AlphaLogD Determination: A New Generation Reversed-Phase HPLC Method to Measure Lipophilicity on Neutral and Basic Small and Beyond-Rule-of-Five Compounds

Lipophilicity can be measured with different methods, such as Shake-Flask or liquid chromatography. HPLC presents the advantage of overcoming solubility issues and therefore extending the range of lipophilicity to high values. A specific HPLC method, called ELogD, had been developed 20 years ago on a C16-amide stationary phase, enhancing hydrophobic and hydrogen bond interactions to mimic octanol-water partition. The emergence of novel stationary phases added to the need for a less complex mobile phase has led to the development of a new HPLC assay called alphaLogD, applicable to neutral and basic compounds at pH 7.4, that combines superficially porous particles, a high number of equilibriums between solutes and stationary phase, and results in a lower number of isocratic methods to determine the logk’w at a higher throughput. Statistical studies have been run to successfully evaluate the alphaLogD method compared to the Shake-Flask method and to allow this lipophilicity measurement into the so-called Beyond-Rule-of-5-molecules space.

Determinations of Labile Nutrients (Methylcobalamin, 5-Methyltetrahydrofolate, and Trans Menaquinone-7) in Oil- and Beadlet-Filled, Gastric-Resistant Capsule

Objectives The dietary supplement industry has grown substantially, and advances in technology have played important roles in the development of new forms of dietary supplements and delivery systems. While it is recommended to use compendia methods for testing dietary supplements, the analytical methods in place are often developed for single ingredients or simple matrices and are not always fit to be used for solving modern matrix challenges. Oil- and beadlet-filled, gastric acid-resistant capsule (OBGRC) is designed to improve nutrient delivery and absorption. However, it is challenging to accurately analyze labile nutrients in OBGRCs because nutrients within different physical locations are difficult to separate, and lengthy sample preparations lead to nutrient degradation. We developed and validated analytical methods to determine methylcobalamin (MeB12), 5-methyltetrahydrofolate (5-MTHF), and trans menaquinone-7 (MK-7) in OBGRCs. Methods MeB12 and 5-MTHF were extracted by completely dissolving the OBGRC shells using aqueous buffers and then applying hexane-facilitated liquid-liquid extraction to remove the oil phase and determined by RP-HPLC using superficially porous particles columns. For MK-7 analysis, the whole OBGRC was cut to open and the oil phase was collected to dissolve in a mixture of dimethyl sulfoxide/tetrahydrofuran/ethanol before analyzed by RP-HPLC with fluorescence detection after on-line, post-column zinc reduction. The methods were subjected to single-laboratory validations according to the United States Pharmacopeia (USP) guidelines for linearity, suitability, detection limits, specificity, accuracy, and precision. Results The methods achieved chromatographic resolution of target analytes without the expensive requirement of ultra-high pressure liquid chromatography and/or mass spectrometry. The methods have wide analytical ranges (from at least 50% to 250% of the input concentrations), high precision (repeatability relative standard deviations ranging from 1.04% to 4.90%), and high accuracy (spike recovery rate ranging from 89.2% to 108%). Conclusions All three methods passed USP method validation criteria and have the potential to be widely adopted to analyze MeB12, 5-MTHF, and MK-7 in similar matrices for quality control purposes. Funding Sources The research funding was provided by Ritual and Eurofins.

A Comparative Study of Advanced Stationary Phases for Fast Liquid Chromatography Separation of Synthetic Food Colorants

Food analysis demands fast methods for routine control and high throughput of samples. Chromatographic separation enables simultaneous determination of numerous compounds in complex matrices, several approaches increasing separation efficiency and speed of analysis were involved. In this work, modern types of column with monolithic rod or superficially porous particles were employed and compared for determination of eight synthetic food dyes, their chromatographic performance was evaluated. During method optimization, cyano stationary phase Chromolith Performance CN 100 × 4.6 mm and Ascentis Express ES-CN 100 × 4.6 mm, 5 µm were selected for the separation of polar colorants. The separation was performed by gradient elution of acetonitrile/methanol and 2% water solution of ammonium acetate at flow rate 2.0 mL min−1. Mobile phase composition and the gradients were optimized in order to enable efficient separation on both columns. The method using fused-core particle column provided higher separation efficiency, narrow peaks of analytes resulted in increased peak capacity and shortening of analysis time. After the validation, the method was applied for analysis of coloured beers, soft drinks and candies.