Biocompatible SPME fibers for direct monitoring of nicotine and its metabolites at ultra trace concentration in rabbit plasma following the application of smoking cessation formulations.

The ultra-trace determination of nicotine and its 4 major metabolites (cotinine, nornicotine, norcotinine and anabasine) from rabbit plasma was achieved by a newly developed solid phase microextraction-liquid chromatography-tandem mass spectrometry method. Extraction of the target analytes was performed with hydrophilic/lipophilic balance-polyacrylonitrile SPME fibers. Dual fiber extraction was necessary to guarantee improved recovery at parts-per-trillion levels. Liquid chromatographic analysis was achieved in a 6-min run using a C18 (1.9 µm C18, 50 mm x 2.1 mm) column with a mobile phase flow rate of 0.4 mL/min. Tandem mass spectrometry was used for detection and quantification in positive electrospray ionization (ESI+) mode for all the targeted analytes. Two stable isotope-labeled internal standards were used for signal correction and accurate quantification. The mass spectrometer with laminar flow ion flux transport, guaranteed improved signal stability, minimal contamination of the ion guide and reproducibility into the first quadrupole analyzer. The method was validated in line with the Food and Drug Administration (FDA) guidelines for bioanalytical method validation. The results met the acceptance criteria as proposed by the FDA: accuracy was tested at 0.35, 10 and 75 µg L - 1 and ranged between 98.3-112.2% for nicotine, 94.1-101.9% for cotinine, 94.7-107.0% for nornicotine, 81.1-107.2% for norcotinine and 94.3-115.2% for anabasine, with precision up to 14.2%. Stability tests indicated that all the targeted analytes were stable in the desorption solution for at least 1 week. LOQs ranged from 0.05 to 1 µg L-1. The method was successfully applied to analyze plasma samples obtained from rabbits following transdermal application of a smoking cessation formulation loaded with solid lipid nanoparticles containing a nicotine-stearic acid conjugate.

E-cigarette liquids: Constancy of content across batches and accuracy of labeling.

AIMS: To assess whether bottles of refill liquids for e-cigarettes were filled true to label, whether their content was constant across two production batches, and whether they contained impurities. METHODS: In 2013, we purchased on the Internet 18 models from 11 brands of e-liquids. We purchased a second sample of the same models 4months later. We analyzed their content in nicotine, anabasine, propylene glycol, glycerol, ethylene glycol and diethylene glycol, and tested their pH. RESULTS: The median difference between the nicotine value on the labels and the nicotine content in the bottles was 0.3mg/mL (range -5.4 to +3.5mg/mL, i.e. -8% to +30%). For 82% of the samples, the actual nicotine content was within 10% of the value on the labels. All models contained glycerol (median 407mg/mL), and all but three models contained propylene glycol (median 650mg/mL). For all samples, levels of anabasine, ethylene glycol and diethylene glycol were below our limits of detection. The pH of all the e-liquids was alkaline (median pH=9.1; range 8.1 to 9.9). The measured content of two batches of the same model varied by a median of 0% across batches for propylene glycol, 1% for glycerol, 0% for pH, and 0.5% for nicotine (range -15% to +21%; 5th and 95th percentiles: -15% and +10%). CONCLUSIONS: The nicotine content of these e-liquids matched the labels on the bottles, and was relatively constant across production batches. The content of propylene glycol and glycerol was also stable across batches, as was the pH.

Trace level determination of pyrethroid and neonicotinoid insecticides in beebread using acetonitrile-based extraction followed by analysis with ultra-high-performance liquid chromatography-tandem mass spectrometry.

Beebread is among the matrices suspected of contaminating honeybee. To better understand this contamination, the study aimed to develop an efficient, sensitive and reliable analytical method for the trace analysis of pesticides in beebread. This study focuses specifically on the insecticides pyrethroids and neonicotinoids and some of their metabolites. It describes the development and validation of an original analytical approach that consists of one simple extraction method based on modified QuEChERS followed by a selective and sensitive analysis by UHPLC-MS/MS to determine the target compounds in beebread. The method was validated using a quadratic fit. RSD values below 20% were obtained, except for 5-hydroxy-imidacloprid and imidacloprid at 0.5 ng/g, which exhibited RSDs of 25% and 21%, respectively. The intra-day precision was less than 10% for many of the investigated compounds. The inter-day precision varied between 2% and 36%, depending on the compound and the concentration. The recoveries varied from 53% to 119%, with averages of 83, 81 and 77% for the extraction of beebread samples spiked at 0.5, 5 and 10 ng/g, respectively. The LOD values for all the substances were below ng/g, with the exception of 6-chloronicotinic acid (LOD=1.7 ng/g). The method was then applied to the analysis of 32 beebread samples and revealed the presence of 7 of the target substances. The most frequently detected pesticides belonged to the neonicotinoid family and were generally present at low concentrations, but in some cases exceeded 170 ng/g (acetamiprid and thiacloprid). Some pyrethroids were also detected (lambda-cyhalothrine and bifenthrine), but at very low levels.

Aromatase inhibitors in cigarette smoke, tobacco leaves and other plants.

A chance observation that cigarette smoke interferes with the aromatase assay led us to investigate tobacco leaf and smoke extracts for the presence of aromatase inhibitors. The highest inhibitory activity was found in the basic fraction of cigarette smoke. Further purification of this fraction led to the identification of N-n-octanoylnornicotine. Synthesis and testing of a series of acylated nornicotines and anabasines for their ability to inhibit aromatase showed an interesting correlation of activity with the length of the acyl carbon chain, with maximum activity at C-11. The acylated derivatives showed activity which was significantly greater than that of nicotine and anabasine. In vivo studies in rats indicated that administration of this inhibitor delayed the onset of NMU-induced breast carcinoma and altered the estrus cycle. These in vivo studies suggest that tobacco alkaloid derivatives exert their effects by suppression of the aromatase enzyme system. Toxicity studies indicated relatively low toxicity with LD50 for N-n-octanoylnornicotine = 367 mg/kg body weight. When extracts from thirty five varieties of vegetables, plant leaves, and fruits were analyzed, seventeen showed quantitatively significant aromatase inhibition which was comparable to that of green tobacco leaf, suggesting that naturally occurring substances may affect endocrine function through aromatase inhibition.

Teratogenicity in swine of the tobacco alkaloid anabasine isolated from Nicotiana glauca.

Nicotiana glauca, wild tree tobacco, induces arthrogrypotic congenital defects in piglets similar to those induced by Nicotiana tabacum, common tobacco. The present work was conducted to isolate the principal alkaloid of N. glauca, anabasine, in large quantity and good purity and to test the teratogenicity of the compound in pigs. The isolated compound was established to be anabasine and to be of suitable purity by chemical characterization. It proved to be teratogenic. Typical arthrogrypotic defects were induced in 21 of 26 offspring (three of three litters) when dams ingested 2.6 mg of the compound per kg body weight twice daily during the 43rd-53rd days of gestation. Of three dams dosed with 1.66 g/kg/day of the dried plant material during the 43rd-53rd days, one delivered deformed offspring representing one-third of all offspring in that group. These arthrogrypotic defects induced by anabasine were indistinguishable clinically from defects induced by either N. glauca or N. tabacum. In addition, anabasine at a dose of 2.6 mg/kg twice daily or N. glauca plant material at 1.66 gm/kg daily induced cleft palate in over three-fourths of offspring (100% of litters) when dams ingested either during the 30th-37th days of gestation or during longer periods that included those days.