oleandrin and bufalin

The microparticle enzyme immunoassay (MEIA for digoxin (Abbott Laboratories, Abbott Park, Ill) requires no sample pretreatment and is widely used in clinical toxicology laboratories for monitoring serum digoxin concentrations. One advantage of the new MEIA is the lower cross-reactivities with such cross-reactants as digitoxin, oleandrin, and bufalin compared with the fluorescence polarization immunoassay (FPIA)for digoxin. Digitoxin, oleandrin, and bufalin showed positive cross-reactivity with MEIA and FPIAs for digoxin in the absence of the primary analyte, digoxin. A surprising finding was that digoxin concentrations were falsely decreased by these cross-reactants when serum pools containing digoxin were supplemented with various concentrations of these cross-reactants and when digoxin concentrations were measured by the MEIA. In contrast, digoxin concentrations were falsely elevated when measured by the FPIA. For example, when a serum pool containing 2.15 nmol/L of digoxin was supplemented with 129.5 nmol/L of bufalin, the apparent digoxin concentrations were 1.45 nmol/L with the MEIA and 3.00 nmol/L with the FPIA. Taking the advantage of only 25% protein binding of digoxin and more than 95% protein binding of digitoxin and bufalin, we demonstrated that monitoring free digoxin instead of total digoxin eliminated negative interference of digoxin by these cross-reactants in the MEIA and positive interference in the FPIA. Although oleandrin is also strongly bound to serum protein, high concentrations of oleandrin still modestly affect the free digoxin assay for both MEIA and FPIAs.

Topics: Bufanolides; Cardenolides; Cardiac Glycosides; Cardiotonic Agents; Cross Reactions; Digitoxin; Digoxin; False Positive Reactions; Fluorescence Polarization Immunoassay; Humans; Immunoenzyme Techniques; Microspheres

Oleandrin plant poisoning is common in children and the plant extract is used in Chinese medicines. The toxicity is due to oleandrin and the deglycosylated metabolite oleandrigenin. Bufalin and cinobufotalin (toad cardiac toxins) are also widely used in Chinese medicines like Chan SU, and Lu-Shen -WU. Severe toxicity from bufalin after consumption of toad soup has been reported. Taking advantage of structural similarities of these toxins with digitoxin, we demonstrated that these compounds can be rapidly detected in blood by the fluorescence polarization immunoassay for digitoxin. The cross reactivities of these compounds with digoxin assay were much lower. For example, when a drug free serum was supplemented with 10 microg/ml of oleandrin, we observed 127.7 ng/ml of digitoxin equivalent but only 2.4 ng/ml of digoxin equivalent concentration. Digibind neutralized all cardiac toxins studied as evidenced by significant fall of free concentrations. When aliquots of serum pool containing 50.0 microg/ml of oleandrin were supplemented with 0, 10.0, 25.0, 50.0, 100, and 200 microg/ml of digibind, the mean free concentrations were 30.6, 23.3, 16.0, 10.7, 7.8 and 5.5 microg/ml respectively. Similarly, with 50.0 microg/ml of oleandrigenin (total concentration: 36.2 ng/ml), the free concentration was 14.5 ng/ml digitoxin equivalent in the absence of digibind and 5.4 ng/ml in the presence of 200 microg/ml of digibind. In another specimen containing 500 ng/ml bufalin (total concentration: 156.9 ng/ml), the free concentration was 8.6 ng/ml in the absence of digibind and none detected in the presence of 100.0 microg/ml digibind. Because such neutralization may also occur in vivo, digibind may be useful in treating patients exposed to these toxins.

Topics: Bufanolides; Cardenolides; Cardiotonic Agents; Chromatography, High Pressure Liquid; Cross Reactions; Digitoxin; Digoxin; Humans; Immunoassay; Immunoglobulin Fab Fragments; Mass Spectrometry; Neutralization Tests