Machtejevas, Egidijus

In this work, monolithic silica columns with the C4, C8, and C18 chemistry and having various macropore diameters and two different mesopore diameters are studied to access the differences in the column efficiency under isocratic elution conditions and the resolution of selected peptide pairs under reversed-phase gradient elution conditions for the separation of peptides and proteins. The columns with the pore structural characteristics that provided the most efficient separations are then employed to optimize the conditions of a gradient separation of a model mixture of peptides and proteins based on surface chemistry, gradient time, volumetric flow rate, and acetonitrile concentration. Both the mesopore and macropore diameters of the monolithic column are decisive for the column efficiency. As the diameter of the through-pores decreases, the column efficiency increases. The large set of mesopores studied with a nominal diameter of 25 nm provided the most efficient column performance. The efficiency of the monolithic silica columns increase with decreasing n-alkyl chain length in the sequence of C18 < C8 < C4. The resolution of proteins and peptides by reversed-phase gradient liquid chromatography on n-octadecyl, n-octyl, and n-butyl bonded monolithic silica columns is optimized. The results obtained imply the use of acetonitrile concentration gradient up to 75% for n-octadecyl and n-octyl bonded monolithic silica columns, and the use of acetonitrile concentration gradient up to 85% for n-butyl bonded monolithic silica columns. With the respect to the gradient times and flow rates, the optimum conditions are the best with n-octyl and n-butyl bonded monolithic silica columns, where the range of optimum gradient times is up to 30 min and mobile phase flow rates in the range of 0.5–1 ml/min [...].

Chromatographic enantiomer separations of different oxazepine indole derivatives were performed using a molecularly imprinted polymer. A 5aR,12R,13S-trans-6,6-dimethyl-12,13-dihydro-6H-5a,13-methanoindolo[2,1-b][1,3] naphthoxazepine-12-carboxamide enantiomer derivative was used as a template and the resultant polymer has shown enantiomer recognition for series of template related compounds. The mechanistic description of the chiral discrimination process is scrutinised, comparing the discrimination between the different conformations and substituents of the oxazepine indoles.

Continuous beds with immobilised human serum albumin as a chiral selector were synthesised in fused-silica capillaries in a modified fashion using additives during the protein allylation and polymerisation steps. Acetyl salicylic acid and L-tryptophan were used as additives to interact with the active centres that are responsible for chiral recognition, thus preserving them during the protein coupling procedure. The beds synthesised under modified conditions exhibit higher resolutions and enantioselectivities in capillary liquid chromatography, especially when L-tryptophan was used as an additive. Monolithic adsorbent synthesised with L-tryptophan possesses much higher efficiency when capillary electrochromatography was employed.

The continuous bed technique with its attractive features, such as fritless design, one-step in situ synthesis, low back pressure and no need for pressurising the electrode vessels to suppress bubble formation was applied to form polyrotaxane-based stationary phases for capillary electrochromatography (CEC). Rotaxanes are synthesized from two classes of substances, namely linear reactive monomers and inert cyclic compounds. Upon polymerisation, a gel forms with the cyclic molecules mechanically immobilized (see Fig. 1). We have employed this simple approach, using charged derivatives of cyclodextrins in order to introduce charged groups into continuous beds and thus render them appropriate for electrochromatography. The self-assembly of supramolecular structures to form rotaxanes during the synthesis of the continuous beds is treated. The electroosmotic and chromatographic properties of the various polyrotaxane-based stationary phases synthesized are discussed, as well as the synthesis of the continuous beds, including how to affect their porosity and its influence on the efficiency of the electrokinetic separation. The applicability of the rotaxane-based continuous bed is demonstrated by separation of model compounds by reversed- and normal-phase chromatography. A separation of enantiomers is also presented. This experiment is of particular interest because it indicates that the interaction with the cavity of β-cyclodextrin (β-CD) is not a fundamental requirement for enantioseparations.

Ištisiniai sorbentai su imobilizuotu žmogaus serumo albuminu kaip chiraliniu selektoriumi buvo susintetinti lydyto kvarco kapiliaruose (vidinis skersmuo 250 mm, išorinis skersmuo 375 mm), panaudojant vienpakopę sintezę iš akrilinių komonomerų ir alilinto baltymo. L-kinurenino ir L-triptofano sąveika su susintetinta HSA stacionaria faze stipriai kito priklausomai nuo mobilios fazės joninės jėgos, metanolio koncentracijos, pH ir injektuoto bandinio tūrio / koncentracijos bei temperatūros. Sulaikymo faktoriaus pokyčiai L-enantiomerui buvo visais atvejais ryškesni. Buvo nustatyta, kad chromatografinių sąlygų pokyčiai gali įtakoti analičių sąveiką su baltymo stacionaria faze. Buvo tirta mobilios fazės linijinio greičio įtaka efektyvumui, selektyvumui ir skiriamajai gebai skirstant enantiomerus. Aktyvių centrų žmogaus serumo albumino kolonėlėje tankiui nustatyti buvo naudotas frontalinės analizės metodas. Atsikartojamumo tyrimai atskleidė enantioselektyvumo ir skiriamosios gebos didėjimą per 400–600 eksploatacijos valandų ir vėlesnę šių parametrų stabilizaciją.

A chiral ligand-exchange phase for capillary electrochromatography based on continuous bed technology was developed. The chiral stationary phase is prepared by a one-step in situ copolymerization procedure using methacrylamide, piperazine diacrylamide, vinylsulfonic acid and N-(2-hydroxy-3-allyloxypropyl)-L-4-hydroxyproline. These chiral continuous beds are inexpensive and easy to prepare. They also have several advantages over silica-based packed capillaries. Since the bed is covalently attached to the capillary wall, no frit is required. The applicability of this new approach to the chiral separation of underivatized amino acids is demonstrated.