New column for high performance Liquid chromatography (HPLC)

New stationary phases for high performance liquid chromatography has been prepared by the reaction of Bromophenol blue salt solution with Amberlite CG-400 (ClP P) resin . The stability of the new attached functional group on the resin was studied against different solvents such as benzene, ethanol, acetonitrile, acetone, methanol, chloroform, and petroleum ether as well as NaOH and HCl solutions. The new prepared resin was found stable and no depletion of the Bromophenol blue moity from the resin was noticed with the above materials .The FTIR analysis has configured the attachment of Bromophenol on the resin as well as the physical appearance of the new resins. The new stationary phase has been packed into stainless steel column. The resin capacity of newly prepared resin has calculated. The resin capacity for the Amberlite- bromophenol blue salt was found to average 5.27 meq./g which is more than the Amberlite resin, 3.95 meq./g. The chromatographic performance of the packed column was characterized. The number of plate numbers N, height equivalent of a theoretical plates H, capacity factors Ḱ, selectivity factors α, and peaks
asymmetry, and Resolution RRsR were measured by analyzing different analytes on the new columns using different mobile phase compositions and flow rates. These parameters were measured from analysis of some amino acids such as Histidine, Phenylalanine, Tryptophane, and Tyrosine. The analysis of amino acids using Amberlite – Bromophenol blue column has performed using %10 acetonitrile in water as mobile phase. The average value of plate numbers N was192.085. The plate heights H was averaged 0.089 cm , capacity factors Ḱ were ranged from 1.8 – 2.42. Separation factors α were ranged from 0.86-1.3 , and peak asymmetry value were range from 1.02-1.98 . Resolution Rs were ranged from 0.92-4.15 . R.S.D were ranged from(0.06-0.10)% , recovery were ranged from (95.84-96.88)% , The relative errors were ranged from (-3.12 - -4.64) % . Calibration curves for all analyzed compound were linear from their detection limits to at least 50 ppm with correlation coefficient ranged from 0.9989-0.9996. The detection limits were ranged from 0.05 - 0.25 ppm at signal to noise ratio of three or more.