Cannabinoids

Cannabinoids are practically stable during storage, with concentration changes attributed to oxidation, temperature effects, and lipophilic binding. Studies prior to 1980 focused on the biologically active Δ9-tetrahydrocannabinol (THC) and showed that it was oxidized to produce cannabinol, cannabidiol, hydroxyquinone, and polymers formed through phenolic coupling. Yields of these products depended on the solution pH, amount of oxygen, and presence of free-radical compounds. The rate of oxidation was more dependent on the storage medium than on atmosphere or temperature. While THC in dry forms chloroform, hexane, and carbon tetrachloride deteriorated about 10% each month, ethanolic solutions were stable for more than 75 days at room temperature and for more than 1 year at 0-5 °C.

THC and 11-Nor-Δ9-Tetrahydrocannabinol-9-Carboxylic Acid in Blood

Johnson et al. reported THC in blood and plasma was stable at room temperature for 2 months but dropped 90% in 6 months. At 4 °C and -10 °C, concentrations of drug test THC in the blood did not change over the entire Month period. A separate study by Wong et al. showed that, when stored between -20 °C and 5 °C, THC concentrations had decreased notably after 17 weeks in blood samples and after 19 weeks in serum samples. Atmospheric conditions and repeated freezing and thawing seemed to have little effect. Over a 1-day period, elevated temperatures (20 – 60 °C) had no effect on THC concentrations.

Being a lipophilic molecule, THC binds to hydrophobic surfaces, which can cause a reduction in concentrations in solutions stored in certain types of containers. Garrett and Hunt reported adsorption of THC to rubber stoppers but Johnson et al. found no adsorption to rubber stoppers in their blood collection tubes. Christophersen studied the stability of blood in unsilanized glass and polystyrene tubes over 4 days at room temperature and over 4 weeks at -20 BIG. Blood concentrations of THC were stable in the glass tubes but 60 to 100% of the THC stored in the plastic containers was lost. This reduction was due principally to adsorption to the plastic that could not be prevented by binding proteins in the blood. Absorption of THC by collection devices was also a problem for oral fluid specimens. Although THC sticks to unsilanized glass, this apparently was prevented by proteins in blood that bind over 97% of THC. Metabolites are generally more stable than THC in blood.

In the study cited above, Johnson et al. showed the concentrations of THC and 11-hydroxy-tetrahydroca~nabinol in blood and plasma were significantly lower after 6 months at room temperature, but 11-nor-Δ9-tetrahydrocannabinol-carboxylic acid (THCCOOH) concentrations did not change GOB. McCurdy et al. confirmed the stability of THCCOOH for a 30-day period and added that heparin, fluoride, or EDTA in the blood at collection produced equivalent results. Skopp et al. examined the stability of the glucuronide conjugate of THCCOOH by adding the compound to drug-free plasma to produce a final concentration of 300ng/mL.

Different portions were stored in glass containers at -20 °C, 4 °C, and 20 °C. The glucuronide was stable in plasma stored at -20 °C for 10 days. Over the same time period, there was degradation of the glucuronide at 4 °C and 20 °c in an apparent first-order kinetic process.

THCCOOH in Urine

Many investigations since 1980 have focused on the stability of THCCOOH the principle urinary metabolite of THC, because of its importance in drug testing. According to Fuhrmann and Szasz, THCCOOH is stable for 2 years at 4 °C in pre-filtered urine containing sodium aside preservative. In another unpublished study, THCCOOH in unfiltered urine was stable for over 1 year at 4 °C and -20 °C, but samples began to deteriorate after 10 weeks at room temperature.

Giardino found that urine spiked with THCCOOH concentration of 75ng/mL stored in a Nalgene® polyethylene container at 2 to 8 °C had less than a 10% decrease over a 78 day period. Multiple studies have tracked the degradation of THCCOOH in spiked urine specimens. A number of reasons for this loss have been proposed: adsorption to the container or to solid matter, concentration of this amphipathic molecule in the foam created during mixing or urine pH. In the previously discussed unpublished study, THCCOOH in unfiltered urine began to deteriorate after 10 weeks at room temperature.

Dextraze et al. showed a 27% loss of THCCOOH resulting from adsorption to the glass container. This loss can be prevented by the addition of albumin. There was an 89% loss of THCCOOH in foaming urine that can be reversed by adding a deforming agent 2-octonol. The presence of albumin did not eliminate entrapment of THCCOOH in the foam.

Craft et al. found that there was a 25% loss of THCCOOH during Iyophilization, but that the final Iyophilized material was stable for over 1 year at refrigeration temperature. Cody reported that THCCOOH in unfiltered urine decreased by 60 to 84% in polypropylene shipping tubes, plastic urine-collection containers, unsilanized glass tubes, and silanized glass tubes that had been gently rocked for 24 h at room temperature.

When refrigerated the loss in plastic bottles was 7 to 25% after 1 week. Stout et al. examined the reduction in THCCOOH concentrations ~ urine specimens stored either in polypropylene or polyethylene containers at 4 °C and 25 °C over a 1-week period. At 4 °C, there was a rapid initial decrease (within 1 h) in urine concentrations in both types of containers. After this initial decrease (14-17% maximum), the urine concentrations were stable over the rest of the period. At 25 °C, there was less than a 5% decrease in concentration in the polypropylene and no significant decrease in polyethylene containers. The authors hypothesized that at the lower temperature, THCCOOH was less soluble in urine, resulting in greater adherence to the plastic.

Jamerson et al. found greater degradation of THCCOOH at pH 4.6 than at neutral or slightly alkaline pH. Skopp and Potsch spiked THCCOOH glucuronide to urine specimens and stored at -20 °C, 4 °C, and 20 °C for 10 days. The glucuronide was stable at -20 °C and at 4 °C over this period. At 20°C, degradation of the glucuronide to THCCOOH occurred within 2 days. A number of studies have also looked at the stability of THCCOOH in authentic urine specimens. Cody compared results from 900 collected urine specimens to results from samples prepared with commercial THCCOOH. Before freezing at -20 °C, the concentration of total THCCOOH, free and conjugated, was determined by gas chromatography-mass selective detection GC-MS. The samples were thawed and reanalyzed using the same methodology at intervals ranging from 1 week to 2 years.

Selected samples had deteriorated significantly and the deterioration could not be correlated with storage time or pH. Prepared controls analyzed by GC MS were stable up to 6 months when stored frozen. One conclusion of this study was that the results from these prepared samples failed to predict loss of THCCOOH in real samples. An anecdotal finding of this study was that a set of controls inadvertently left at room temperature for 1 week before analysis had lower values. Gere et al. presented results for patient samples similar to Cody’s results. Romberg and Past retested 85 urine specimens positive for THCCOOH that were stored frozen for up to 1 year following the initial analysis. The average decrease in concentration of the retested specimens was 24%.

A normal bell-shaped distribution was observed, with a range of concentration changes between 30 and 80%. The influence of factors such as microbial growth and precipitate in urine has not been reported. In a stabilizer study published by Golding et al., 40 urine specimens positive for THCCOOH were stored at -18 °C, 4°C, and 20-25 °C for 10 days. At the frozen temperature, there was an average decline of approximately 20% in THCCOOH concentrations after 3 years. At room temperature, there was a similar decline after 10 days. At 4 °C, there were minimal losses <10% after 4 weeks. Skopp and Potsch tracked the stability of both free and corrugated THCCOOH in 38 specimens at -20 °C, 4 °C, and 20 °C for 15 days. Initially, 18 specimens tested positive for free THCCOOH.

After 2 days of storage at room temperature, free THCCOOH was detected in 4 additional specimens; after 15 days, 7 more specimens were positive. The glucuronide was more stable at 4 °C than at 20 °C, but decreases were observed over the 15 day period. Under frozen conditions, both free and corrugated THCCOOH were stable after 15 days.

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