Specific high performance liquid chromatographic determination of the molecular weight and concentration of hyaluronic acid in complex mixtures by labelled hyaluronate binding proteins

Abstract

A simple High performance liquid chromatographic (HPLC) method for the specific determination of the molecular weight and concentration of hyaluronic acid (HA) in complex mixtures has been developed. Hyaluronate-binding proteins isolated from bovine cartilage labelled by 125I or fluoresceinisothiocyanate were used as specific markers. The specific binding affinities of the markers were compared and were found to have association constants of 1.6 x 10(7) M-1 and 1.2 x 10(7) M-1 respectively. The HA levels and molecular weight distributions can be easily determined in the range 10-500 ng/mL in complex mixtures by the use of markers, molecular sieving HPLC columns and appropriate detectors. It has been demonstrated clearly that the method is useful for the highly specific determination of the parameters in complex biological samples such as serum and synovial fluids and is recommended for clinical applications.

Degradation of high-molecular-weight hyaluronan by hydrogen peroxide in the presence of cupric ions

“…Since H 2 O 2 plus Cu 2+ is an especially efficient system for degradation of high-molecularweight polysaccharides including HA, [21][22][23][24] and since the presence of both hydrogen peroxide and copper ions have been claimed in the rheumatic joint, 25-27 the reaction of H 2 O 2 with CuCl 2 could be of (pathophysiological) importance. Previously, H 2 O 2 was applied as the analytical reagent for the determination of trace transition metal ions using their catalytic effect on hydrogen peroxide decomposition (Eq.…”

Degradation of high-molar-mass hyaluronan by an oxidative system comprising ascorbate, Cu(II), and hydrogen peroxide: Inhibitory action of antiinflammatory drugs—Naproxen and acetylsalicylic acid

“…Changes of η of the HA solution during degradation studies could be of advantage in classifying the efficacy of various substances to act as antioxidants [42][43][44]. In principle, the changes of the dynamic viscosity of the polymer solution can be monitored easily by rotational viscometry, since the solution "flow" behavior is responsive to properties such as the polymer molar mass and molar mass distribution.…”

Determination of sodium hyaluronate in pharmaceutical formulations by HPLC–UV

“…The literature survey shows that several methods [11] , [12] , [13] , [14] , [15] , [16] , [17] , [18] , [19] , [20] , [21] , [22] , [23] , [24] , [25] , [26] , [27] , [28] like enzymatic, Carbopac PA1 chromatography, chemiluminescence, digestion, gas–liquid chromatographic, on-line HPLC/ESI–MS; HPLC UV–vis methods have been reported for the determination of sodium hyaluronate with derivatization. Most of the reported methods are by derivatization, gel permeation chromatography or digestion.…”