Effect of pH additive and column temperature on kinetic performance of two different sub-2 μm stationary phases for ultrafast separation of charged analytes

The performance characteristics of separation were studied for small pharmaceuticals and larger charged molecules (peptides) in various mobile phase conditions on two 5-cm long narrow bore columns packed with 1.7 μm core-shell and totally porous particles respectively. The effect of temperature and pH additives (formic acid, trifluoroacetic acid, ammonium formate, ammonium acetate and ammonia) on column efficiency was investigated through a kinetic study based upon data obtained under gradient elution conditions. Sample peak capacities were calculated and compared in all studied conditions for a sample of ten representative peptides having masses ranging from 500 to 2000 Da. The elevation of temperature was found to be significantly beneficial. The effect of flow-rate on peak shape was also investigated. Ammonium acetate at neutral pH led to the best results in terms of both efficiency and peak capacity. It was found that column performance was strongly dependent on the type of stationary phase, especially in acidic medium.