Elimination kinetics of domoic acid from the brain and cerebrospinal fluid of the pregnant rat.

Domoic acid (DA) causes neurological effects in multiple species upon exposure, including status epilepticus in pregnant sea lions and an epileptic disease state that commonly develops in juveniles. This study aims to define brain toxicokinetic parameters in the pregnant rat in the larger context of maternal-fetal toxin transfer. Specifically, Sprague-Dawley rats were exposed to a low observable effect level of 1.0 mg DA/kg intravenously at gestational day 20, and plasma, brain, and cerebrospinal fluid (CSF) samples were taken at discrete time points over 24 h. Domoic acid concentrations were determined by a tandem LC/MS method recently optimized for brain tissue and CSF. Data showed that 6.6% of plasma DA reached the brain, 5.3% reached the CSF, and DA levels were nearly identical in both brain and CSF for 12 h, remaining above the threshold to activate isolated hippocampal neurons for 2 h. The calculated terminal half-life of CSF was 4 h, consistent with the time for complete CSF regeneration, suggesting that CSF acts as a mechanism to clear DA from the brain.

Toxicokinetics of domoic acid in the fetal rat.

Domoic acid (DA) is a potent neurotoxin that has both marine wildlife and human health impacts, including developmental effects during prenatal exposure in rodent models. However, little is known regarding DA toxicokinetics in the fetal unit during maternal-fetal transfer. Tissue distribution and toxicokinetics of DA were investigated in pregnant rats and their pups just prior to birth at gestational day 20. Pregnant Sprague Dawley rats were given an intravenous dose of 1.0 mg DA/kg and samples of maternal plasma, fetal plasma, placenta, amniotic fluid and fetal brain were taken at intervals over 24 h. Toxicokinetic parameters were determined using WinNonLin software analysis. Maternal plasma DA log concentration-time curves fit a two compartment pharmacokinetic profile, with alpha and beta half-lives of elimination of 26.9 and 297 min, respectively. Placenta had a C(max) of 752 ng/mL and a terminal half-life of 577 min. Maternal-fetal transfer between the plasma compartments was 31% with a fetal plasma C(max) of 86 ng/mL at 60 min and terminal half-life of 553 min. Amniotic fluid and fetal brain had overall averages of 27±12 ng/mL and 8.12 ng/g, respectively, and did not show evidence of elimination over 24 h. The longer fetal retention of DA, particularly in amniotic fluid, indicates that the fetus may be continually re-exposed during gestation, which could potentially lead to a disease state even at small exposure dose. This has implications for the California sea lions (Zalophus californianus), which exhibit an epilepsy-like disease that arises months after DA producing blooms.

Optimization of solid-phase extraction and liquid chromatography-tandem mass spectrometry for the determination of domoic acid in seawater, phytoplankton, and mammalian fluids and tissues.

We previously reported a solid-phase extraction (SPE) method for determination of the neurotoxin domoic acid (DA) in both seawater and phytoplankton by liquid chromatography-tandem mass spectrometry (LC-MS/MS) with the purpose of sample desalting without DA pre-concentration. In the present study, we optimized the SPE procedure with seawater and phytoplankton samples directly acidified with aqueous formic acid without addition of organic solvents, which allowed sample desalting and also 20-fold pre-concentration of DA in seawater and phytoplankton samples. In order to reduce MS contamination, a diverter valve was installed between LC and MS to send the LC eluant to waste, except for the 6-min elution window bracketing the DA retention time, which was sent to the MS. Reduction of the MS turbo gas temperature also helped to maintain the long-term stability of MS signal. Recoveries exceeded 90% for the DA-negative seawater and the DA-positive cultured phytoplankton samples spiked with DA. The SPE method for DA extraction and sample clean-up in seawater was extended to mammalian fluids and tissues with modification in order to accommodate the fluid samples with limited available volumes and the tissue extracts in aqueous methanol. Recoveries of DA from DA-exposed laboratory mammalian samples (amniotic fluid, cerebrospinal fluid, plasma, placenta, and brain) were above 85%. Recoveries of DA from samples (urine, feces, intestinal contents, and gastric contents) collected from field stranded marine mammals showed large variations and were affected by the sample status. The optimized SPE-LC-MS method allows determination of DA at trace levels (low pg mL(-1)) in seawater with/without the presence of phytoplankton. The application of SPE clean-up to mammalian fluids and tissue extracts greatly reduced the LC column degradation and MS contamination, which allowed routine screening of marine mammalian samples for confirmation of DA exposure and determination of fluid and tissue DA concentrations in experimental laboratory animals.