Packed in‐tube SPME–LC–MS/MS for fast and straightforward analysis of cannabinoids and metabolites in human urine

Abstract

Cannabinoids are pharmacologically active compounds present in cannabis plants, which have become important research topics in the modern toxicological and medical research fields. Not only is cannabis the most used drug globally, but also cannabinoids have a growing use to treat a series of diseases. Therefore, new, fast, and efficient analytical methods for analyzing these substances in different matrices are demanded. This study developed a new packed-in-tube solid-phase microextraction (IT-SPME) method coupled to liquid chromatography with tandem mass spectrometry (LC-MS/MS), for the automated microextraction of seven cannabinoids from human urine. Packed IT-SPME microcolumns were prepared in (508 µm i.d. × 50 mm) stainless-steel hardware; each one required only 12 mg of sorbent phase. Different sorbents were evaluated; fractional factorial design 2 4−1 and a central composite design were employed for microextraction optimization. Under optimized conditions, the developed method was a fast and straightforward approach. Only 250 µl of urine sample was needed, and no hydrolysis was required. The sample pretreatment included only dilution and centrifugation steps (8 min), whereas the complete IT-SPME-LC-MS/MS method took another 12 min, with a sample throughput of 3 samples h −1 . The developed method presented adequate precision, accuracy and linearity; R 2 values ranged from 0.990 to 0.997, in the range of 10-1000 ng ml −1 . The lower limits of quantification varied from 10 to 25 ng ml −1 . Finally, the method was successfully applied to analyze 20 actual urine samples, and the IT-SPME microcolumn was reused over 150 times.

The Determination of Cannabinoids in Urine Samples Using Microextraction by Packed Sorbent and Gas Chromatography-Mass Spectrometry

“…The current method achieves lower or similar LODs than the literature for: CBD [ 38 , 39 , 40 , 41 , 42 ]; THC [ 33 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 ]; CBN [ 41 ]; 11-OH-THC [ 33 , 38 , 39 , 40 , 41 , 42 , 43 , 44 ]; and THC-COOH [ 38 , 46 , 47 ]. The same LLOQs are reported for: CBD [ 39 , 40 , 41 , 42 , 48 , 49 ]; THC [ 33 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 48 , 49 ]; CBN [ 39 , 40 , 41 , 48 , 49 ]; 11-OH-THC [ 33 , 39 , 40 , 41 ,…”

“…When using a GC–MS system [ 41 , 43 , 44 ], the authors report greater LODs and LLOQs than those obtained with the herein proposed technique. The same happens when using systems with greater sensitivity, such as LC–MS/MS [ 38 , 39 , 40 , 42 , 45 , 46 , 47 , 48 , 49 ]. Nonetheless, some authors report lower LLOQs than those obtained with our MEPS–GC–MS, mostly using techniques with greater sensitivity, including GC–MS/MS [ 50 , 51 , 52 , 53 ] and LC–MS/MS [ 38 , 39 , 42 , 46 , 54 , 55 , 56 , 57 , 58 , 59 ]; several of these approaches also used larger sample volumes.…”

“…Micro-solid-phase extraction (µ-SPE) [ 39 ], molecularly imprinted solid-phase extraction (MISPE) [ 43 , 45 ], disposable pipette extraction (DPX) [ 42 , 55 , 63 ], dispersive liquid–liquid microextraction (DLLME) [ 53 , 60 ], packed in-tube solid-phase microextraction (IT-SPME) [ 48 ], and hollow fiber membrane solvent microextraction (HFMSME) [ 64 ] are other miniaturized sample preparation techniques that have been applied to urine samples to extract cannabinoids. However, extraction efficiencies were not reported for some of the latter, namely DPX [ 63 ], SMPE [ 48 ], and DLLME [ 53 ].…”

Analysis of Cannabinoids in Biological Specimens: An Update

“…Likewise, Morisue Sartore et al [63] developed a new packed-in-tube solid-phase microextraction (IT-SPME) technique coupled to LC-MS/MS to automatically extract THC, THCV, THC-OH, THC-COOH, THC-COOH-gluc, CBN, and CBD from urine samples. Thus, both metabolites and neutral cannabinoids were analysed.…”

Modern automated microextraction procedures for bioanalytical, environmental, and food analyses

“…Although it required several days for sample pre‐treatment, including oxidative stress (1 day), deamidation (3–7 days), and glycation (7 days), the mD‐LC‐MS analysis time was under 2 h and required only 10 μg of protein. Finally, in 2022, Sartore and coworkers [109] developed a packed IT‐SPME‐LC‐MS/MS for the simultaneous determination of cannabinoids and metabolites in urine. Using 508 μm id × 50 mm stainless‐steel hardware, the authors prepared the packed IT‐SPME micro columns, which required only 12 mg of commercial extractive phase each.…”