Transcript Ethylglucuronide (EtG) assessed by two methods following

ETHYLGLUCURONIDE (ETG)
ASSESSED BY TWO METHODS
FOLLOWING FRAGRANCE AND HAND
SANITIZER EXPOSURE IN MEN AND
WOMEN
Mollie Starkie, Pharm.D. Candidate
Mercer University
College of Pharmacy and Health Sciences
BACKGROUND
COOH
O
O
OH
HO
CH3
HO
COOH
O
HO
O
OH
HO
Ethylglucuronide
CH3
BACKGROUND
 EtG

Enzyme multiplied immunoassay
technique (EMIT)


Semi-quantitative laboratory screen for
urinary EtG
Liquid chromatography-mass
spectrometry-mass spectrometry
(LC/MS/MS)


analysis
Confirmatory test for urinary EtG
Positive EtG concentrations: range of
100-500 ng/mL
BACKGROUND
 Cologne
and perfume in US contains
78-83% alcohol by volume.
 Hand sanitizer can contain 62%
alcohol by volume.
 Exposure to such products would
increase ethanol exposure.
OBJECTIVES
 Primary

objective:
Evaluate whether positive urine EtG results
were detected following:
1) fragrance exposure alone
 2) combined fragrance plus hand sanitizer exposure in men
and women


when evaluated with a screen (EMIT analysis) and
confirmatory test (LC/MS/MS analysis)
 Secondary


objectives:
Comparison of EMIT analysis to LC/MS/MS
analysis
Detection of gender differences between the
two analyses
HYPOTHESIS
 Combined
fragrance (cologne or perfume)
and hand sanitizer exposure is more likely
to produce positive EtG screen and
confirmation results than the fragrance
exposure alone.
METHODS
METHODS
Days 1-7:
Abstinence
from alcohol
consumption
& use of
cologne,
perfume, or
hand
sanitizer
Day 13:
• Hand sanitizer
applied every 15
minutes for 8
hours
• Urine samples
collected at end
of 8 hour period
& 4 hours later
Day 8:
• Begin BID
fragrance
application
behind ear
• Begin
collection of
first
morning
urine void
Day 18:
End
fragrance
application
Day 21:
End
collection
of urine
URINE SAMPLE DIVISION
Urine Sample
Tube A
•Refrigerated
•EMIT analysis every 3-4
days
Tube B
•Frozen
•LC/MS/MS analysis at
the end of study
DEFINITIONS
 POSITIVE
RESULT: EtG concentration
> 100 ng/mL
 FALSE POSITIVE: EMIT positive screen
result not confirmed by LC/MS/MS
confirmation analysis
 FALSE NEGATIVE: Negative EMIT
screen but positive LC/MS/MS confirmation
result
STATISTICS
 Mixed

Assess difference between male and
female positive results
Mixed

model repeated measures
model Chi Square
Comparison of probability of having EtG
concentrations >100 ng/mL by gender
 Pearson
r coefficient and Kappa
agreement

Evaluated correlation and association of
EMIT and LC/MS/MS analyses
RESULTS
RESULTS
4
males & 4 females provided 128 urine
samples
 Range for EtG values:
EMIT: 0-719 ng/mL
 LC/MS/MS: 0-711 ng/mL

 Mean
scores of EMIT and LC/MS/MS
 EtG positive = > 500 ng/mL, 100%
agreement
 EtG positive = >100 ng/mL, 37% agreement
(r = 0.42, p<0.001)
 Females less likely to test EMIT positive
RESULTS
Men (n=40)
Women (n=39*)
Total (n=79)
Men (n=24)
Women (n=25*)
Total (n=49)
Men (n=64)
Women (n=64)
Total (n=128)
Chart 1: Positive Results
Fragrance Alone Positive Results
EMIT (%)
7 (17.5)
LC/MS/MS (%)
0 (0)
0 (0)
0 (0)
7 (8.9)
0 (0)
Fragrance and Hand Sanitizer Positive Results
EMIT (%)
LC/MS/MS (%)
8 (33)
4 (16.7)
5 (20)
13 (26.5)
Total Positive Results
Total Positives
EMIT (%)
LC/MS/MS(%)
Men (n=64)
15 (23.4)
4 (6.3)
Women (n=64)
5 (7.8)
3 (4.7)
20 (15.6) Total (n=128)
7 (5.5)
3 (12)
9 (18.4)
False
Positives (%)
False
Positives
False11
Positives
(17.2) (%)
11 (17.2)
2 (3.1)
2 (3.1)
13 (10.2)
13 (10.2)
*On the morning of hand sanitizer application, 1 female subject provided first
morning sample after hand sanitizer exposure.
Graph of EMIT vs. LC/MS/MS analyses of female subject
800.0
Legend:
Gray background: EtG levels < 100
ng/mL
Green background: EtG levels 100-500
ng/mL
Brown background: EtG levels > 500
ng/mL
700.0
600.0
EtG concentration (ng/mL)
500.0
400.0
EMIT
LC/MS/MS
300.0
200.0
100.0
0.0
1
-100.0
2
3
4
5
6
7
8
9
10
Collection Number
11
12
13
14
15
16
CONCLUSIONS





Study participants abstained from alcohol beverages
during the study.
Positive EtG concentrations > 100 ng/mL were found
following exposure of fragrance alone and fragrance
and hand sanitizer when evaluated with both
analyses.
False positive results occurred more frequently in
males.
In laboratory screens the biomarker EtG detects
ethanol exposure from sources other than oral
alcoholic intake and should not be used as a sole
method to determine covert alcohol intake with
positive levels defined as > 100 ng/mL.
Further research should be done to determine if false
positives and false negatives results would occur with
concentrations > 500 ng/mL.
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QUESTIONS ??