In the study presented, a simple analytical method for the direct determination of glycine in immunoglobulins by hydrophilic interaction liquid chromatography was developed. The HPLC separation was performed using a SeQuant ZIC-HILIC column (250 mm × 4.6 mm i.d, 5 μm) with the isocratic mobile phase consisting of ammonium formate (20 mM) and acetonitrile (30:70, v/v), and the flow rate set at 0.8 mL/min. UV detection was carried out at a wavelength of 210 nm. The procedure was validated for specificity, precision, linearity, accuracy, limit of detection, limit of quantitation, and robustness. The calibration curve was found to be linear within the concentration range of 1.2-3.6 mg/mL. RSD values for intra-day and inter-day precision were in the range of 0.66 to 1.84%. The limit of quantification and limit of detection were 0.10 mg/mL and 0.03 mg/mL, respectively. The developed chromatographic method was applied for the glycine analysis in various immunoglobulins.
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