Potassium hyaluronate

Mitra et al.AS have also re-refined the structures of the regular (26) and relaxed 4-fold sodium hyaluronate (27) helices for direct comparison with their own analysis of the relaxed 4-fold helix of potassium hyaluronate (29). Within experimental error, their final models strongly support the original results, which have been described here. 

Fig. 30. — Packing arrangement of 4-fold antiparallel double helices of potassium hyaluronate (32). (a) Stereo view of a unit cell approximately normal to the line of separation of the two helices. The two chains in each duplex, drawn in open and filled bonds for distinction, are linked by not only direct hydrogen bonds, but also water bridges. Inter double-helix hydrogen bonds are mediated between hydroxymethyl and iV-acetyl groups. Potassium ions (crossed circles) at special positions have only a passive role in the association of hyaluronate chains.

The X-ray diffraction patterns obtained from potassium hyaluronate films prepared from solutions of pH 3.0-4.0 were representative of the patterns from Rb+, Cs+, and NH4 forms. At 90-98% r.h., the unit cell is tetragonal, with a = b = 1.714 nm, and c = 3.28 nm. Each chain is a 4(— 0.82) helix, and two such chains fonn an antiparallel-stranded double-helix. There are two double helices in the unit cell, and the space group is 14,22. There are no intramolecular hydrogen-bonds. 

Figure 1 Example of an x-ray fiber diffraction pattern suitable for analysis using AXIS. The spots are discrete, there is no layer line streaking, and the background scatter may be seen clearly. The sample (potassium hyaluronate) was slightly tilted out of the plane perpendicular to the x-ray beam.

The current method for the hyaluronidase assay described in the United States Pharmacopeia (USP) [132] is based on the inability of hydrolyzed potassium hyaluronate to form a complex precipitate with proteins from added serum, reflected in a decreased turbidity of the reaction mixture (measured after 30 min). The method is, from the enzymological point of view, not well defined since it does not actually evaluate the kinetics of the hydrolysis of the substrate. An assay with end-point determination is only valid if the reaction rate does not change during this reaction time. We found that only with the two lowest test concentrations (0.15 and 0.3 IU) was this condition fulfilled, while with the three higher test concentrations the reaction is not linear. Commercially available hyaluronates can be contaminated with chondroitin sulfates. They are more acidic than hyaluronic acid itself and hence can form better protein complexes and influence the turbidity. In a suitability test of the USP [133], the substrate must pass both an inhibitor content test and a turbidity-production test. The assumption is made that... 

Binding parameters for potassium hyaluronate solutions in phosphate buffers calculated by Equs. (2), (3), (4), and (5) from data in Ref. 2. 

Figure 1. Binding parameters for potassium hyaluronate in phosphate buffer (pKa = 7.2) based on Refs. 1 and 2 and demonstrating conformational change. Dotted curve is...

Hyaiuronic Acid. See Hyaluronic acid Hyaluronic acid, dimethylsilylene ester. See Dimethylsilanol hyaluronate Hyaluronic acid, potassium salt. See Potassium hyaluronate... 

Potassium hexanitrocobaltate III. See Cobalt potassium nitrite Potassium hyaluronate CAS 31799-91-4... 

Polyvinyl alcohol Polyvinyl butyral Polyvinyl imidazolinium acetate Polyvinyl laurate Polyvinyl methyl ether Potassium acetate Potassium carrageenan Potassium hyaluronate PPG-12/SMDI copolymer PPG-26/TDI copolymer Procollagen Propylene glycol diundecanoate PVM/MA copolymer PVM/MA copolymer, butyl ester PVM/MA copolymer, ethyl ester PVM/MA copolymer, isopropyl ester PVM/MA decadiene crosspolymer PVP... 

New experimental evidence has shown that glass capillaries filled with potassium hyaluronate solutions do not contain electrochemical activity gradients between the ends of the capillaries. Depending on the method of extraction, hyaluronic acid from rabbit skin has a molecular weight between 1.7 x 10 and 1.3 x 10. ... 

The method of Byers and co-workers (17) is especially suited for the isolation of hyaluronate from human cords on a large scale because of its simplicity and good yields. Partially acetone dried cords are digested with trypsin, the digest is dialyzed and filtered, and potassium hyaluronate is precipitated by McClean s method. It must be pointed out, however, that the material obtained by this method contains approximately 30% of chondroitin sulfuric acid. Hyaluronate has also been extracted with Hayem s solution and precipitated from the extract with acetone (57). This method leads to a product of very high salt content and cannot be recommended. 

Several methods allow the preparation of purified hyaluronate from umbilical cords in a matter of a day or two (88,188). The procedure of Jensen (88) is a model of simplicity. Minced, alcohol-dried cords are washed with 90% acetic acid, and extracted with water at pH 7. Potassium hyaluronate is obtained from the extract by two successive alcohol precipitations according to McClean. Preparations of high viscosity are obtained in good yields (89). 

Substrate Solution. Purified sodium or potassium hyaluronate is dissolved in the pH 6 buffer to a concentration of 0.4 mg. per milliliter. This solution is stored under... 

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