Development, validation, and uncertainty measurement of HPLC-DAD method for determination of some free amino acids in infant formula and medical food products for inborn errors of metabolism.

This research aims at determining some free amino acids in amino acid-based infant formulas and amino acid-modified medical foods for inborn errors of metabolism to prove their quality. A method based on high-performance liquid chromatography and diode array detection was developed and validated. Then, overall uncertainty was estimated by the bottom-up approach. Applying the weighted least squares regression method suggested good linearity with coefficient of determinations ≥ 0.9960. The limits of detection were calculated between 0.01 and 0.28 µg/mL. The most repetitive recovery values were obtained in the range of 91-108%, with RSDs ≤ 15%. The expanded uncertainties were below 20% for most amino acids. The contributions of linear regression and repeatability are two main factors in estimating overall uncertainty. The results offer this method as a simple and easy procedure for analyzing free amino acids in seven powdered medical foods designed for phenylketonuria, maple syrup urine disease, methylmalonic, and propionic acidemia.

Clogging sensitivity of flow distributors designed for radially elongated hexagonal pillar array columns: a computational modelling.

Flow distributor located at the beginning of the micromachined pillar array column (PAC) has significant roles in uniform distribution of flow through separation channels and thus separation efficiency. Chip manufacturing artifacts, contaminated solvents, and complex matrix of samples may contribute to clogging of the microfabricated channels, affect the distribution of the sample, and alter the performance of both natural and engineered systems. An even fluid distribution must be achieved cross-sectionally through careful design of flow distributors and minimizing the sensitivity to clogging in order to reach satisfactory separation efficiency. Given the difficulty to investigate experimentally a high number of clogging conditions and geometries, this work exploits a computational fluid dynamic model to investigate the effect of various design parameters on the performance of flow distributors in equally spreading the flow along the separation channels in the presence of different degrees of clogging. An array of radially elongated hexagonal pillars was selected for the separation channel (column). The design parameters include channel width, distributor width, aspect ratio of the pillars, and number of contact zone rows. The performance of known flow distributors, including bifurcating (BF), radially interconnected (RI), and recently introduced mixed-mode (MMI) in addition to two new distributors designed in this work (MMII and MMIII) were investigated in terms of mean elution time, volumetric variance, asymmetry factors, and pressure drop between the inlet and the monitor line for each design. The results show that except for pressure drop, the channel width and aspect ratio of the pillars has no significant influence on flow distribution pattern in non-clogged distributors. However, the behavior of flow distributors in response to clogging was found to be dependent on width of the channels. Also increasing the distributor width and number of contact zone rows after the first splitting stage showed no improvement in the ability to alleviate the clogging. MMI distributor with the channel width of 3 µm, aspect ratio of the pillars equal to 20, number of exits of 8, and number of contact zones of 3 exhibited the highest stability and minimum sensitivity to different degrees of clogging.

Validated determination of losartan and valsartan in human plasma by stir bar sorptive extraction based on acrylate monolithic polymer, liquid chromatographic analysis and experimental design methodology.

In our previous work, a new monolithic coating based on vinylpyrrolidone-ethylene glycol dimethacrylate polymer was introduced for stir bar sorptive extraction. The formulation of the prepared vinylpyrrolidone-ethylene glycol dimethacrylate monolithic polymer was optimized and the satisfactory quality of prepared coated stir bar was demonstrated. In this work, the prepared stir bar was utilized in combination with ultrasound-assisted liquid desorption, followed by high-performance liquid chromatography with ultraviolet detection for the simultaneous determination of losartan (LOS) and valsartan (VAS) in human plasma samples. In a comparison study, the extraction efficiency of the prepared stir bar was accompanied much higher extraction efficiency than the two commercial stir bars (polydimethylsiloxand and polyacrylate) for both target compounds. In order to improve the desorption efficiency of LOS and VAS, the best values for effective parameters on desorption step were selected systematically. Also, the effective parameters on extraction step were optimized using a Box-Behnken design. Under the optimum conditions, the analytical performance of the proposed method displayed excellent linear dynamic ranges for LOS (24-1000 ng mL-1) and VAS (91-1000 ng mL-1), with correlation coefficients of 0.9998 and 0.9971 and detection limits of 7 and 27 ng mL-1, respectively. The intra- and inter-day recovery ranged from 98 to 117%, and the relative standard deviations were less than 8%. Finally, the proposed technique was successfully applied to the analysis of LOS and VAS at their therapeutic levels in volunteer patient plasma sample. The obtained results were confirmed using liquid chromatography-mass spectrometry. The proposed technique was more rapid than previously reported stir bar sorptive extraction techniques based on monolithic coatings, and exhibited lower detection limits in comparison with similar methods for the determination of LOS and VLS in biological fluids. The obtained results were demonstrated that the lower selectivity of UV in comparison with MS detection was rectified by appropriate sample preparation through proposed extraction method to eliminate as many interfering compounds as possible.

Determination of Carvedilol Enantiomers in Pharmaceutical Dosages by SBSE-HPLC Based on Diastereomer Formation.

A sensitive, selective and simple method for the simultaneous determination of carvedilol enantiomers in aqueous solution has been developed using stir bar sorptive extraction (SBSE) followed by high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection. This method is based on the reaction of carvedilol enantiomers with (-)-menthyl chloroformate (MCF) after extraction by the SBSE method to produce diastereomeric derivatives. The separation was achieved by use of a C18 analytical column and the influence of mobile phase composition on the enantioseparation of carvedilol was studied. The applicability of two sorptive phases, poly(methyl methacrylate/ethyleneglycol dimethacrylate) (PA-EG) and polydimethylsiloxane, were tested for extraction of carvedilol enantiomers from aqueous samples. The obtained results showed excellent linear dynamic ranges and precisions for each of them. The least limit of detection for (S)- and (R)-carvedilol obtained 8 and 11 µg L(-1), respectively, using the PA-EG sorptive phase. Inter- and intra-mean recoveries were also satisfactory, ranging from 98 to 103%, with coefficient of variation in the range of 1-5% at three fortified levels using a PA-EG coated stir bar. The proposed SBSE (PA-EG)-MCF derivatization-HPLC-UV method was successfully applied to enantioselective analysis of carvedilol in water and pharmaceutical dosages, confirming the application of this method.