Analytical method validation for assay determination of cannabidiol and tetrahydrocannabinol in hemp oil infused products by RP-HPLC.

A simple quantitative reverse phase high performance liquid chromatographic (RP-HPLC) method has been developed and validated for assay determination of cannabidiol and tetrahydrocannabinol in hemp oil infused products. The RP-HPLC method was developed and optimized for the mobile phase composition, flow rate, column selection and detector wavelength. An isocratic elution of samples were performed on SOLAS 100 Å C18 150 mm × 4.6 mm, 5 µm column with a mobile phase containing 75/25 acetonitrile/water v/v, with a flow rate of 1.5 mL/min by using an ultraviolet-visible (UV/Vis) detector operating at 214 nm. The RP-HPLC method was validated to meet regulatory requirements which covers specificity, accuracy, range, linearity, precision, system suitability and robustness. The validated assay test method was applied successfully to quantify cannabidiol and tetrahydrocannabinol in commercial hemp oil infused products such as tablets, soft gel capsules, plant extract oils, oral drops, tincture, and beverage enhancers. All the test results were found acceptable as per ICH guidelines, and this confirmed the feasibility of this method for its intended use in regular quality control and assay of cannabidiol and tetrahydrocannabinol in hemp oil infused products.

Synthesis and characterisation of non-bonded 1.7µm thin-shell (TS1.7-100nm) silica particles for the rapid separation and analysis of uric acid and creatinine in human urine by hydrophilic interaction chromatography.

A rapid chromatographic method for the simultaneous determination of uric acid (UA) and creatinine (Cr) in human urine is described, using a non-bonded 1.7µm thin-shell (TS1.7-100nm) silica particle prepared by the seeded-growth approach. The new shell particle was characterised by scanning electron microscopy (SEM), transmission electron microscopy (TEM), dynamic light scattering (DLS) and BET analysis. TEM reveals 1.5µm solid core and 100nm shell thickness. DLS shows polydispersity index <0.2. Expanded pore size of 88Å and specific surface area of 78m2/g were determined by BET. Chromatographic results demonstrate that UA, Cr and hypoxanthine (Hyp, as internal standard) can be separated in less than 1min on the in-house packed TS1.7-100 column (4.6 ID x 100mm), using chromatographic conditions with mobile phase 70% acetonitrile, 10mM ammonium acetate buffer, pH 6.78, flow rate 1.25ml/min and UV detection at 254nm. A linear relationship between the ratio of the peak area of the standard UA and Cr to that of the internal standard (Hyp) and the concentration of standards was obtained for both UA and Cr with the square of the correlation coefficients, R2=0.998 for both renal biomarkers. The calculated detection limits were 0.03µg/ml and 0.05µg/ml for UA and Cr respectively. Urine samples tested were found to contain UA and Cr in the concentration range of 782-1206µg/ml and 535-862µg/ml respectively. The recovery ranges for spiked urine standards were 85.7-93.2% for UA and 91.9-94.6% for Cr and the relative standard deviations (RSD) for both biomarkers were 3.05% and 0.88% respectively. The developed rapid HILIC method can have application in determining the concentrations of UA and Cr for early prediction in patients with developing disease conditions, including acute kidney injury (AKI).

Sub-2-µm seeded growth mesoporous thin shell particles for high-performance liquid chromatography: Synthesis, functionalisation and characterisation.

Nanometer control over the porous shell thickness of sub-2-µm-shell particles is investigated. Three seeded growth mesoporous thin shell particles for HPLC were prepared, with 0.05µm (or 50nm) porous shell layers: particle sizes 1.5µm (solid core diameters 1.4µm), 1.7µm (solid core diameter 1.6µm), 1.9µm (solid core diameter 1.8µm) and compared with a fourth 1.7µm particle (solid core diameter 1.4µm) surrounded by 0.15µm (or 150nm) porous shell thickness. The thin shell particles were functionalised using a mono-functional octadecyldimethylchlorosilane ligand (C20H43SiCl) under optimised reflux conditions and packed in-house in narrow bore columns (2.1 I.D.×50mm) denoted as TS1.5-50-C18, TS1.7-50-C18, and TS1.9-50-C18 respectively. The synthesised thin shell particles and bonded materials were comprehensively characterised using scanning electron microscopy (SEM), transmission electron microscopy (TEM), dynamic light scattering (DLS), zeta potential, BET analysis, elemental analysis (CHN), thermogravimetric analysis (TGA) and diffuse reflectance infrared Fourier transform (DRIFT) spectroscopy. Experimental data from inverse size exclusion chromatography (ISEC) was used to measure external, internal and total column porosities. Five probe analytes (uracil, naphthalene, acetophenone, benzene and toluene) were chosen for the chromatographic performance analysis of these columns. Column evaluation and measurements of height equivalent to a theoretical plate (HETP) data were performed on naphthalene using 55% acetonitile in water. The retention coefficients for the thin shell particles (TS1.9-50-C18, TS1.7-50-C18, TS1.5-50-C18) were in the range 1.26-1.35 and 5.6 for the core-shell particle (EiS1.7-150-C18). The minimum reduced plate heights range from 3.89 to 4.26 for the thin shell particles and 2.03 for the core-shell particle.