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Effect of the shampoo Ultra Clean on drug concentrations in human hair

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The influence of the special shampoo Ultra Clean (Zydot Unlimited, Tulsa, Oklahoma) on the results of hair analyses was investigated. Hair samples from persons (n = 14) with a known history of drug abuse were collected at autopsy. The hair samples were divided into separate strands which were analyzed both after washing with Ultra Clean and without treatment. Hair analyses were performed by methanol extraction under sonication, purification by solid phase extraction and GC/MS in SIM mode according to routine procedures for tetrahydrocannabinol (THC), cocaine, amphetamine, methylenedioxyamphetamine (MDA), methylenedioxymethamphetamine (MDMA), methylenedioxyethylamphetamine (MDE), heroin, 6-monoacetylmorphine (6-MAM), morphine, codeine, dihydrocodeine and methadone. All drugs originally present in the hair fibers were still detected after a single application of Ultra Clean. However, a slight decrease in drug concentrations could mostly be observed e.g. cocaine (n = 10) -5%, 6-MAM (n = 12) -9%, morphine (n = 12) -26%, THC (n = 4) -36%. The findings clearly demonstrated that drug substances had not been sufficiently removed from human hair by a single Ultra Clean treatment to drop their concentrations below the limit of detection of the analytical method applied.
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Abstract The influence of the special shampoo Ultra
Clean (Zydot Unlimited, Tulsa, Oklahoma) on the results
of hair analyses was investigated. Hair samples from per-
sons (n = 14) with a known history of drug abuse were
collected at autopsy. The hair samples were divided into
separate strands which were analyzed both after washing
with Ultra Clean and without treatment. Hair analyses were
performed by methanol extraction under sonication, pu-
rification by solid phase extraction and GC/MS in SIM
mode according to routine procedures for tetrahydrocan-
nabinol (THC), cocaine, amphetamine, methylenedioxy-
amphetamine (MDA), methylenedioxymethamphetamine
(MDMA), methylenedioxyethylamphetamine (MDE), her-
oin, 6-monoacetylmorphine (6-MAM), morphine, codeine,
dihydrocodeine and methadone. All drugs originally pre-
sent in the hair fibers were still detected after a single
application of Ultra Clean. However, a slight decrease in
drug concentrations could mostly be observed e.g. co-
caine (n = 10) –5%, 6-MAM (n = 12) –9%, morphine (n =
12) –26%, THC (n = 4) –36%. The findings clearly
demonstrated that drug substances had not been suffi-
ciently removed from human hair by a single Ultra Clean
treatment to drop their concentrations below the limit of
detection of the analytical method applied.
Key words Hair · Drug testing Hair analysis ·
Manipulations · Shampoo effect
Introduction
Hair testing for drugs of abuse has been established as a
routine method in drug monitoring [1–3]. Since a positive
drug test is usually connected with considerable legal or
economic consequences, e.g. the loss of the driving
license, drug abusers try to attain negative test results.
Besides complete shaving, common manipulations are
bleaching or hair dyeing. Ultra Clean, a commercially
available hair care product, is recommended to remove
“medications, chemical build-up and other unwanted im-
purities from within the hair shaft”. The manufacturer
(Zydot Unlimited, Tulsa, Oklahoma) offers a “money
back guarantee” if the product should fail. The aim of the
present study was to investigate the effect of Ultra Clean
on drug concentrations in human hair samples obtained
from chronic drug abusers. The hair samples were ana-
lyzed after washing with Ultra Clean and without any
treatment. Hair analysis was performed by a
methanol/sonication extraction procedure [4, 5] followed
by a further purification step using solid phase extraction.
Material and methods
Hair samples
Hair samples from persons (n = 14) with a known history of drug
abuse were collected at autopsy. The hair samples were collected
from the vertex posterior region. A strand (about 5mm in diame-
ter) was cut as close as possible to the scalp, fixed with string and
enveloped. Root and tip of the hair strand were marked. The fibers
were investigated for morphological parameters including hair
color. The samples were stored under dry conditions at room tem-
perature until analysis.
Sample treatment
Each of the 14 hair samples was divided by length into 4 strands,
2 of which were treated with Ultra Clean prior to analysis. The re-
maining two hair strands were subjected to analysis without any
pretreatment. The cleansing product Ultra Clean, consisting of
shampoo, purifier and conditioner (see Table1), was applied ac-
cording to the directions for use given by the manufacturer. The
J. Röhrich · S. Zörntlein · L. Pötsch · G. Skopp ·
J. Becker
Effect of the shampoo Ultra Clean on drug concentrations
in human hair
Int J Legal Med (2000) 113:102–106 © Springer-Verlag 2000
Received: 21 January 1999 / Received in revised form: 19 April 1999
ORIGINAL ARTICLE
J. Röhrich () · S. Zörntlein · L. Pötsch · J. Becker
Institut für Rechtsmedizin,
Johannes Gutenberg-Universität Mainz, Am Pulverturm 3,
D-55131 Mainz, Germany
G. Skopp
Institut für Rechtsmedizin und Verkehrsmedizin,
Ruprecht-Karls-Universität Heidelberg, Voßstrasse 2,
D-69115 Heidelberg, Germany
hair was first wetted and half of the shampoo was applied for 3min
(step 1). Second the purifier was thoroughly distributed on the hair
and left to work for 3min. Subsequently, the remainder of the
shampoo was applied again for 3min (step 3) before the condi-
tioner was used in the fourth step. After each step the hair was
rinsed well with warm water. Shampoo, purifier and conditioner
were applied and distributed on the hair strands with a small brush.
The hair samples were finally air dried.
Hair extraction
The proximal 4cm of the hair strands were used for analysis. The
hair segment was placed in a polypropylene vial and washed for
5min each with water (5ml), acetone (5ml) and hexane (5ml). Af-
ter drying, the hair was cut into small pieces of about 1mm and
40–80mg was used for analysis. The hair was transferred to a
polypropylene vial and 4ml methanol and 100µl of the internal
standard (IS) mixture (2ng/µl of amphetamine-D
11
, MDA-D
5
,
MDMA-D
5
, MDE-D
6
, cocaine-D
3
, morphine-D
3
, codeine-D
3
and
methadone-D
9
; 0.2ng/µl of THC-D
3
) were added. The closed vial
was sonicated for 4h at 50°C.
Purification
The methanol was evaporated and the residue was dissolved in
7ml of phosphate buffer (0.1M, pH6) containing 400mg of bovine
serum albumin. The mixture was then applied to a solid phase ex-
traction column (Bakerbond SPE C18, 500mg), which had been
conditioned by flushing with 2ml of methanol and 2ml of phos-
phate buffer (0.1M, pH6). The column was rinsed with 1ml of
0.1M acetic acid and dried for 10 min under vacuum. THC was
first eluted with 3ml dichloromethane/acetone (1:1; v/v), followed
by elution of amphetamines, opiates and cocaine with 3ml di-
chloromethane/2-propanol/ammonia (40:10:1, v/v/v). Both ex-
tracts were evaporated under a slight stream of nitrogen at 30°C.
Derivatization
THC was methylated with methyl iodide. First 150µl of a mixture
of dimethylsulfoxide and 60% aqueous tetrabutylammonium hy-
droxide solution (98:2, v/v) was added to the extract. Subse-
quently, 50µl methyl iodide was added and the mixture was vor-
texed. After 5min at room temperature,350µl 0.1N hydrochloric
acid was added. The methylated THC (THC-Me) was then ex-
tracted with 2 ×1ml isooctane. The organic layer was separated
and the solvent evaporated at 30°C in a slight nitrogen stream. For
GC/MS analysis the dry residue was dissolved in 50µl of water-
free ethyl acetate.
Amphetamines and opiates were derivatized by adding 50µl of
pentafluoropropionic anhydride (PFPA) to the extract and the mix-
ture was incubated at 70°C for 30min. The excess pentafluoropro-
pionic anhydride was evaporated at room temperature in a slight
nitrogen stream. For GC/MS analysis the dry residue was dis-
solved in 50µl of water-free ethyl acetate.
GC/MS analysis
For GC/MS analysis of amphetamines, opiates and cocaine a HP-
5 MS capillary column was used. The carrier gas was He (constant
flow: 1 mL/min), the injection volume 1 µl (splitless injection),
the injector temperature 250°C and the transfer line temperature
280°C. The oven temperature program was 2 min isothermally at
60°C, 40°C/min to 170°C, 8°C/min to 270°C and 13min isother-
J. Röhrich et al.: Effect of Ultra Clean on drug concentrations 103
Table 1 Ingredients of the cleansing product Ultra Clean
Components Ingredients
Shampoo (tube 1) Sodium laureth sulfate, aloe vera, cocamido-
propyl betaine, cocamid DEA, sodium PCA,
tetrasodium EDTA, panthenol, citric acid,
sodium thiosulfate, methylparaben, DMDM
hydantoin, sodium chloride, fragrance, coloring
Purifier (tube 2) Aloe vera, propylene glycol, EDTA, potas-
sium sorbate, methylparaben, carbomer-940,
triethanolamine, coloring
Conditioner Aloe vera, geranium maculatum, comfrey,
(tube 3) grapefruit juice, hydrolized animal protein,
N-octadecanol, cetyl trimethyl ammonium
bromide, N-hexadecyl alcohol, methyl-
paraben, propylparaben, fragrance, coloring)
Table 2 Ions measured in
SIM-mode and retention times
of the analyzed compounds as
well as the respective internal
standards (IS)
Compound Ions Retention time
Amphetamine-D
11
-PFP (IS) m/z = 128 (target), 194, 292 5.37 min
Amphetamine-PFP m/z = 91, 118 (target), 190, 281 5.40 min
MDA-D
5
-PFP (IS) m/z = 136 (target), 167, 330 7.20 min
MDA-PFP m/z = 135 (target), 162, 190, 325 7.22 min
MDMA-D
5
-PFP (IS) m/z = 208 (target), 344 8.10 min
MDMA-PFP m/z = 135, 162, 204 (target), 339 8.12 min
MDE-D
6
-PFP (IS) m/z = 224 (target), 359 8.41 min
MDE-PFP m/z = 135, 162, 218 (target), 353 8.44 min
Methadone-D
9
(IS) m/z = 78 (target), 303, 318 12.31 min
Methadone m/z = 72 (target), 223, 294, 309 12.37 min
Cocaine-D
3
(IS) m/z = 85, 185 (target), 306 12.98 min
Cocaine m/z = 82, 182 (target), 303 12.99 min
Morphine-D
3
-2PFP (IS) m/z = 417 (target), 580 13.56 min
Morphine-2PFP m/z = 414 (target), 415, 577 13.58 min
Dihydrocodeine-PFP m/z = 282 (target), 445 13.86 min
Codeine-D
3
-PFP (IS) m/z = 285 (target), 448 13.97 min
Codeine-PFP m/z = 284, 300, 447 (target) 14.00 min
6-MAM-PFP m/z = 204, 361, 414 (target), 473 14.71 min
Heroin m/z = 268, 310, 327 (target), 369 16.98 min
THC-D
3
-Me (IS) m/z = 331 (target), 316, 248 14.55 min
THC-Me m/z = 328 (target), 313, 285, 245 14.58 min
mally at 270°C. EI ionization (70eV) was used. The ions listed in
Table2 were measured in the selected ion monitoring (SIM) mode
(dwell time per ion: 30ms). For analysis of THC the same GC/MS
conditions were applied. The measured ions for THC-Me and
THC-D
3
-Me are also listed in Table2.
Quantification and validation data
For quantification the peak areas of the ions specified as “target”
(see Table2) were used. Quantification was based on peak area ra-
tios relative to the respective internal standard (IS). A 5-point cal-
ibration curve was obtained by measuring spiked hair samples
(100mg) containing 10, 50, 100, 200 and 400ng of the analytes.
The calibrations were linear in the range tested and the correlation
coefficients were > 0.99 for all compounds. The intra-assay preci-
sion was determined by analyzing five spiked hair samples (100mg)
containing 200ng of each drug in one series. The intra-assay coef-
ficients of variation (CV) ranged from 7% to 26% (listed in Table 3).
Inter-assay precision data were obtained from analyses of the
spiked hair samples (200ng), performed on six different days. The
day-to-day coefficients of variation (CV) ranged from 8% to 29%
and the limit of detection (signal-to-noise ratio = 3) was < 0.01
ng/mg for all compounds. Each test series included a drug-free hair
sample as negative control.
Instrumentation and reagents
Instrumentation: Gas chromatograph HP 6890 with auto-sampler
(Hewlett-Packard, Palo Alto, Calif.), mass-spectrometer HP 5973
(Hewlett-Packard), capillary column HP-5 MS (30m, 0.25mm in-
ternal diameter, 0.25µm film thickness; Hewlett-Packard). For son
i-
cation the Elma T 460/H ultrasonic bath (Elma, Singen, Germany)
was used, the 50ml polypropylene vials with screw caps were pur-
chased from Falcon (Lincoln Park, N.J.). All solvents and reagents
were analytical grade. Methanol, acetone, hexane, ethyl acetate,
dichloromethane, isooctane, ammonia, dimethylsulfoxide, tetra-
butylammonium hydroxide, acetic acid and 2-propanol were pur-
chased from E. Merck (Darmstadt, Germany), methyl iodide,
pentafluoropropionic anhydride and bovine serum albumin from
Sigma-Aldrich (Deisenhofen, Germany), solid phase extraction
columns from Mallinckrodt Baker (Griesheim, Germany) and all
drug standard solutions as well as deuterated compounds from Ra-
dian (Austin, Tex.).
Results
It has been already pointed out that each of the 14 hair
samples was divided into 4 strands. Two strands were
treated with Ultra Clean prior to analysis, whereas the re-
maining two strands were analyzed without any treat-
ment. This procedure made it possible to run the test se-
ries in duplicate producing two independent analytical re-
sults for each sample of treated and untreated hair. For
evaluation of the analytical data obtained, the mean val-
ues of the two independently determined drug concentra-
tions were used to reduce the influence of statistical ef-
fects. Therefore all concentrations listed in Tables 4–12
under “Without treatment” and “After Ultra Clean” are
mean values of two independently determined values.
The drug concentrations in the untreated hair samples
were compared to those measured in the corresponding
samples after washing with Ultra Clean. The relative
change in concentration after Ultra Clean was given as a
percentage of the concentration in the untreated sample.
Only those changes which exceeded the intra-assay coef-
ficient of variation (%) were considered to be a decrease
or increase. No effect was assumed if the relative change
was in the range of the intra-assay coefficient of variation.
Table3 summarizes the results and the respective intra-
assay coefficients of variation (CV) of the tested com-
pounds are listed. The mean values and medians of
changes in concentration due to Ultra Clean treatment
were calculated from the percentage differences given in
Tables 4–12. Decreases in concentration after Ultra Clean
are marked by “(d)”, increases by “(i)” and no significant
104 J. Röhrich et al.: Effect of Ultra Clean on drug concentrations
Table 3 Summary of results
*mean value for MDA,
MDMA, MDE; d = decrease;
i = increase; n = no significant
effect
No. of posi- Intra-assay Mean vealue Median of
tive findings CV (%) of change in change in con-
concentration (%) centration (%)
THC 4 8 –36 (d) –51 (d)
Amphetamine 6 10 –41 (d) –51 (d)
MDA/MDMA/MDE 6 7* –9 (d) –11 (d)
Cocaine 10 13 –5 (n) –20 (d)
Heroin 6 20 –19 (n) –51 (d)
6-MAM 12 21 –9 (n) –18 (n)
Morphine 12 12 –26 (d) –22 (d)
Codeine 13 11 –30 (d) –27 (d)
Dihydrocodeine 7 26 11 (i) –32 (d)
Table 4 Changes in THC con-
centration Without After Ultra Differ-
treatment Clean ence
(ng/mg) (ng/mg) (%)
0.03 0.04 35
0.04 0.01 –78
0.57 0.28 –51
0.61 0.29 –51
Table 5 Changes in ampheta-
mine concentration Without After Ultra Differ-
treatment Clean ence
(ng/mg) (ng/mg) (%)
0.11 0.05 –55
0.12 0.07 –44
0.13 0.15 16
0.34 0.17 –48
0.74 0.33 –56
0.94 0.39 –59
changes by “(n)”. The number of positive findings in the
untreated samples for the tested compounds is also given
in Table3. All drugs were also detected in the correspond-
ing hair samples which were treated by Ultra Clean prior
to analysis, so that no difference in the qualitative analyt-
ical results could be observed. The influence of Ultra
Clean on methadone concentrations was not evaluated be-
cause only two of the hair samples were positive.
The detailed results of the concentration changes of
THC, amphetamine, MDA, MDMA, MDE, cocaine,
heroin, 6-MAM, morphine, codeine and dihydrocodeine
are presented in Tables 4–12. The methylenedioxyam-
phetamine derivatives MDA, MDMA and MDE were
combined for the evaluation of Ultra Clean effects on
their concentrations (Table6). Therefore the average intra-
J. Röhrich et al.: Effect of Ultra Clean on drug concentrations 105
Table 6 Changes in MDA, MDMA and MDE concentration
Compound Without treat- After Ultra Difference
ment (ng/mg) Clean (ng/mg) (%)
MDMA 0.02 0.03 61
MDA 0.10 0.10 4
MDA 0.10 0.03 –71
MDE 0.34 0.38 14
MDMA 0.37 0.27 –25
MDMA 0.46 0.30 –36
Table 7 Changes in cocaine
concentration Without After Ultra Differ-
treatment Clean ence
(ng/mg) (ng/mg) (%)
0.08 0.18 120
0.33 0.11 –66
0.65 0.20 –69
2.24 1.18 –47
2.29 2.77 21
2.30 4.25 84
3.94 0.35 –91
4.31 6.07 41
7.59 5.63 –26
23.75 20.19 –15
Table 8 Changes in heroin
concentration Without After Ultra Differ-
treatment Clean ence
(ng/mg) (ng/mg) (%)
0.07 0.02 –75
0.13 0.26 100
0.42 0.56 34
0.44 0.22 –50
0.46 0.14 –69
0.52 0.25 –52
Table 9 Changes in 6-MAM
concentration Without After Ultra Differ-
treatment Clean ence
(ng/mg) (ng/mg) (%)
0.03 0.08 139
0.07 0.02 –71
0.10 0.02 –76
0.16 0.29 80
0.29 0.42 45
0.31 0.08 –76
1.10 0.90 –19
2.67 1.56 –42
7.01 7.18 2
12.17 5.15 –58
18.32 15.47 –16
21.59 18.00 –17
Table 10 Changes in mor-
phine concentration Without After Ultra Differ-
treatment Clean ence
(ng/mg) (ng/mg) (%)
0.04 0.03 –10
0.06 0.07 5
0.06 0.05 –20
0.06 0.02 –62
0.08 0.07 –8
0.24 0.14 –45
0.25 0.21 –15
0.29 0.15 –47
0.63 0.59 –5
2.03 1.52 –25
2.54 1.49 –41
13.27 8.33 –37
Table 11 Changes codeine in
concentration Without After Ultra Differ-
treatment Clean ence
(ng/mg) (ng/mg) (%)
0.03 0.02 –48
0.04 0.01 –61
0.05 0.02 –52
0.05 0.04 –27
0.07 0.03 –59
0.10 0.08 –24
0.16 0.06 –63
0.20 0.15 –23
0.42 0.36 –14
0.61 0.79 29
1.13 1.20 6
4.32 3.43 –21
6.14 4.22 –31
Table 12 Changes in dihy-
drocodeine concentration Without After Ultra Differ-
treatment Clean ence
(ng/mg) (ng/mg) (%)
0.06 0.02 –68
0.14 0.05 –61
0.31 1.08 253
0.32 0.45 42
0.40 0.10 –76
30.52 35.53 16
53.55 36.45 –32
106 J. Röhrich et al.: Effect of Ultra Clean on drug concentrations
assay coefficient of variation of MDA, MDMA and MDE
(7%) was used for the evaluation as increase or decrease.
A relatively small decrease was found for these com-
pounds (mean value: –9%, median: –11%). This must be
considered as a result of the relatively large increase of
61% in the MDMA concentration observed in one sam-
ple. No distinct tendency was found for cocaine (Table7).
Whereas 6 of the 10 cocaine positive samples showed de-
creasing concentrations, in 4 cases considerable increases
up to 120% could be observed. Remarkable is an enor-
mous increase of dihydrocodeine up to 253% observed in
one sample (Table12). It is obvious that this particular
large increase raised the mean value of the changes in di-
hydrocodeine concentrations up to 11%.
Discussion
The mean values of the percentage changes in concentra-
tion (Table3) showed a decrease after Ultra Clean treat-
ment for all tested compounds with the exception of dihy-
drocodeine which had an average increase of 11%. How-
ever, the average decreases of cocaine, heroin and 6-
MAM concentrations could not be considered as signifi-
cant because they were in the range of the respective in-
tra-assay coefficients of variation, although the median
values listed in Table3 clearly demonstrate a general ten-
dency towards decreasing concentrations after Ultra
Clean treatment. The median values of all compounds, ex-
cept 6-MAM, exhibit a distinct decrease in concentration
exceeding the respective intra-assay coefficient of varia-
tion. Even the median value of the changes in dihy-
drocodeine concentrations showed a decrease of –32%.
The considerable differences between mean values and
medians for cocaine, heroin and dihydrocodeine are obvi-
ously caused by particular samples with high increases in
concentration after Ultra Clean treatment. In one sample
the cocaine concentration was 0.08ng/mg without treat-
ment and 0.18ng/mg after Ultra Clean, corresponding to
an increase of 120% (Table7), heroin increased from 0.13
to 0.26ng/mg (100%) in one case (Table8) and an large
increase in dihydrocodeine concentration of 253% (0.31
to 1.08ng/mg) was found in another sample (Table12).
Generally decreases in the range of about –5 to –50%
were observed after washing with Ultra Clean compared
to the untreated samples.
Increased concentrations after Ultra Clean treatment
were mostly found in hair samples containing relatively
small amounts of drug (see Tables 4–10), e.g. for THC
+35% at a concentration of 0.03ng/mg without treatment
and 0.04ng/mg after washing, MDMA +61% (0.02/0.03ng/
mg), cocaine +120% (0.08/0.18ng/mg) or 6-MAM +139%
(0.03/0.08ng/mg). Therefore, it has to be assumed that in-
creased concentrations were mainly caused by fluctua-
tions of the analytical results, since especially at low con-
centration levels the scattering of analytical measure-
ments has a larger influence on the obtained result than at
higher levels [6]. However, it has to be taken into account
that in these particular samples drug substances may have
possibly penetrated the hair fiber during the intensive
washing procedure like other small molecules [7].
The extent of the general decrease in concentration
was different for the particular compounds. For example
THC and amphetamine showed relatively large average
decreases of –36% or –41%, respectively, whereas the av-
erage decrease for cocaine was only –5 %. In considera-
tion of the relatively small number of 14 samples this
might be statistical effects, but this finding could possibly
be a result of different sites and strengths of binding of the
particular analytes to the various morphological hair com-
ponents [8]. It is known that cocaine in particular exhibits
a high melanin affinity and a considerable tendency for
incorporation into hair [9, 10]. In the present study, corre-
lations between change in concentration and the individ-
ual color or type of hair could not be established.
In conclusion the major result of this study was that all
drugs originally present in the tested hair samples were
still detectable after the application of Ultra Clean. There-
fore, our findings clearly demonstrated that the drugs had
not been sufficiently removed by a single Ultra Clean
treatment to drop their concentration below the limit of
detection of the analytical method applied. Overall, al-
though a general tendency towards a decrease in concen-
tration could be observed, the “special purifying effect” of
this product might possibly not have exceeded those ob-
tained by any “usual” shampoo.
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... Given the principal source of drug levels in hair has been proposed to be through active or passive diffusion from the bloodstream [15,16], these factors related to pharmacokinetics may also affect hair antiretroviral concentration. In addition, previous studies also reported that hair-related characteristics (for example, frequency of hair washing and hair cosmetic treatment) might affect hair concentrations of non-HIV drugs (for example, amphetamine) [17,18]. Given drugs in hair may share similar incorporation and 'wash-out' mechanisms, these hair-related factors may affect hair antiretroviral concentration too. ...
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Background: Hair antiretroviral concentration has served as an innovative and objective measure of antiretroviral adherence. However, some factors (e.g., pharmacokinetics and hair characteristics) may contribute to the variability of hair antiretroviral concentration that may threaten the validity and reliability of the hair measure as a biomarker of adherence. This study aimed to examine the potential factors that may influence the measure of hair antiretroviral concentration. Methods: Hair samples from a cohort of 372 people living with HIV (PLHIV) receiving lamivudine (300 mg/day) in Guangxi, China. Lamivudine concentration was analyzed using liquid chromatography-tandem mass spectrometry. Multivariable linear regression was used to evaluate the associations of hair lamivudine concentration with age, sex, ethnicity, height, weight, body mass index, duration of HIV diagnose, duration of current regimen, dosing schedule, concomitant antiretroviral medications, frequency of hair washing, hair care products use, hair cosmetic treatment, and self-reported adherence. Results: Multivariable models revealed that frequency of hair washing (ß = -0.221, p = 0.001), dosing schedule (ß = 0.141, p = 0.036), and self-reported adherence (ß = 0.160, p = 0.002) were associated with hair lamivudine concentration. Conclusions: We observed that, among those potential factors, hair lamivudine concentration was influenced by frequency of hair washing and dosing schedule. Therefore, frequency of hair washing and dosing schedule should be considered in future research using hair lamivudine concentration as a measure of lamivudine exposure and biomarker of adherence.
... According to the variance-threshold t-test, the concentrations of 6-acetylmorphine and morphine in the hair segments that were grown before withdrawal were inversely proportional to the withdrawal time. These changes in the concentrations may relate to the daily cleaning treatment for all of the tested compounds [19] but more research is needed. ...
Article
In recent years, overseas anti‐obesity drugs including amfepramone have flowed into China through the Internet or personal import by travelers. Amfepramone is controlled in China and is not available as a pharmaceutical product. It is obtainable either through the Internet or imported by individuals across the border. The abuse of amfepramone is causing serious health problems. A method for the detection and quantification of amfepramone and its metabolite cathinone in human hair was developed and fully validated using liquid chromatography–tandem mass spectrometry (LC–MS/MS). Approximately 10 mg of hair was weighed and pulverized with extraction solvent (a mixture of methanol: acetonitrile: 2 mM ammonium formate (pH 5.3) (25:29:46, v/v/v)). The limit of detection (LOD) and the limit of quantitation (LOQ) were 5 and 10 pg/mg, respectively. The method was linear over a concentration range from 10 to 10000 pg/mg. The accuracy varied from ‐9.3 to 2.3%, with acceptable intra‐ and inter‐day precision. The validated method was successfully applied to 17 authentic cases. The amfepramone concentrations ranged from 11.7 to 209 pg/mg, with a median of 30.2 pg/mg, and the hair cathinone concentrations ranged from 11.9 to 507 pg/mg, with a median of 54.0 pg/mg. This is the first report of amfepramone concentrations in human hair from amfepramone users. Cathinone can be incorporated into hair after amfepramone use.
Article
Although it has been accepted by most scientists that drugs circulating in blood are eligible to hair incorporation, this cannot be considered as a general statement. A 42-year old man was found dead in his swimming pool. He was living alone, and seen alive 2 days before by a neighbour. Femoral blood, cardiac blood and hair were collected during body examination. Free morphine was identified in femoral blood at 28 ng/mL, corresponding to his treatment for chronic pain (3 × 5 mg daily for 4 months). However, with a limit of quantitation (LOQ) at 10 pg/mg, segmental hair testing (3 × 1 cm) for morphine was negative. In this paper, the author has reviewed the different factors which can be responsible of this discrepancy. Several variables can influence the detection of a drug in hair and the author has listed reasons that can account for the absence of analytical response in hair after drug administration. The drug may not be incorporated in hair. That is the case for large bio-molecules, such as hormones, which cannot be transferred from the blood capillaries to growing cells of hair. Cosmetic treatments (perming, colouring, bleaching) or environmental aggressions (ultraviolet radiation, thermal application) will always reduce the concentrations. In this case, the lack of morphine detection was attributed to the effects of chlorinated water from the swimming pool. A negative hair result is also a result. However, this can be interpreted in three different ways: 1. the owner of the hair did not take or was not exposed to the specific drug, 2. the procedure is not sensitive enough to detect the drug, or 3. something happened after drug incorporation (cosmetic treatment, environmental influence).
Article
Hair analysis is capable of determining both an individual's long‐term drug history and a single exposure to a drug, which can be particularly important for corroborating incidents of drug‐facilitated crimes. As a source of forensic evidence that may be used in a court of law, it must be credible, impartial and reliable, yet the pathways of drug and metabolite entry into hair are still uncertain. Many variables may influence drug analysis results, most of which are outside of the control of an analyst. An individual's pharmacokinetic and metabolic responses, hair growth rates, drug incorporation routes, axial migration, ethnicity, age and gender, for example, all display interpersonal variability. At present there is little standardization of the analytical processes involved with hair analysis. Both false positives and negative results for drugs are frequently encountered, regardless of whether a person has consumed a drug or not. In this regard, we have categorized these variables and proposed a three‐stage analytical approach to facilitate forensic toxicologists, hair analysis experts, judiciaries and service users in the analytical and interpretation process. The analysis of drugs in hair has the potential to be used as a powerful source of forensic evidence. However, the biological processes involved are still not fully understood. This paper discusses the factors that can influence the outcome of hair analysis results and suggests a three‐stage interpretation of the many variables that an analyst should consider when presenting analytical data.
Article
• Hair testing is a complementary approach to document doping agent(s) use • All prohibited substances but hormones should be detectable in hair • Interest and limitations of hair testing for doping agents are reviewed based on the authors' experience • Although a lot of data are available for drugs of abuse, controlled studies are missing for anabolic steroids, diuretics and some unusual classes of substances
Article
Assessment of antiretroviral (ARV) concentration in hair (hair antiretroviral concentration [HAC]) is one of the latest non-invasive innovations for measuring long-term ARV adherence. We performed a systematic review following Preferred Reporting Items for Systematic Reviews and Meta-analysis guidelines to identify the factors that may affect the validity and reliability of HAC. This review included 25 studies that reported data on the associations of hair concentrations of 10 ARVs with 22 potential factors related to HAC. Notwithstanding scarce data and some inconsistencies, the data from existing studies suggested that (1) HAC was associated with hair types, hair segment position, housing, illegal drugs use, high-risk sexual behaviors, renal function, and genetic factors; (2) HAC was not associated with race/ethnicity, location of sample, ARV side effects, length of ARV treatment, smoking, alcohol use, orange consumption, depression, or anthropometry characteristics; and (3) the relationships of HAC with natural hair color, hair treatment, age, sex, dosing schedule, and liver function need further study. This review of factors related to HAC informed the design, analysis, and interpretation of the future HIV treatment and HIV prevention research utilizing hair concentrations of various ARVs as a biomarker of ARV adherence.
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This paper represents an experimental approach of histology of the human hair fiber in dyeing and diffusion phenomena and its contribution to the interpretation of hair analysis results for drug abuse. Rhodamine B was applied to human hair fibers from either aqueous solution or methanol/ethanol solvent. The experiments were performed on natural hair of different ethnic groups as well as on extensively bleached hair strands. The microscopical study of the pathway of diffusion of rhodamine B into the hair fibers indicated that the reagent had entered the unmodified fibers at the scale edges between the cuticle cells. At the beginning of the diffusion process intercellular diffusion was the preferred route predominantly along the nonkeratinous regions of the cell membrane complex (CMC) and intermacrofibrillar cement. Penetration into the high sulfur regions of the fiber occurred as dyeing in aqueous solution proceeded and resulted in evenly stained cross sections. The dye distribution pattern observed in natural hair exposed to nonaqueous solution showed that rhodamine B did not penetrate the cortex cells as easy as from aqueous solution and selectively stained nonkeratinous regions only. The determination of the amount of dye taken up by the fibers by spectrophotometric analysis demonstrated that samples diffusion generally increased by time and temperature. It also depended on the morphology of the hair sample. The penetration of rhodamine B from aqueous solution was much greater than from methanol/ethanol solvent.
Article
Hair testing is an effective method for identifying chronic drug abusers. The procedure involves a decontamination step, acid hydrolysis in presence of deuterated internal standards, liquid-liquid extraction and analysis by gas chromatography coupled to mass spectrometry. Hair analysis has a wide window of detection ranging from months to years and provides information concerning the severity and pattern of an individual's drug use.
Article
This article reviews the analysis of 31 drugs and drug metabolites in human hair by thin-layer chromatography, high-performance liquid chromatography, gas chromatography, gas chromatography-mass spectrometry and mass spectrometry. The most important detection method after chromatographic separation of the components is the mass spectrometry because of its sensitivity and specificity. Washing steps to exclude external contamination, extraction, derivatization, stationary phases, detection modes and detection limits of the mass spectrometric and gas chromatographic-mass spectrometric procedures are presented in five tables. Additionally, a method for a gas chromatographic-mass spectrometric screening procedure is presented.
Article
We studied the incorporation rates of cocaine and its major metabolites, benzoylecgonine (BE) and ecgonine methylester (EME), into hair from blood. It was demonstrated in our previous report (Nakahara et al. 1992a) that the incorporation rate of cocaine from blood into hair was much higher than those of BE and EME following administration of cocaine. In this study, the incorporation rates of these drugs into rat hair were investigated following independent administration of BE and EME at 10 mg/kg per day for 5 days, and co-administration of cocaine, BE-d3 and EME-d3 at 3, 0.5 and 0.5 mg/kg per day for 10 days. When BE and EME were administered to rats independently, levels of both these drugs were hardly detectable or quite low in hair, though their AUCs in plasma were very high. On co-administration of cocaine, BE-d3 and EME-d3, the deuterium labeled metabolites in hair were negligibly lower in comparison with the unlabeled ones and in particular the concentration of BE in rat hair was significantly higher than that of BE-d3 although the AUC of BE-d3 was higher than that of BE. It was concluded that the incorporation rates of BE and EME into hair were very low in comparison with that of cocaine and most of the BE detected in hair would be a hydrolytic product derived from cocaine in hair matrix after incorporation.
Article
Hair samples taken from 850 individuals with presumed drug abuse were tested simultaneously for delta 9-tetrahydrocannabinol (THC), cocaine, heroin, the primary heroin metabolite 6-monoacetylmorphine (6-MAM) and morphine. The drugs were extracted with methanol under sonication. Compared to other extraction procedures this solvent extraction technique provides high extraction yields and less experimental effort. The analyses were carried out using gas chromatography-mass spectrometry (GCMS) in selected ion monitoring (SIM) mode. This procedure allows the simultaneous detection of amphetamine, methylenedioxyamphetamine (MDA), methylenedioxymethamphetamine (MDMA) and methylenedioxylamphetamine (MDE). THC was found in 104 (12.2%), cocaine in 230 (27%) and 6-MAM in 141 (16.6%) samples. In addition to 6-MAM, morphine was detected in 87 (10.2%) and heroin in 38 samples (4.5%). The concentrations found were in a range 0.009-16.7 ng/mg for THC, 0.037-129.68 ng/mg for cocaine, 0.028-79.82 ng/mg for 6-MAM, 0.045-53.14 ng/mg for heroin and 0.011-7.800 ng/mg for morphine. The statistical distribution of the drug concentrations compared with the self-reported consumption behaviour of the users may possibly lead to a better understanding of the relationship between drug dosage and corresponding concentrations in hair.
Article
To determine the mechanism involved, the incorporation rate (ICR) of drugs into hair was compared to melanin affinity, lipophilicity and membrane permeability. The following 20 drugs were tested; amphetamine, methamphetamine, p-hydroxyamphetamine, p-hydroxymethamphetamine, cocaine, benzoylecgonine, ecgonine methyl ester, morphine, 6-acetylmorphine, phencyclidine, 1-(1-phenyl)-piperidinyl-cyclohexanol, methylenedioxyamphetamine, methylenedioxymethamphetamine, methoxyphenamine, O-desmethyl methoxyphenamine, benzphetamine, norbenzphetamine, deprenyl, lysergic acid diethylamide (LSD) and 11-nortetrahydrocannabinol-9-carboxylic acid (THCA). Their ICRs were represented as the ratios of the drug concentrations in rat hair to AUCs (the areas under the concentration vs. time curves) in rat plasma. Cocaine had the highest incorporation rate understood and there was a 3600 fold difference between the ICR of cocaine and that of THCA, the lowest drug. The melanin affinity of these drugs was determined by incubating a test solution with melanin at 36 degrees C in the dark for 2 h. After incubation and centrifugation, the drug concentration in the filtrate was determined by GC/MS or LC. The drug most affinitive to melanin was cocaine, followed by benzphetamine, phencyclidine, methylenedioxymethamphetamine and LSD. The correlation coefficient between ICR and melanin affinity of the 20 drugs was 0.947 (0.949 excluding THCA). Lipophilicity was calculated from the retention times of HPLC according to Kaliszan's method. Although the correlation coefficient between ICR and lipophilicity was very low (0.201), it rose to 0.770 by removing only THCA. The combination of melanin affinity and lipophilicity brought about a higher correlation (0.979) with the ICRs. Our data also suggested that the higher ICRs of basic drugs than neutral or acidic ones are strongly related to the membrane permeability of the drug based on the pH gradient between blood (pH 7.4) and hair matrix (acidic).
Article
Two GC/MS-procedures for the detection of amphetamine and its methylenedioxy-derivatives (MDA, MDMA and MDE) in hair are presented. In these methods a methanol sonication extraction technique was applied. The extracted drugs were derivatized either with propionic acid anhydride (PSA) or trifluoroacetic acid anhydride (TFA). PSA-derivatives are more stable than TFA-derivatives, but the latter provide more specific mass-spectrometric information, and, therefore, seem to be preferably for amphetamine determination. The detection limit for all compounds was in a range of about 0.01 ng/mg, if at least 50-100 mg of hair were analyzed, independent of the derivatization used.
Article
A biochemical concept for the endogenous incorporation of drug molecules into growing hair is presented. It is based on the principles of transport across biomembranes, on the principles of biotransformation and drug melanin affinity. The approach gives explanations for current observations in hair analysis, which up to date have not been understood sufficiently. Phenomena such as the ratio of parent drug to metabolite in hair, the dependence of incorporation on the physico-chemical properties of the drug, the independence of drug incorporation on active melanogenesis (incorporation into non-pigmented hair) as well as the dependence of drug content on hair pigmentation are elucidated.
Hair analysis for abused drugs is recognized as a powerful tool to investigate exposure of subjects to these substances. In fact, drugs permeate the hair matrix at the root level and above. Evidence of their presence remains incorporated into the hair stalk for the entire life of this structure. Most abusive drugs (e.g. opiates, cocaine, amphetamines, cannabinoids etc.) and several therapeutic drugs (e.g. antibiotics, theophylline, beta 2-agonists, etc.) have been demonstrated to be detectable in the hair of chronic users. Hence, hair analysis has been proposed to investigate drug abuses for epidemiological, clinical, administrative and forensic purposes, such as in questions of drug-related fatalities and revocation of driving licences, alleged drug addiction or drug abstinence in criminal or civil cases and for the follow-up of detoxication treatments. However, analytical and interpretative problems still remain and these limit the acceptance of this methodology, especially when the results from hair analysis represent a single piece of evidence and can not be supported by concurrent data. The present paper presents an updated review (with 102 references) of the modern techniques for hair analysis, including screening methods (e.g. immunoassays) and more sophisticated methodologies adopted for results confirmation and/or for research purposes, with special emphasis on gas chromatography-mass spectrometry, liquid chromatography and capillary electrophoresis.