ИСТИНА

The design of novel mixed-mode stationary phases is a current direction of development in the field of HPLC. In their structure there are various functional groups that make it possible to use such separation materials in different chromatography modes: reverse phase (RP HPLC), ion-exchange (IEC) and hydrophilic interaction liquid (HILIC) chromatography. Majority of mixed-mode stationary phases are based on silica, which is stable in a limited pH range (from 2 to 8). As a result, such phases can’t be used with strongly acidic or strongly basic mobile phases. A promising approach is the transition from silica to a copolymer of styrene and divinylbenzene (PS–DVB) with a high degree of cross-linking, which is stable over a whole pH range and is compatible with organic solvents. One of the types of mixed-mode stationary phases for HPLC is zwitterionic stationary phases containing both positively and negatively charged groups. These stationary phases are highly hydrophilic, biocompatible, and can be used for simultaneous separation of anions and cations. In this work, mixed-mode zwitterionic stationary phases based on modified PS–DVB were synthesized. The modification was carried out in two ways: acylation with reductive amination with methylamine and epoxidation followed by amination with methylamine. Polyelectrolyte chains were prepared from 1,4-butanediol diglycidyl ether (1,4-BDDGE) and iminodiacetic acid, as well as from 1,4- BDDGE and methyl glycine. Additionally, positively charged chains formed from 1,4-BDDGE and dimethylamine were sequentially grafted to form an external functional anion-exchanging layer. The synthesized resins were investigated in three retention modes: RP HPLC, IEC and HILIC. It was noted that grafting of positively charged chains leads to a hydrophilicity increase of the stationary phases in the HILIC mode, but an efficiency decrease in the RP HPLC mode. A mixture of 14 anions containing standard anions, lactate, bromate and anions of 5 phosphonic acids was separated in IEC mode in 32 min in a gradient elution mode. In the RP HPLC mode, the ability to separate a mixture of 7 alkylbenzenes in 20 min (with an efficiency of up to 15,000 tp/m), 3 fat-soluble vitamins in 15 min (with an efficiency of up to 3,000 tp/m), as well as a mixture of phenol and 5 of its derivatives in 15 min (with efficiency up to 9,000 tp/m) was shown. The stationary phases in HILIC mode made it possible to separate 5 water-soluble vitamins in 12 min in isocratic elution mode (with an efficiency of up to 14,500 tp/m) and 6 water-soluble vitamins in 9 min in gradient elution mode (with an efficiency of up to 13,500 tp/m), 4 nitrogenous bases in 3.5 min (with an efficiency of up to 10,000 tp/m) and 10 amino acids in 24 min (with efficiency up to 21,000 tp/m) in isocratic elution mode. This work was supported by the Russian Science Foundation (project No 23-73-01145)