Optimization and Validation of a Multiresidue Method for Screening of 126 Pesticide Residues in Herbal Raw Materials and Extracts Used as Ingredients in Ayurvedic Medicines and Dietary Supplements.

BACKGROUND: Testing for pesticide levels in herbal products is an important aspect in determining product safety. Plants and their extracts are widely used as ingredients in botanical dietary supplements and traditional medicines. The extracts of plants, especially those prepared out of organic solvents, are rich in secondary metabolites and pigments, and adequate clean-up is required since the extracts completely dissolve in organic solvents. OBJECTIVE: The study aims at reporting a multiresidue analytical method for 126 different pesticides in raw material biomass as well as extracts of plants, which are widely used as ingredients in ayurvedic medicines as well as dietary supplements using LC-MS/MS and GC-MS/MS with a rugged sample preparation technique for accurate results. METHOD: QuEChERS (quick, easy, cheap,effective, rugged, and safe) procedure, gel permeation chromatography (GPC), GPC coupled with solid phase extraction (SPE), and liquid-liquid extraction (LLE) coupled with SPE sample preparation methods were compared against each other for suitability to test pesticides in selected herbal raw materials and their alcoholic and aqueous extracts. The standard addition method was used for quantifying the level of pesticides below 10 µg/kg. RESULTS: Single laboratory validation for sample preparation involving GPC and SPE resulted linearity in the range of 2.5-500 ng/mL, average intraday and interday precision of 6.6% RSD, and average recovery (spiked at 10 µg/kg) of 92% for all analytes tested. The method was repeatable with different analysts and days. CONCLUSIONS: The sample preparation technique combining GPC and SPE as well as LLE and SPE was the most suitable for the selected herbal alcoholic extracts, whereas any of the regular techniques involving LLE, SPE, and QuEChERS were suitable for raw material biomass as well as aqueous extracts. HIGHLIGHTS: The method was found to be capable of determining selected pesticides in the selected matrixes at 10 µg/kg concentration. Provision of recycling solvents used in the GPC+SPE method was adopted to make the method environmentally friendly.

Determination of Withanolides in Withania somnifera by Liquid Chromatography: Single-Laboratory Validation, First Action 2015.17.

An LC method was developed and validated in 2007 for analyzing Withania somnifera raw material (root) and dried extracts for withanolide content, including withanoside IV, withanoside V, withaferin A, 12-deoxywithastromonolide, withanolide A, and withanolide B. The method involved the extraction of the analytes with methanol, their subsequent filtration, and then analysis on a C18 column with an acetonitrile gradient and UV detection. Single-laboratory validation yielded linearity generally in the range of 20 to 200 µg/mL for each analyte, with a repeatability precision of RSD < 3% in most cases, and recovery in the range of 90 to 105%. These results compare well with the performance criteria recently detailed in AOAC Standard Method Performance Requirement 2015.007. The method was shown to be rugged with respect to different analysts, equipment, and days of analysis, and the sample solution was shown to be stable for 24 h at room temperature after extraction. The method was reviewed by the AOAC Expert Review Panel on Dietary Supplements (Set 2 Ingredients) and approved for First Action Official MethodSM status.

Withania somnifera prevents morphine withdrawal-induced decrease in spine density in nucleus accumbens shell of rats: a confocal laser scanning microscopy study.

Opiate withdrawal is associated with morphological changes of dopamine neurons in the ventral tegmental area and with reduction of spine density of second-order dendrites of medium size spiny neurons in the nucleus accumbens shell but not core. Withania somnifera has long been used in the Middle East, Africa, and India as a remedy for different conditions and diseases and a growing body of evidence points to its beneficial effects on a number of experimental models of neurological disorders. Recently, many studies focused on the potential neuritic regeneration and synaptic reconstruction properties of its methanolic extract and its constituents (withanolides). This study investigates whether morphine withdrawal-induced spine reduction in the nucleus accumbens is affected by the administration of a Withania somnifera extract. To this end, rats were chronically treated with Withania somnifera extract along with morphine or saline and, upon spontaneous (1 and 3 days) or pharmacologically precipitated withdrawal, their brains were fixed in Golgi-Cox stain for confocal microscopic examination. In a separate group of animals, Withania somnifera extract was administered during three days of spontaneous withdrawal. Withania somnifera extract treatment reduced the severity of the withdrawal syndrome when given during chronic morphine but not during withdrawal. In addition, treatment with Withania somnifera extract during chronic morphine, but not during withdrawal, fully prevented the reduction of spine density in the nucleus accumbens shell in spontaneous and pharmacologically precipitated morphine withdrawal. These results indicate that pretreatment with Withania somnifera extract protects from the structural changes induced by morphine withdrawal potentially providing beneficial effects on the consequences related to this condition.