TestingDivision 1.2 "Organic Chemical Analysis; Reference Materials"Richard‐Willstätter‐Strasse 11, DE‐12489 Berlinroland.becker@bam.de

Dr. Frank Sporkert Centre Universitaire Romand de Médecine LégaleUnité de Toxicologie et Chimie ForensiquesRue du Bugnon 21, CH‐1011 Lausannefrank.sporkert@chuv.ch

Further InformationAvailability of hair reference materials for validation and internal quality control:

www.medichem.de

FAEE Results15 laboratories participated in FAEE analysis (9 returns).

The concentrations covered by 2 authentic, 1 "blank" and 3 soakedsamples were in the range between 89 and 3,218 pg/mg total FAEE.However, the number of returns for FAEE analysis was too small toassess any influence of procedural variants on the results.

Due to the large amount of data, only the FAEE samples A, B and E aredisplayed without the results of additional calibration:

MaterialsPreparation of several hair samples by three different procedures withspecial emphasis on

comparable concentrations

comparable homogeneity

preserving the structural integrity of the hair fibers

The homogenization has been realized by cutting the hair strands veryprecisely into pieces of 1 mm length by using a cut technique developedby MEDICHEM.

Additional preparation of several standard solutions and spiked hairextracts prepared from blank hair.

EtG Results34 laboratories participated in EtG analysis (29 returns).

The spiked sample, assuming that its extraction characteristics beingmost similar to a powdered hair sample, has shown the slightestanalytical problems, displayed by the highest findings for mostlaboratories (mean 49.3 pg/mg) in combination with the lowest robustreproducibility standard deviation (RSD 14.5%). The RSD increased incase of the authentic samples (means 38.7 and 68.6 pg/mg) to 35.6%respectively 27.8% and has reached its highest level for the soakedsample (mean 58.0 pg/mg) with 51.1%.

All produced ring test samples (authentic, soaked, spiked) displayedsufficient and comparable homogeneity in the range of RSD 2.8% to7.0%. Problems as reported for the preparation of quality controlsamples from authentic or soaked hair in the recent literature [1] couldbe avoided using a specifically designed homogenization procedure.

Conspicuously, the reproducibility SD in case of the aqueous solutions isin the range already observed for the hair samples.

EtG concentrations obtained after pulverization have been significantly higher than those obtainedfrom cut hair. Furthermore, pulverization improves the reproducibility, as observed for all types ofmaterial. Suggesting a decreasing quantity of easily extractable EtG, starting from the spiked sampleover both authentic samples to the soaked sample, which has been washed repeatedly afterincorporation to eliminate remaining EtG on the hair surface, the difficulties of extraction appear toincrease analogously. Sonication does not guarantee a complete extraction of EtG. Pulverizationappears to improve the EtG recovery in accordance with recent literature reports [2, 3].

19 laboratories additionally reported EtG quantification results in hair samples A to D. These samplesare calibration‐based with fortified extracts from blank hair and have been provided together:

Calibration‐based fortified extracts from blank hair do not provide a means to improve EtG analysis.

Comparison of Extraction Methods and Sample Types

Error bars: U(mean)

Standard Calibration ‐ Sample EtG B  Additional Calibration ‐ Sample EtG B

Insa Lô1, Roland Becker2, Frank Sporkert3