Gy. Szókán

HPLC analysis of anthraquinone derivatives in madder root (Rubia tinctorum) and its cell cultures

Abstract A sensitive and reproducible RP-HPLC method was developed for the characterization of madder root and its cell cultures extracts and for the determination of anthraquinone derivatives as glycosides (ruberythric acid, lucidin primveroside) and aglycones (alizarin, lucidin, purpurin) in them. The fingerprint chromatographic patterns of the extracts were obtained on C18 silica column, using gradient with acetonitrile-water-acidic buffer eluent system. For quantitative measurements, the anthraquinone derivatives were separated by isocratic elution in the same type of mobile phase. The chief components were identified and quantitatively determined. The natural plant and its cell suspension cultures were compared to each other. The preparative fractionation of the extracts was achieved by gel chromatography, HPLC and selective extractions with solvent series, solid-phase extraction techniques and SFE.

Comparison of thin-layer chromatography and overpressured layer chromatographic techniques for the separation of ascorbigen and 1'-methylascorbigen

Abstract Simple and efficient methods are described for the separation of biologically active ascorbigen and 1′-methylascorbigen by normal-phase TLC, conventional as well as new personal overpressured layer chromatographic (OPLC) systems using the same eluent. Results obtained with these techniques were compared on the basis of the separation parameters with special emphasis on the presentation of the advantages of the automated personal OPLC system. For the characterization of the personal OPLC system dependence of average plate height and resolution on the external pressure, front velocity and development distance was examined on TLC and HPTLC sorbent layers using ascorbigen and 1′-methylascorbigen as model compounds.

Liquid Chromatographic Analysis and Separation of Polypeptide Components from Honey Bee Venoms

Abstract Reversed-phase HPLC on different columns with acetonitrile-water-trifluoroacetic acid eluent system was used to characterize honey bee venoms and to separate and determine quantitatively its peptide components.

Antitumor effect of lysine-isopeptides

Isopeptides (ε-peptides) of lysine, with a given Mw and low polydispersity (10–400 units), were synthesized to study the relationship between their chemical structure and biological effect. The designed compounds were of high purity, low polydispersity and high stereochemical purity. The effect of the compounds was tested on a human erythroleukemia cell line (K-562) and on four transplantable mouse tumors (L1210 lymphoid leukemia, P38 macrophage derived tumor, Ehrlich ascites carcinoma, Lewis lung tumor /LLT/). In case of the L1210 and P388 tumors and the Ehrlich carcinoma, survival of the animals was used as an indicator of the effect. In case of the Lewis lung tumor, the number and size of metastases in the lung and/or liver of treated and untreated mice were used as indicators. The polymers of polymerisation degree 80–120 (Mw 10.2–15.4 KD) showed the strongest antiproliferative effect both on K562 cells and the tumors growing in vivo. This effect was manifest with a significantly higher survival rate as compared to the control (L1210, P38, Ehrlich ascites), furthermore, by a decrease in the number and size of liver and lung metastases (LLT).

HPLC enantioseparation of amino acids and their protected derivatives used in peptide synthesis with pre-column chiral derivatization

SummaryIndirect chiral separation methods based on enantiomeric derivatizations were developed in order to monitor optical purity of uncoded amino acids and a new series of amino acid derivatives. Marfeys reagent was used for derivatization of amino groups: whilst boxyl groups were derivatised with (1R, 2R)-or (1S, 2S)-2-amino-1(4-nitrophenyl)-1,3-propanediol reagents were used, respectively. The separations of diastereomeric derivatives formed via derivatization were optimized in RP-HPLC and NP-HPLC systems.

Structure determination and synthesis of lysine isopeptides influencing on cell proliferation

The clavicepamines are lysine‐rich basic proteins isolated from saprophytic culture of ergot (Claviceps purpurea), having human pharmacological importance.

HPLC determination of enantiomeric purity of protected amino acid derivatives used in peptide synthesis

Abstract An improved RP-HPLC method on ODS-Hypersil column with pre-column derivatization with Marfeys reagent were used to monitor racemization in N-, C- and/or side-chain protected amino acid derivatives by separation of a series of new diastereoisomeric Marfeys compounds. Chromatographic samples were obtained by partial deprotection of different starting materials. In a simple two-step procedure (deprotection and derivatization) the compounds of amino acids formed stable diastereomeric derivatives having facile resolutions.

ON-LINE CD DETECTION IN CHIRAL SEPARATION OF SPIRO-λ4-SULFANES

Enantiomers of chiral spiro-λ4-sulfanes with equatorial diaryl and axial bis(acyloxy), (alkoxy)-(acyloxy) or (acylamino)-(acyloxy) S-substituents were separated on a Kromasil-based chiral sorbent, namely on O,O′-bis(3,5-dimethylbenzoyl)-N,N′- diallyl-L-tartardiamide silica CSP, by using n-hexane (or n-heptane) – dioxane (or 2-propanol) isocratically mixed mobile phases. Direct chiral separation was monitored by a home-made HPLC-CD system consisting of a circular dichroism (CD) spectrophotometer (Jobin-Yvon Model III Dicrograph) and a high performance liquid chromatograph. The system was used in conjunction with HPLC-UV to study the chemical and stereochemical purity of the samples. By the stopped-flow technique CD spectra were measured in the 350–230 nm spectral range. Monitoring at a selected wavelength enabled high-sensitivity detection. Racemic spiro-λ4-sulfanes were also separated into pure enantiomers on preparative scale. Regardless of the structural type of diaryl-spiro-λ4-sulfanes, the first-elute...

Chromatographic studies on the racemization of thiopeptides

Abstract It was found by chromatographic, CD and NMR methods, that the thionation of piperazine-2,5-diones [cyclo(Aaa1-Aaa2) → cyclo (Aaat 1-Aaat 2 (Aaa=-NH-CHR-CO-; Aaat=-NH-CHR-CS-)] or piperazine-2,5-onthiones [cyclo(Aaat 1-Aaa2) → cyclo (Aaat-Aaat)] and, occasionally, even the spontaneous cyclization of endo-thiodipeptide esters [H-Aaat-Aaa-OR] result in enantiomeric (Aaa or Aaa=Gly) or diastereomeric mixtures of piperazine monothiones or dithiones. The diastereoisomers were separated by semipreparative HPLC and their quantitative product distribution was determined by an optimized HPLC method on Hypersil-silica column with CH2Cl2-EtOAc eluent mixtures. Isocratic RP-HPLC on ODS-Hypersil column and pre-column derivatization with 1-flu-oro-2,4-dinitrophenyl-5-L-alanine amide (Marfeys reagent) were used to monitor the racemization of Ala and Pro residues and to determine the ratio of enantiomers. Thionation of urethane protected dipeptide esters or dethionation of the corresponding endoth-iodipeptide de...

RP-HPLC assay for 1,2-5,6-dianhydro-3,4-disuccinyl-galactitol and its metabolites in blood plasma and liver

SummaryA method involving pre-column derivatization and HPLC assay is described for measuring submicrogram quantities of 1,2-5,6-dianhydro-3,4-disuccinyl-galactitol (1,2-5,6-dianhydro-3,4-bis(carboxypropionyl)-galactitol), an effective cytostatic drug and its metabolites in blood plasma and liver homogenate. The substance and its metabolites were derivatized with sodium pentamethylene-dithiocarbamate to form different bis(dithiocarbamoyl) esters, which can be detected by UV absorbance at 254 and 280 nm. The directly derivatized products were then extracted into CHCl3, and after sample preparation resolved by RP-HPLC on SAS-Hypersil column.