Maternal-fetal biomarkers of

prenatal exposure to ethanol

Simona Pichini

National Observatory on Alcohol, tobacco,

smoking, doping and gambling

Istituto Superiore di Sanit

Rome, Italy

Indirect Maternal biomarkers of gestational ethanol consumption

-Interview with the mother (the most used,

the most unreliable!)

-Ethanol concentrations in expired air, blood

and urine (very rare)

- Glutamyltransferase (GGT)

-Aspartate aminotransferase (AST)

-Mean corpuscular volume (MCV)

-N-Acetyl--Hexosaminidase (Beta-Hex)

-apolipoprotein J

-Total sialic acid

-Hydroxytryptophol 5-(5-HTOL)

-Phosphatidyl Ethanol (PEth)

-Carbohydrate Deficient Transferrin (CDT)

ACUTE RECENT CHRONIC

DA

YS

Indirect

direct

MARKERS OF ALCOHOL ABUSE

FAEEs (EtOH + fatty acids Synthetase FAEEs)

O

O

CH3CH3

O

O

CH3CH3

O

O

CH3CH3

O

O

CH3CH3

O

O

CH3CH3

O

O

CH3CH3

O

O

CH3CH3

ethyl palmitate

ethyl palmitoleate

ethyl stearate

ethyl oleate

ethyl linoleate

ethyl linolenate

ethyl arachinodate

Ethylglucuronide and Ethylsulfate

EtOH

CH3

O

HOH

HH

H

H

OOH

OH

COOH

UDP-glucuronosyltransferase

-D-ethylglucuronide (EtG)

C8H14O7 (222.1945)

S OH

O

O

OCH3

sulfotransferase

ethylsulfate (EtS)

C2H6O4S (126.1313)

Alcohol dehydrogenase

aldehyde dehydrogenase acetic acid Krebs cycle

Accounting for 0.1% ingested ethanol

Placenta

Fetus

ABSORPTION

DISTRIBUTION

/

METABOLISM

EXCRETION

cord

blood

meconium

Newborn

breast milk

hair

Drug

Drug/Metabolites

Blood/Plasma

Gastrointestinal Tract

faeces

Skin and Lungs

bronchial

secretions/

expired air

Liver

bile

Lungs

Extra-cellular fluids

and matrices:

tears, nails,

cerebrospinal fluid,

interstitial fluid,

seminal fluid,

vitreous humour,

pericardial fluid,

saliva, sweat, hair

amniotic fluid

Kidney

urine

Metabolites

D/M

D/M D/M

D/M

D/M

D/M

D/M

D/M

D/M Urine

D/M

Direct ethanol metabolites in maternal biological

matrices

Fatty acid ethyl esters (FAEEs).

- FAEEs in blood account for acute consumption.

- FAEEs in hair account for chronic consumption as

a function of hair length and segmental analysis

(performed only in two labs worldwide).

Ethyl glucuronide (EtG) and Ethyl sulfate (EtS).

- EtG and EtS in blood account for recent

consumption.

- EtG and EtS in urine increase the window of

detection to the previous 3-4 days.

- EtG and EtS in hair account for chronic

consumption as a function of hair length and

segmental analysis

Biological Matrices

related to gestation-

delivery-neonatal

period

Amniotic Fluid

Placenta

Fetal hair

Meconium

Cord blood

Neonatal urine

Ethyl glucuronide and ethyl sulfate in placenta and fetal tissues

by LC-MS/MS: biomarkers of placental ethanol transport

Luca Morini*1, Maria Falcn2, Simona Pichini3, Oscar Garcia-

Algar4, Paolo Danesino1, Angelo Groppi1 and Aurelio Luna2

1Department of Legal Medicine, Forensic and Pharmaco-Toxicological Science, University of Pavia, Italy; 2Deparment of Legal Medicine, Universidad de Murcia, Murcia, Spain; 3Istituto Superiore di Sanit, Rome, Italy; 4Unitat de Recerca Infancia i Entorn (URIE), Institut Municipal dInvestigaci Mdica-Hospital del Mar, Barcelona, Spain.

http://milano.blogosfere.it/images/ecografia01.jpg

Real samples

EtG EtS Placenta

ng/g

Fetal Tissues

ng/g

Hair

pg/g

Placenta

ng/g

Fetal Tissues

ng/g

S 1-S5

Similarly to amniotic fluid and

placenta, cord blood objectively

assess the acute exposure, which

occurs at the end of the

pregnancy.

Cord blood is obtained during

delivery and thus provides

information on the last days

before it.

Neonatal urine is obtained the first

day(s) of life and can account

for the previous 60-72 hours.

As previously

reported, meconium

and fetal hair

objectively assess a

chronic exposure to

drugs during fetal life,

starting from second

trimester of pregnancy

in case of meconium

whereas hair grows

only during the third

trimester of

pregnancy.

FAEEs in neonatal hair

Caprara DL, Klein J, Koren G.

Baseline measures of fatty acid etil esters in hair of

neonates born to abstaining or mild social drinking

mothers.

Ther Drug Monit. 2005 Dec;27(6):811-5.

hair samples of infants born to women with problems of

excessive alcohol consumption should be analyzed to

check the possible difference in the concentrations of

FAEEs

AN IMPORTANT LIMITATION IS THE AMOUNT OF

NEONATAL HAIR WHICH CAN BE COLLECTED (< 10

mg)

EtG and EtS in neonatal hair?

All hair samples from newborns tested < LLOQ for HEtG.

Morini L, Marchei E, Vagnarelli F, Garcia Algar O, Groppi A, Mastrobattista L, Pichini S.

Ethyl glucuronide and ethyl sulfate in meconium and hair-potential biomarkers of intrauterine exposure to ethanol.

Forensic Sci Int. 2010 Mar 20;196(1-3):74-7. Epub 2010 Jan 8.

http://www.ncbi.nlm.nih.gov/pubmed/16306859?ordinalpos=8&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSumhttp://www.ncbi.nlm.nih.gov/pubmed/16306859?ordinalpos=8&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSumhttp://www.ncbi.nlm.nih.gov/pubmed/16306859?ordinalpos=8&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSumhttp://www.ncbi.nlm.nih.gov/pubmed/16306859?ordinalpos=8&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSumhttp://www.ncbi.nlm.nih.gov/pubmed/16306859?ordinalpos=8&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSumhttp://www.ncbi.nlm.nih.gov/pubmed/16306859?ordinalpos=8&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSumhttp://www.ncbi.nlm.nih.gov/pubmed/16306859?ordinalpos=8&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSumhttp://www.ncbi.nlm.nih.gov/pubmed/20060246http://www.ncbi.nlm.nih.gov/pubmed/20060246http://www.ncbi.nlm.nih.gov/pubmed/20060246http://www.ncbi.nlm.nih.gov/pubmed/20060246http://www.ncbi.nlm.nih.gov/pubmed/20060246http://www.ncbi.nlm.nih.gov/pubmed/20060246

The first bowel movement a baby has is called meconium.

Meconium is composed of amniotic fluid, mucus, lanugo

(the fine hair that covers the baby's body), bile, and cells

that have been shed from the skin and the intestinal tract.

Meconium is thick, greenish black, sticky and sterile.

The Meconium Project: An Italian-Spanish

Joint Study to Assess Exposure to Illicit Drugs

During Pregnancy and Birth Outcomes in a

mediterranean city (2004-2008)

Istituto Superiore di Sanit, Roma, Italy

IMIM- Hospital del Mar, Barcelona, Spain

We found in 1209 meconium samples of mother-infant dyads attending the

Hospital during 2002-2004 a prevalence of prenatal exposure to 2.6%

Cocaine, 4.7% heroin and 5.3% cannabis and.

45% daily maternal ethanol measured by FAEEs in

meconium

Chan et al., JPET 2004

Fatty acid ethyl esters (FAEEs) in meconium

Positive cut-off:

7 FAEEs> 2 nmol/g meconium

Are there EtG + EtS in meconium?

2007-2009

MECONIUM SAMPLES OBTAINED FROM

Similar sociodemographic (Urban) and ethnic (Caucasians with 40%

immigrants from North Africa, South America and Asia)

characteristics

BARCELONA, SPAIN

BCN

REGGIO EMILIA, ITALY

RE

Meconium

Samples Statistics

EtG

(nmol/g) EtS

(nmol/g)

FAEEs > 2

nmol/g

(n=44)

Mean 2.04 0.14

SD 3.92 0.61

Min 0.11 0.01

Max 21.94 3.47

Median 0.21 0.02

FAEEs < 2

nmol/g

(n=133)

Mean 0.63 0.02

SD 1.19 0.03

Min 0.02 1.01

Max 7.11 0.2

Median 0.06 0.01

P value* 0.0002 0.009

* In relation to samples FAEE >2

EtS

Measurable

in 52%

samples

EtG

Measurable

in 84%

samples

Comparison of EtG and EtS levels in meconium

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3

4

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