S1 certification of alpha-endosulfan, beta-endosulfan, and endosulfan sulfate in a candidate certified reference material (organochlorine pesticides in tea) by isotope dilution gas chromatography-mass spectrometry.

This paper presents the certification of alpha-endosulfan, beta-endosulfan, and endosulfan sulfate in a candidate tea certified reference material (code: GLHK-11-03) according to the requirements of the ISO Guide 30 series. Certification of GLHK-11-03 was based on an analytical method purposely developed for the accurate measurement of the mass fraction of the target analytes in the material. An isotope dilution mass spectrometry (IDMS) method involving determination by (i) gas chromatography-negative chemical ionization-mass spectrometry (GC-NCI-MS) and (ii) gas chromatography-electron ionization-high-resolution mass spectrometry (GC-EI-HRMS) techniques was employed. The performance of the described method was demonstrated through participation in the key comparison CCQM-K95 "Mid-Polarity Analytes in Food Matrix: Mid-Polarity Pesticides in Tea" organized by the Consultative Committee for Amount of Substance-Metrology in Chemistry in 2012, where the study material was the same as the certified reference material (CRM). The values reported by using the developed method were in good agreement with the key comparison reference value (KCRV) assigned for beta-endosulfan (727 ± 14 µg kg(-1)) and endosulfan sulfate (505 ± 11 µg kg(-1)), where the degree of equivalence (DoE) values were 0.41 and 0.40, respectively. The certified values of alpha-endosulfan, beta-endosulfan, and endosulfan sulfate in dry mass fraction in GLHK-11-03 were 350, 730, and 502 µg kg(-1), respectively, and the respective expanded uncertainties, due to sample inhomogeneity, long-term and short-term stability, and variability in the characterization procedure, were 27 µg kg(-1) (7.8 %), 48 µg kg(-1) (6.6 %), and 33 µg kg(-1) (6.6 %).

Determination of Aconitum alkaloids in dietary supplements and raw botanical materials by liquid chromatography/UV detection with confirmation by liquid chromatography/tandem mass spectrometry: collaborative study.

An interlaboratory study was conducted to evaluate a method for the determination of 3 Aconitum alkaloids, viz., aconitine, mesaconitine, and hypaconitine, in raw botanical material and dietary supplements. The alkaloids were extracted with diethyl ether in the presence of ammonia. After cleanup by solid-phase extraction to remove matrix interferences, the alkaloids were determined by reversed-phase liquid chromatography (LC)/UV detection at 235 nm with confirmation by LC/tandem mass spectrometry (MS/MS). A total of 14 blind duplicates were successfully analyzed by 12 collaborators. For repeatability, the relative standard deviation (RSDr) values ranged from 1.9 to 16.7%, and for reproducibility, the RSDR values ranged from 6.5 to 33%. The HorRat values were all <2 with only one exception at 2.3. All collaborating laboratories had calibration curves with correlation coefficients of >0.998. In addition, 6 collaborators performed the confirmation and were able to verify the identities of the alkaloids by using LC/MS/MS.

Preparation of reference material for organochlorine pesticides in a herbal matrix.

The development of reference material for four organochlorine pesticides, namely hexachlorobenzene and three isomers of hexachlorocyclohexane (alpha-hexachlorocyclohexane, beta-hexachlorocyclohexane and gamma-hexachlorocyclohexane), in a ginseng root sample is presented. Raw materials (Panax ginseng) were purchased from a local market and confirmed to contain certain levels of incurred organochlorine pesticide residues by a validated gas chromatography-mass selective detection method. A total of more than 300 bottles each containing 25 g of samples were prepared after the materials had been freeze-dried, milled and thoroughly mixed. The homogeneity and stability of samples from randomly selected bottles were verified and the reference values were characterized using a highly precise isotope dilution gas chromatography-mass spectrometry (ID-GCMS) method that was recently developed by our laboratory. The purity of standard organochlorine chemicals was determined against certified reference materials to establish the accuracy of the ID-GCMS analysis. The concentrations (+/- expanded uncertainty) of hexachlorobenzene, alpha-hexachlorocyclohexane, beta-hexachlorocyclohexane and gamma-hexachlorocyclohexane in the reference material were 0.198 +/- 0.015, 0.450 +/- 0.022, 0.213 +/- 0.011 and 0.370 +/- 0.032 mg kg(-1), respectively. A portion (70 bottles) of the samples was also used in a proficiency testing (PT) scheme for assessing the testing capabilities of field laboratories. The consensus mean values of the PT obtained from the 70 participants were on the same order but deviated by -2.7 to -14.1% from those of the assigned reference values. Because of the wide spread of participants' data (relative standard deviation ranging from 44 to 56%), the PT results were not included in the calculation of the assigned values of the reference materials. The materials served as suitable reference materials to ascertain the quality control and validation processes for the determination of organochlorine pesticides in herbal matrices.

Interlaboratory comparison for the determination of five residual organochlorine pesticides in ginseng root samples by gas chromatography.

An interlaboratory comparison study for the determination of 5 residual organochlorine pesticides (hexachlorobenzene and 4 hexachlorocyclohexane isomers) in ginseng root was performed. This program [Asia Pacific Laboratory Accreditation Cooperation (APLAC) T049] was the first of its kind for an herbal matrix and involved the participation of 70 laboratories from 26 countries worldwide. Consensus mean values were computed statistically from the reported results, which were eventually used to assess the performance of individual laboratories in terms of the z-scores. The distribution of analytical data was found to be widespread, with standard deviation ranging from 43.9 to 55.9%, and the result patterns obtained were similar to those residue pesticide programs using other matrixes. Although the estimation of measurement uncertainty is a crucial requirement for all quantitative tests for laboratories that meet the requirements of International Organization for Standardization/International Electrotechnical Commisssion (ISO/IEC) 17025, some laboratories in this program had difficulties and showed unfamiliarity with respect to that quality criterion. It was recommended that laboratories review and rectify the situation promptly so that they would have a better understanding of measurement uncertainty or the test service provided.

Application of isotope dilution gas chromatography-mass spectrometry in analysis of organochlorine pesticide residues in ginseng root.

A highly accurate and precise method based on isotope dilution gas chromatography-mass spectrometry was developed for the determination of five matrix-bound organochlorine pesticides, namely, hexachlorobenzene and hexachlorocyclohexanes (alpha-, beta-, delta-, and gamma- isomers), in a reference sample of Panax gingseng. Identification of the analytes was confirmed under selective ion monitoring mode by the presence of two dominant ion fragments within the specific time windows (+/-1% of the relative retention time with respect to the calibration standards) and matching of relative ion intensities of the concerned ions in samples and calibration standards (within +/-5%). Quantification was based on the measurement of concentration ratios of the natural and isotopic analogues in the sample and calibration blends. To circumvent the tedious iterative process of exact isotope matching that is often used in isotope dilution mass spectrometry analysis, a single-point calibration procedure was adopted with the isotopic amount ratios in the sample and calibration blends close to unity (0.9-1.1). Under the described approach, intraday and interday repeatability of replicate analyses of organochlorine pesticides in the ginseng root sample were below 1.4%. The expanded relative uncertainty ranging from 4.0 to 6.5% at a coverage factor of 2 was significantly lower than those of conventional gas chromatographic methods using other calibration techniques (internal or external standards). A deviation of less than 2.0% from the certified values was achieved when applying the developed method to determine hexachlorobenzene, alpha-, and beta-hexachlorocyclohexane in a certified reference material (CRM), BCR-CRM 115. Because of the unavailability of relevant CRMs of herbal origins, the concerned ginseng root sample, after verification of the "true values" of the concerned organochlorine pesticides by the valid primary method, is suitable for serving as an in-house reference material for quality assurance and method validation purposes.

Identification and characterisation of the chinese herb Langdu by LC-MS/MS analysis.

An LC-MS/MS method has been developed for the identification of three species of herb used as the traditional Chinese medicine Langdu, namely Stellera chamaejasme L., Euphorbia ebracteolata Hayata and E. fischeriana Steud. As these herbs contain different mixtures of marker compounds, they could be unambiguously differentiated from each other by comparing their respective characteristic segmental multiple reaction monitoring profiles. The profiles indicated that S. chamaejasme contained daphnetin, skimmetine, stellerin, chamaechromone and neochamaejasmin, E. fischeriana contained ebracteolata compound B, ingenol, jolkinolide B and fischeriana A, whilst E. ebracteolata contained ebracteolata compounds B and C along with ingenol. These results were confirmed from the respective MS/MS spectra. The method has been successfully applied to differentiate these herbs from the related species Alocasia macrorrhiza (L.) Schott and E. kansui Liou.

Determination of ginsenosides (ginseng saponins) in dry root powder from Panax ginseng, Panax quinquefolius, and selected commercial products by liquid chromatography: interlaboratory study.

Twelve collaborating laboratories assayed 4 products, namely, Panax ginseng, Panax quinquefolius, and 2 ginseng products, for 6 ginsenosides: Rb1, Rb2, Rc, Rd, Re, and Rg1. Collaborators also received a negative control for the recovery study. Pure ginsenosides were provided as reference standards for the liquid chromatography (LC) analysis and the system suitability tests. The LC analyses were performed on the methanol extract using UV detection at 203 nm. For P. ginseng, individual ginsenosides were consistent in their means; repeatability standard deviations (RSDr) ranged from 4.17 to 5.09% and reproducibility standard deviations (RSDR) ranged from 7.27 to 11.3%. For P. quinquefolius, the Rb1 and Rb2 ginsenosides were higher and lower in concentration than P. ginseng, with RSDr values of 3.44 and 6.60% and RSDR values of 5.91 and 12.6% respectively, and other analytes at intermediate precisions. For ginseng commercial products, RSDr values ranged from 3.39 to 8.12%, and RSDR values ranged from 7.65 to 16.5%. A recovery study was also conducted for 3 ginsenosides: Rg1, Re, and Rb1. The average recoveries were 99.9, 96.2, and 92.3%, respectively. The method is not applicable for the determination of Rg1 and Re in ginseng product at levels <300 mg/kg.

Analysis of proprietary Chinese medicines for the presence of toxic ingredients by LC/MS/MS.

This paper presents an LC/MS/MS approach for simultaneous qualitative and quantitative analysis of proprietary Chinese medicine products for the presence of toxic ingredient compounds. The target compounds include three C(19)-diterpenoid alkaloids, two quinolizidine alkaloids, two indole alkaloids and four bufadienolide steroids. They were recognized as active compounds of several toxic/potent herbal materials commonly used in the preparation of some proprietary medicine products. These toxic/potent herbal materials include Radix Aconiti Lateralis, Radix Sophorae Tonkinensis, Semen Strychni and Venenum Bufonis. In the analysis, the LC/MS/MS system was set to record the MS spectra and respective multiple reaction monitoring (MRM) signals for different target compounds after they were separated and eluted from the column. The MS spectra were used for the qualitative analysis whereas the MRM signals for the quantitative analysis. In this study, totally 12 proprietary medicine products were tested and found to contain some of the target compounds at different levels.