Exhaled breath has recently been proposed as a new possible matrix for drugs of abuse testing. A key drug is cannabis, and the present study was aimed at investigating the possibility of detecting tetrahydrocannabinol and tetrahydrocannabinol carboxylic acid in exhaled breath after cannabis smoking. Exhaled breath was sampled from 10 regular cannabis users and 8 controls by directing the exhaled breath by suction through an Empore C₁₈ disk. The disk was extracted with hexane/ethyl acetate, and the resulting extract was evaporated to dryness and redissolved in 100 µL hexane/ethyl acetate. A 3-µL aliquot was injected onto the LC-MS-MS system and analyzed using positive electrospray ionization and selected reaction monitoring. In samples collected 1–12 h after cannabis smoking, tetrahydrocannabinol was detected in all 10 subjects. The rate of excretion was between 9.0 and 77.3 pg/min. Identification of tetrahydrocannabinol was based on correct retention time relative to tetrahydrocannabinol-d₃ and correct product ion ratio. In three samples, peaks were observed for tetrahydrocannabinol carboxylic acid, but these did not fulfill identification criteria. Neither tetrahydrocannabinol or tetrahydrocannabinol carboxylic acid was detected in the controls. These results confirm older reports that tetrahydrocannabinol is present in exhaled breath following cannabis smoking and extend the detection time from minutes to hours. The results further support the idea that exhaled breath is a promising matrix for drugs-of-abuse testing.

Hepatocellular carcinoma is often treated with a combination of doxorubicin and embolization, exposing it to high concentrations and hypoxia. Separation of the possible synergistic effect of this combination in vivo is difficult. Here, treatment with doxorubicin, under hypoxia or normoxia in different liver cancer cell lines, was evaluated. Liver cancer cells HepG2, Huh7, and SNU449 were exposed to doxorubicin, hypoxia, or doxorubicin + hypoxia with different duration. Treatment response was evaluated with cell viability, apoptosis, oxidative stress, and summarized with IC₅₀. The protein profile of a 92-biomarker panel was analyzed on cells treated with 0 or 0.1 µM doxorubicin during 6 or 72 h, under normoxia or hypoxia. Hypoxia decreased viability of HepG2 and SNU499. HepG2 was least and SNU449 most tolerant to doxorubicin treatment. Cytotoxicity of doxorubicin increased over time in HepG2 and Huh7. The combination of doxorubicin + hypoxia affected the cells differently. Normalized protein expression was lower for HepG2 than Huh7 and SNU449. Hierarchical clustering separated HepG2 from Huh7 and SNU449. These three commonly used cell lines have critically different responses to chemotherapy and hypoxia, which was reflected in their different protein expression profile. These different responses suggest that tumors can respond differently to the combination of local chemotherapy and embolization.

A bioalanytical method for the quantification of tacrolimus (TAC) and 3 metabolites, 13-O, 15-O, and 31-O-demethylated TAC (M-I, M-III, and M-II) in human whole blood using liquid chromatography, electrospray ionization, tandem mass spectrometry (LC-ESI-MS/MS) was developed and validated. The analytes were extracted from 85 μL of blood by protein precipitation followed by solid-phase extraction and a concentration step. The analytes and the internal standard (IS, ascomycin) were separated on a C18 column using a slow gradient mobile phase elution, with an analysis time of 3.3 minutes. The ammonium-adduct ions with transitions of m/z 821.5 > 768.7 (TAC), 807.5 > 754.7 (M-I, M-III, M-II), and 809.4 > 756.7 (IS) were measured in selected reaction monitoring mode using electrospray ionization. Measuring ranges were 0.1-50 ng/mL for M-II, M-III, and TAC and 0.15-39 ng/mL for M-I. Imprecision in quantification was 50% for all analytes. The sample's stability was proven for 1 month at -20°C and 72 hours at room temperature. Three freeze-thaw cycles had no significant effect on the stability. The prepared samples were stable at least 16 hours at 8°C. Analysis of 53 patient samples resulted in average concentrations of 7.2 for TAC, 0.8 for M-I, 0.4 for M-III, and 0.2 ng/mL for M-II. The total metabolite concentration was 17% (4%-52%) of the TAC concentration. The TAC concentration measured by LC-MS/MS was 36.1% ± 27.1% lower than by immunochemical (enzyme multiplied immunoassay technique) analysis. When adding the metabolite crossreactivity in the presence of TAC, the difference between the 2 methods was still 29.8% ± 28.3%, indicating that the overestimation of TAC concentration of enzyme multiplied immunoassay technique compared with liquid chromatography-tandem mass spectrometry cannot only be ascribed to the demethylated metabolites. An LC-ESI-MS/MS method for the quantitative analysis of TAC and 3 metabolites, using a 2-step sample preparation was successfully developed, validated, and applied on 53 patient samples.

The biopharmaceutical properties of doxorubicin delivered via two drug-delivery systems (DDSs) for the palliative treatment of unresectable hepatocellular carcinoma were reviewed with relation to the associated liver and tumor (patho) physiology. These two DDSs, doxorubicin emulsified with Lipiodol (R) and doxorubicin loaded into DC Bead (R) are different regarding tumor delivery, release rate, local bioavailability, if and how they can be given repeatedly, biodegradability, length of embolization and safety profile. There have been few direct head-to-head comparisons of these DDSs, and in-depth investigations into their in vitro and in vivo performance is warranted.

Gastrointestinal (GI) mucus is continuously secreted and lines the entire length of the GI tract. Essential for health, it keeps the noxious luminal content away from the epithelium. Our aim was to characterize the composition and structure of mucus throughout the various GI segments in dog. Mucus was collected from the stomach, small intestine (duodenum, jejunum, ileum), and large intestine (cecum, proximal and distal colon) from dogs. Composition was determined by multi-omics. Structural properties were investigated using cryoSEM and rheology. GI mucus contained 74-95% water and maintained a pH around 6.5. The proteome was similar across the different GI segments. The highest abundant secreted gel-forming mucin in the gastric mucus was mucin 5AC, whether mucin 2 had highest abundance in the intestinal mucus. Lipid and metabolite abundance was generally higher in the jejunal mucus than the colonic mucus. CryoSEM microscopy revealed smaller pore size in small intestinal mucus, which increased in the large intestine. All mucus samples showed shear-thinning behavior and characteristics of gel-like structure. In conclusion, the mucus is a highly viscous and hydrated material. These data provide an important baseline for future studies on human and canine intestinal diseases and the dog model in drug absorption.