EDTA (Ethylenediaminetetraacetic acid) is amino acid based compound which is widely used in industrial cleaners, detergents, photography, agrochemicals, textile, and food industries. EDTA serves as preservative in packaged food, vitamins and pharmaceutical formulations. It is used in organic and analytical laboratories as scavenger of metals, buffer solution, complexometric titration, masking agent for metal determination, etc.
EDTA is very polar compound with strong chelating properties. Low UV activity and strong binding to metal ions makes HPLC analysis of EDTA very difficult. Reproducible method for determination of wide range on concentration of EDTA in various formulations is developed using Primesep D anion-exchange mixed mode column. Copper sulfate is used for visualization purposes to increase sensitivity of the method. EDTA forms UV active complex with some metal ions. This method can be used for highly accurate quantitation of EDTA in different mixtures and composition. Method demonstrates controllable retention and perfect peak shape. Equilibration of the column prior to analysis requires special attention.
If multiple injections produce different retention times, your column might not be equilibrated properly. Primesep D columns have a large ionic capacity towards anions. The shipping solvent for the Primesep D columns is ACN/water/TFA. The end point of gradient has only 4 mmol of sulfate ions. It takes over 5 hours to equilibrate a column with such low concentration. Before running your experiments you need to wash your column with 20% ACN and 0.2% of sulfuric acid for 1 hour. After you replace previous buffer/additive, you can set up your experiment. If you don’t change ionic modifier in your mobile phase the equilibration time is equivalent of 4-5 column volumes.
EDTA, or Ethylenediaminetetraacetic acid, is widely used in textile, pulp/paper, food and pharmaceutical industries. It has application as chelating agent and preservative. EDTA molecule is very hydrophilic and contains two basic groups and four carboxylic groups. It has tendency to bind to metals, making analysis of EDTA by HPLC very challenging. EDTA is analyzed in the presence of copper sulfate as visualization agent on a Primesep SB column. The method can be used for EDTA determination in various mixtures and compositions using UV detector.
HILIC Separation of Common Preservatives - Citric Acid, Ascorbic Acid and EDTA
Citric acid, ascorbic acid, and EDTA are commonly used in food and pharmaceutical industry as preservatives. These compounds are very polar in nature. They are weak organic acids with limited UV activity. Retention and separation is achieved on HILIC mixed-mode Obelisc N column. All three compounds are retained by combination of strong HILIC and strong anion-exchange mechanisms. Separation can be monitored by ELSD, LC/MS, UV or Corona CAD. In contrast to other HILIC column, Obelisc N has two ionizable groups basic and acidic which provide ion-exchange interaction in addition to hydrophilic interaction. This allows to use less acetonitrile for HILIC separation.
EDTA Standards Solution A:
For the preparation of the EDTA standard solution, 5 mg of EDTA was accurately weighed and transferred into a 5 mL volumetric flask and dissolved in water with sonication. The EDTA stock solution (1 mg/mL) should be stored in a cold, dark place and can be used for upto a week to prepare standards of required concentration.
Iron(III) chloride Solution B:
The standard stock solution of Iron(III) chloride (10 mg/ml) was prepared in water. 50 mg of FeCl3 was accurately weighed and transferred into a 5 mL volumetric flask and dissolved in water, with sonication if needed.
General procedure for Ferric EDTA complex analysis:To make a sample for analysis mix 100 µL Solution A (or unknown sample) with 100 µL Solution B and 800 µL of water. Place this mixture in a plastic HPLC vial for analysis. Setup instrument and column according to the method provided.