ABSTRACT
An enzyme-linked immunosorbent assay is reported for monitoring thevetin, an active constituent of the highly poisonous plant Thevetia nerifolia. A thevetin-BSA conjugate was employed as the immunogen and the antibodies raised in rabbits were used for the development of an ELISA. Penicillinase served as the marker enzyme and its conjugation to thevetin by the periodate method is reported for the first time. The present ELISA method could detect 2 ng/ml of thevetin. Cross-reactivity studies with structural analogues and other phytotoxins and drugs of common occurrence in clinical and forensic toxicology established the superiority of the ELISA over the existing analytical methods for determining thevetin in various biospecimens.
Subject(s)
Cardenolides/analysis , Enzyme-Linked Immunosorbent Assay/methods , Animals , Antibody Specificity , Cardenolides/chemistry , Cardenolides/immunology , Cardenolides/isolation & purification , Cross Reactions , Evaluation Studies as Topic , Glycosides/chemistry , Glycosides/immunology , Haptens , Hormones/chemistry , Hormones/immunology , Penicillinase , Pharmaceutical Preparations/chemistry , Plants, Toxic/chemistry , Rabbits , Reproducibility of Results , Seeds/chemistry , Sensitivity and SpecificityABSTRACT
Cleistanthus collinus is a highly toxic plant frequently implicated in suicidal and homicidal poisoning cases referred to our laboratory. An enzyme-linked immunosorbent assay (ELISA) is reported in this paper for the quantitation of the active principles of C. collinus. This method is highly reproducible and sensitive to detect as low as 2 ng/ml of cleistanthin B, a toxic constituent of C. collinus. The ELISA can be successfully applied for the specific identification of C. collinus poisoning in clinical and forensic toxicology.
Subject(s)
Enzyme-Linked Immunosorbent Assay/methods , Glucosides/toxicity , Plant Poisoning , Animals , Immunotoxins , RabbitsABSTRACT
A method is reported for the solid-state fluorodensitometric determination of haptens in hapten-protein conjugates. The applicability of the method is shown using the bovine serum albumin conjugate of cleistanthin B, a toxic glycoside of the plant Cleistanthus collinus. This non-destructive technique, which is relatively simple, sensitive, rapid and versatile, can be used for the determination of protein conjugates of fluorogenic haptens.
Subject(s)
Glycosides/metabolism , Haptens/metabolism , Lignans , Plants, Toxic/metabolism , Serum Albumin, Bovine/metabolism , Chromatography, Thin Layer , Spectrometry, Fluorescence , Spectrophotometry, UltravioletABSTRACT
Enzyme-linked immunosorbent assay is reported for the estimation of cleistanthin A, a major constituent of the toxic plant Cleistanthus collinus. Rabbit antibodies were obtained by immunisation with cleistanthin A hemisuccinate-BSA conjugate and the ELISA developed thereupon could detect cleistanthin A at as low a concentration as 3 ng/ml. Cross-reactivity studies with structural analogs as well as with other phytotoxins and drugs of common occurrence established the suitability of the ELISA to specifically monitor the C. collinus marker molecules in emergency clinical and forensic cases. The simplicity and specificity make the ELISA superior to the other available techniques.
Subject(s)
Enzyme-Linked Immunosorbent Assay/methods , Glycosides/analysis , Lignans , Cross Reactions , Enzyme-Linked Immunosorbent Assay/statistics & numerical data , Evaluation Studies as Topic , Humans , Plant Poisoning/diagnosis , Plants, Toxic , Sensitivity and Specificity , Toxins, Biological/analysisABSTRACT
Spectrofluorometric quantitation of a lignan lactone, diphyllin, and its glycosides cleistanthin A and cleistanthin B, the active principles of the poisonous plant Cleistanthus collinus, is reported for the first time. Thin layer chromatographic resolution of the components precedes the fluorometric measurements. The method is highly reproducible and sensitive to 0.1 microgram/mL for diphyllin and to 1 microgram/mL for cleistanthins A and B in ethanol. The technique is employed for the assay of several biospecimens of forensic and clinical importance.