LC-MS/MS analysis of acetaminophen and caffeine in amniotic fluid

The intake of several substances by pregnant women could be
hazardous to the fetus and mother's health: many substances can
cross the placenta and reach the fetal compartment, causing adverse
outcomes. Consequently, to accurately measure the presence of
xenobiotics in fetal matrices, sensitive and specific bioanalytical
methods are necessary: this would allow the assessment of fetal
exposure to substanceswhich, although licit, can be dangerous for the
fetal and child's growth. The aim of this study was to develop and
validate a liquid chromatography tandem mass spectrometry method
for the simultaneous determination and quantitation of caffeine and
acetaminophen in amniotic fluid. Amniotic fluid is a quite complex
biological matrix and, as such, it requires a purification step prior to
analysis. The extraction method has been optimized by comparing
three different commercially available SPE cartridges (Supel™ Select
HLB, Phenomenex Strata C18-E, and Agilent ABS Elut-NEXUS), and a
liquid/liquid extraction with acetonitrile. A reverse-phase HPLC with a
C18 column and gradient elution programwas used. MS detectionwas
carried out in MRM mode. Quantitation was performed using the
internal standard method. Validation parameters were very satisfactory.
The high selectivity and sensitivity of the method (LOQ < 9.5 ng
mL1, and LOD < 3.3 ngmL1) allowed us to determine target analytes
even in small amounts. Precision, matrix effect, and stability were also
evaluated. The whole validated method has finally been applied to the
analysis of 194 real samples of human amniotic fluid obtained from
pregnant women (15–21 weeks of gestation) in order to monitor the
effective intake of target analytes: 96% of the examined women
consumed caffeine during pregnancy while a lower percentage (20%)
showed acetaminophen intake. The whole procedure is simple and
easy to perform with minimal sample preparation and short analysis
time.