Innovative development and validation of an HPLC/DAD method for the qualitative and quantitative determination of major cannabinoids in cannabis plant material.

GC is commonly used for the analysis of cannabis samples, e.g. in forensic chemistry. However, as this method is based on heating of the sample, acidic forms of cannabinoids are decarboxylated into their neutral counterparts. Conversely, HPLC permits the determination of the original composition of plant cannabinoids by direct analysis. Several HPLC methods have been described in the literature, but most of them failed to separate efficiently all the cannabinoids or were not validated according to general guidelines. By use of an innovative methodology for modelling chromatographic responses, a simple and accurate HPLC/DAD method was developed for the quantification of major neutral and acidic cannabinoids present in cannabis plant material: Delta9-tetrahydrocannabinol (THC), THC acid (THCA), cannabidiol (CBD), CBD acid (CBDA), cannabigerol (CBG), CBG acid (CBGA) and cannabinol (CBN). Delta8-Tetrahydrocannabinol (Delta8-THC) was determined qualitatively. Following the practice of design of experiments, predictive multilinear models were developed and used in order to find optimal chromatographic analytical conditions. The method was validated following an approach using accuracy profiles based on beta-expectation tolerance intervals for the total error measurement, and assessing the measurements uncertainty. This analytical method can be used for diverse applications, e.g. plant phenotype determination, evaluation of psychoactive potency and control of material quality.

Can cocaine use be evaluated through analysis of wastewater? A nation-wide approach conducted in Belgium.

AIMS: Cocaine is the second most-used illicit drug world-wide and its consumption is increasing significantly, especially in western Europe. Until now, the annual prevalence has been estimated indirectly by means of interviews. A recently introduced and direct nation-wide approach based on measurements of the major urinary excreted metabolite of cocaine, benzoylecgonine, in wastewater is proposed. DESIGN: Wastewater samples from 41 wastewater treatment plants (WWTPs) in Belgium, covering approximately 3,700,000 residents, were collected. Each WWTP was sampled on Wednesdays and Sundays during two sampling campaigns in 2007-08. Samples were analysed for cocaine (COC) and its metabolites, benzoylecgonine (BE) and ecgonine methylester (EME) by a validated procedure based on liquid chromatography coupled with tandem mass spectrometry. Concentrations of BE were used to calculate cocaine consumption (g/day per 1000 inhabitants) for each WWTP region and for both sampling campaigns (g/year per 1000 inhabitants). FINDINGS: Weekend days showed significantly higher cocaine consumption compared with weekdays. The highest cocaine consumption was observed for WWTPs receiving wastewater from large cities, such as Antwerp, Brussels and Charleroi. Results were extrapolated for the total Belgian population and an estimation of a yearly prevalence of cocaine use was made based on various assumptions. An amount of 1.88 tonnes (t) per year [standard error (SE) 0.05 t] cocaine is consumed in Belgium, corresponding to a yearly prevalence of 0.80% (SE 0.02%) for the Belgian population aged 15-64 years. This result is in agreement with an earlier reported estimate of the Belgian prevalence of cocaine use conducted through socio-epidemiological studies (0.9% for people aged 15-64 years). CONCLUSIONS: Wastewater analysis is a promising tool to evaluate cocaine consumption at both local and national scale. This rapid and direct estimation of the prevalence of cocaine use in Belgium corresponds with socio-epidemiological data. However, the strategy needs to be refined further to allow a more exact calculation of cocaine consumption from concentrations of BE in wastewater.

Spatial and temporal variations in the occurrence of cocaine and benzoylecgonine in waste- and surface water from Belgium and removal during wastewater treatment.

Estimates of cocaine consumption are currently resulting from population surveys, consumer interviews and crime statistics. A new approach ("sewage epidemiology") based on the analysis of cocaine (COC) and its metabolite, benzoylecgonine (BE), in water samples was applied to 10 river sites and 30 wastewater treatment plants (WWTPs) in Belgium. Each river site was sampled twice, during the summer of 2007 and the winter of 2007-2008, while each WWTP was sampled on a Sunday and a Wednesday, during the summer-autumn of 2007 and the winter of 2007-2008. This sampling strategy allowed for the evaluation of spatial and seasonal variations in the occurrence of COC and BE in waste- and surface water. WWTP Brussel-Noord was sampled for 19 consecutive days to evaluate daily and weekly variations in the presence of COC and BE in wastewater. For 7 WWTPs, influent and effluent water samples were collected to investigate the removal of COC and BE during the wastewater treatment process. Analysis of water samples was performed using solid-phase extraction and liquid chromatography coupled to tandem mass spectrometry. Measured concentrations were further converted into an amount of used cocaine, called cocaine equivalents, as previously described in the literature. Results showed no significant difference in cocaine use between the investigated seasons. A constant cocaine consumption was observed during the week (Monday-Friday) with peaks during the weekend for WWTP Brussel-Noord. The COC/BE ratio in water samples was significantly higher during winter, most probably due to a slower hydrolysis of cocaine in low-temperature water. COC and BE were removed in the investigated WWTPs with a removal efficiency of >93%.

Cocaine and metabolites in waste and surface water across Belgium.

Cocaine abuse, a growing social problem, is currently estimated from population surveys, consumer interviews and crime statistics. A new approach based on the analysis of cocaine (COC) and metabolites, benzoylecgonine (BE) and ecgonine methyl ester (EME), in water samples was applied to 28 rivers and 37 waste water treatment plants in Belgium using solid-phase extraction and liquid chromatography coupled to tandem mass spectrometry. While EME was undetectable, COC and BE were detectable with concentrations ranging from <1 to 753 ng/L and <1 to 2258 ng/L, respectively. BE concentrations were employed to calculate the local amount of abused cocaine. The highest values (up to 1.8 g/day cocaine per 1000 inhabitants) were found in large cities and during weekends. The estimation of cocaine abuse through water analysis can be executed on regular basis without cooperation of patients. It also gives clear geographical information, while prevention campaigns can easily be implemented and evaluated.