Appetite stimulation with cannabis-based medicine and methods for assessment of glomerular filtration in older patients with medical illness: A study protocol.

BACKGROUND AND AIM: Malnutrition in older patients is linked to poor appetite. Cannabis-based medicine may have orexigenic properties in older patients, but this has to our knowledge never been investigated. In older patients, uncertainty applies to the accuracy of estimated glomerular filtration rate (eGFR) based on creatinine, which is crucial for medication prescribing. In older patients with poor appetite, the study aims (1) to assess the efficacy of Sativex® (8.1-mg delta-9-tetrahydrocannabinol [THC] and 7.5-mg cannabidiol [CBD]) to stimulate appetite and (2) to compare the performance of various GFR-estimates and measured-GFR (mGFR) for determining gentamicin clearance utilizing population pharmacokinetic (popPK) modelling methods. METHODS AND OBJECTIVES: This study is composed of two substudies. Substudy 1 is an investigator-initiated single-center, double-blinded, randomized, placebo-controlled, superiority, cross-over study. Substudy 1 will recruit 17 older patients with poor appetite, who will also be invited to substudy 2. Substudy 2 is a single-dose pharmacokinetics study and will recruit 55 patients. Participants will receive Sativex® and placebo in substudy 1 and gentamicin with simultaneous measurements of GFR in substudy 2. The primary endpoints are as follows: Substudy 1-the difference in energy intake between Sativex® and placebo conditions; substudy 2- the accuracy of different eGFR equations compared to mGFR. The secondary endpoints include safety parameters, changes in the appetite hormones, total ghrelin and GLP-1 and subjective appetite sensations, and the creation of popPK models of THC, CBD, and gentamicin.

Therapeutic drug monitoring of flecainide in serum using high-performance liquid chromatography and electrospray mass spectrometry.

High-performance liquid chromatography with electrospray mass spectrometry (LC-MS) was used for analysis of the drug flecainide in serum. The clean-up was performed by solid-phase extraction, and an aromatic ring positional isomer was used as internal standard. Results from method validation on spiked serum samples showed excellent reproducibility; intra- and inter-assay variations (C.V.% and %Bias) were less than 6% within the therapeutic concentration range of the drug (0.2-1.0 microg/ml). Linearity was demonstrated from 0.05 to 2.0 microg/ml. The limit of detection and quantification was 0.025 and 0.05 microg/ml, respectively. Due to the high selectivity of the mass spectrometric detection, no interferences were observed. Results from clinical samples (n=18) from patients in treatment with Tambocor (flecainide acetate) showed excellent correlation with parallel data obtained from a method based on high-performance liquid chromatography (HPLC) with fluorescence detection after liquid/liquid extraction. The chromatographic separation of flecainide and internal standard was improved compared to earlier HPLC methods. The methodology is simple, accurate and requires only 0.25 ml of sample. It is a well suited method for routine therapeutic drug monitoring in a hospital or clinical chemistry laboratory.

Plasticizers in total diet samples, baby food and infant formulae.

The plasticizers di-n-butylphthalate (DBP), butylbenzylphthalate (BBP), di-2-(ethylhexyl)phthalate (DEHP) and di-2-(ethylhexyl)adipate (DEHA) were analysed in 29 total diet samples, in 11 samples of baby food and in 11 samples of infant formulae. In all of the total diet samples the presence of one or more of the plasticizers was demonstrated. Maximum and minimum mean concentrations in the total diet samples were: 0.09-0.19 mg DBP/kg, 0.017-0.019 mg BBP/kg, 0.11-0.18 mg DEHP/kg and 0.13-0.14 mg DEHA/kg. One or more of the phthalates was also found in about 50% of the samples of baby food as well as in infant formulae. The calculated mean maximum intakes of the individual compounds from the total diet samples were below 10% of the restrictions proposed by the EU Scientific Committee for Food (SCF), and the spread in individual intakes was considerable. DEHP was the plasticizer determined most frequently and contributed the highest fraction of its tolerable daily intake (TDI). Hence, the maximum calculated intake of DEHP from single samples of the foodstuffs analysed could be up to one-third of the TDI. The calculated mean intake of DEHA was about 1% of the TDI with a maximum value of 13% of the TDI. Violations of the restrictions proposed by the EU Scientific Committee for Food (SCF) in the form of TDI values or specific migration limits were not found in this investigation.

Determination of 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid in urine using high-performance liquid chromatography and electrospray ionization mass spectrometry.

High-performance liquid chromatography with electrospray ionization mass spectrometry was used to determine 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) in urine. After basic hydrolysis of conjugates, the compound was extracted using SPEC-PLUS-3ML-C18 solid-phase extraction columns. A deuterium labelled internal standard (d3-THC-COOH) was added prior to hydrolysis. Separation was performed on a reversed-phase Zorbax Eclipse XDB-C8 analytical column (150x3.0 mm I.D.) using a gradient program from 60 to 80% acetonitrile (4 mM formic acid) at a flow-rate of 0.5 ml/min. The compounds were detected by single ion monitoring of m/z 345 and m/z 348 for the protonated molecules [THC-COOH+H]+ and [d3-THC-COOH+H]+, respectively. The precision and accuracy were tested on spiked urine samples in the range 2.5-125 ng/ml. The mean recovery was 95% (n = 58), coefficients of variations were 2.2-4.3% and the limit of detection 2 ng/ml. Diagnostic qualifying ions of THC-COOH (m/z 327 and m/z 299) and d3-THC-COOH (m/z 330) were generated using up-front collision-induced dissociation. The relative ion intensities in clinical samples (n = 21) were within +/-20% deviation compared with standards. Using this tolerance and the presence of the ions m/z 327 and m/z 299 at the correct retention times as the acceptance criteria for identification of THC-COOH positive samples, the limit of detection was 15 ng/ml. The LC-MS method complies with the current recommendations on drugs of abuse testing, in which mass spectrometric detection is emphasized.

Validation of urine drug-of-abuse testing methods for ketobemidone using thin-layer chromatography and liquid chromatography-electrospray mass spectrometry.

High-performance thin-layer chromatography (TLC) with visual detection (post-chromatographic derivatization) was used in screening for the drug ketobemidone in human urine samples. High-performance liquid chromatography with electrospray mass spectrometry (LC-ESI-MS) was used for final confirmation of the result. The clean-up was performed by mixed-mode solid-phase extraction, and nalorphine was used as internal standard. A screening cut-off for TLC was established at 0.2 microg/ml. The mean recovery for LC-MS was 91% (n=60) with coefficients of variation (C.V.) in the range of 7 to 16%. Qualifying fragment ions of ketobemidone (m/z 190, 201 and 230) were generated by up front collision-induced dissociation (CID) on a single quadrupole instrument. Relative ion intensities were within +/- 15% deviation compared with standards in the same batch. The limit of detection for LC-MS was 0.025 microg/ml. Positive clinical samples from drug abusers (n=10) had concentrations in the range 0.07 to 3.2 microg/ml, which could be determined by LC-MS without matrix interference. During screening of unknown clinical samples (n=27) the results from TLC was in agreement with LC-MS data. After acid hydrolysis of conjugates in clinical samples the analyte response of ketobemidone and norketobemidone was increased by a factor of approximately two and twelve, respectively. A qualitative GC-MS technique was demonstrated for the detection of the spasmolyticum A29 (N,N-dimethyl-4,4-diphenyl-3-buten-2-amine), which can be found in a preparation combined with ketobemidone (Ketogan).

Specific migration of di-(2-ethylhexyl)adipate (DEHA) from plasticized PVC film: results from an enforcement campaign.

A control campaign on the correct labelling of plasticized PVC film according to current legislation on food contact materials has been performed. Analytical methods based on the isotope dilution technique were developed. For enforcement purposes, the films were exposed to the official food simulant, olive oil, followed by clean-up using size exclusion chromatography and final determination of di-(2-ethylhexyl) adipate (DEHA) by combined capillary gas chromatography and mass spectrometry (GC-MS). In the initial screening, the samples were exposed to the alternative food simulant, isooctane, and DEHA could be determined by GC-MS without further clean-up. A good consistency between results from the two different methods was obtained. During the campaign, 49 samples of PVC films, the majority intended for use in retail shops, were sampled from importers and wholesalers by the Municipal Food Control Units. Initially, all films were screened for the migration into isooctane (exposed 2 h at 40 degrees C) of DEHA and other potentially present low molecular weight plasticizers using full scanning mass spectrometry. Films showing a substantial migration of DEHA were further tested with olive oil according to the declared field of application (exposed for 10 days at 40 degrees C). In 47 of the 49 films the migrate contained a substantial amount of DEHA. In 46 films the migration exceeded the specific migration limit of 3 mg/dm2 after use of the relevant reduction factor given in legislation. However, because of the general uncertainty of the analytical method and because the variation in the thickness of the films was calculated to be 1 mg/dm2, the action limit in this campaign was 4 mg/cm2. A migration higher than this action limit was found in 42 films (89% of the samples) and these films were deemed to be illegal according to their present declared field of application as given by their labelling. In a few cases, some migration of the plasticizer di-n-butyl phthalate was seen.