I. Yu. Ponedel’kina

Modification of Hyaluronic Acid with Aromatic Amino Acids

Hyaluronic acid was modified with aromatic amino acids (5-aminosalicylic, 4-aminosalicylic, anthranilic, and p-aminobenzoic) in the presence of 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide. The modified glycans contained 9–43% of arylamide groups and 10–33% of isoureidocarbonyl groups depending on the nature of the amino acid. Reduction with sodium borohydride allowed the conversion of isoureidocarbonyl groups into hydroxymethyl groups.

Conjugation of Chondroitin Sulfates with Amines

Conjugation of chondroitin sulfates with pharmacologically important amines in a water medium in the presence of 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide was studied. Conjugates with amide and isoureidocarbonyl groups were synthesized. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2008, vol. 34, no. 5; see also http://www.maik.ru

Synthesis of glucosaminoglycans hydroxamic acids

The hydroxamic functional group –C(=O)NHOH is introduced into polysaccharides in order to prepare water-soluble antioxidants [1–3], matrix metalloproteinase inhibitors [2, 4], and iron chelators [5, 6] based on them. The reaction of natural and synthetic alkyl esters of polysaccharides and hydroxylamine base is usually used to synthesize the hydroxamic acids [2, 5, 7]. For this, the content of hydroxamic units is <25% [2]. We showed earlier that the reaction of glycosaminoglycans (GAG) with 1-ethyl-3-[3(dimethylamino)propyl]carbonyldiimide (CDI) formed readily O-(glycosaminoglycanoyl)isoureas that exhibited ester properties [8–10]. Therefore, we studied the reaction of the corresponding O-acylisoureas with alkaline NH2OH in aqueous and alcoholic solution in order to prepare hydroxamic acids of hyaluronic acid (1), dermatan sulfate (2), and heparin (3). The action of a five-molar excess of CDI on GAG 1, 2, and 3 synthesized the corresponding O-acylisoureas 4, 5, and 6. Treatment with an alkaline aqueous solution of NH2OH converted them into hydroxamic acids 7, 8, and 9, respectively (Scheme 1). The appearance in IR spectra of 7–9 of absorption bands at 1552–1559 cm–1 that were characteristic of N–H stretching vibrations in –C(=O)NHOH [7] and the formation of iron–hydroxamate complexes with an absorption maximum at 520 nm [11] were consistent with the formation of hydroxamic units in the GAG structures.

Conjugates of hyaluronic acid and chondroitin sulfates with 4-aminoantipyrine and their analgesic properties

Heteropolysaccharides of the glucosaminoglycan class, e.g. hyaluronic acid (HA) and chondroitin sulfates (CS), are known for their reparative and regenerative properties, on the basis of which several drugs for treating diseases of the musculoskeletal system were developed [1–3]. HA and CS stimulate regeneration of cartilage and bone tissue and exhibit antiinflammatory and analgesic activity. High-molecular-weight HA is the principal component in the sinovial fluid replacements Sinokrom, Sinvisk, etc. These are highly viscous hydrogels for intra-joint administration that act as a lubricant for painful joints [2]. The active substance CS in chondro-protector drugs (Chondroxide etc.) normalizes the exchange of cartilage tissue substances including phosphorus and calcium, decreases the pain level, and increases the flexibility of afflicted joints [3, 4]. In order to enhance the anti-inflammatory and analgesic activity of HA (1) and CS [2, a 3:2 mixture of CS-4 and CS-6], we incorporated into their structures 1-phenyl-2,3-dimethyl-4-aminopyrazol-5-one (4-aminoantipyrine) (3). The corresponding conjugates 4 and 5 (Scheme 1) were synthesized by the reaction of glucosaminoglycans 1 and 2 with amine 3 through the action of 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (CDI) [5].

Oxidation of dermatan sulfate with a NaOCl-NaBr-2,2,6,6-tetramethylpiperidine-1-oxyl reagent in an aqueous medium

The oxidation of primary hydroxyl groups in dermatan sulfate with the NaOCl-NaBr-2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) reagent in an aqueous alkaline medium was carried out for the first time. Modified dermatan sulfates containing hydrated aldehyde (15–50%) and carboxyl (25–100%) groups were obtained.

Acid glycosaminoglycans and their chemical modification

The modern state of chemical modification of hyaluronic acid, chondroitin sulfates, and heparin is considered, and the possible application of modified glycosaminoglycans as potential drugs is discussed.

The in vitro resistance of oxidized hyaluronic acid to testicular hyaluronidase.

Hyaluronic acid belongs to the class of acid glycosaminoglycans, heteropolysaccharides with a linear structure. This is the key compound of the extracellular matrix that performs important functions in the organism. It is composed of repeated units represented by glucuronic acid and N -acetylD -glucosamine. The interest to the derivatives of hyaluronic acid displaying a decreased enzymatic biodegradability has increased recently due to their potential as controlled release drugs [1–3]. We have found that the oxidation of hydroxymethyl groups into carboxyl groups renders hyaluronic acid resistant to testicular hyaluronidase in vitro. Hyaluronic acid ( I ) was obtained by alkaline extraction from the umbilical cord of newborns and purified by anion exchange chromatography on DEAE cellulose as described in [4]. Modified hyaluronic acid II was obtained by treating hyaluronic acid I with sodium hypochlorite in the presence of sodium bromide and a catalytic amount of 2,2,6,6-tetramethylpiperidine-1oxyl (TEMPO) in water [5]:

Chemical modification of heparin

Heparin was modified at carboxyl groups by reaction with several pharmacologically important amino-containing compounds in aqueous medium in the presence of 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide. In dependence on the nature of the amine and the ratio of reagents, conjugates containing 36–100% amide and 0–25% isoureidocarbonyl groups were synthesized. Isoureidoarylamide groups are present, along with amide moieties, in the products of heparin modification by hydroxyl-containing aromatic amines. The conjugate of heparin with p-aminobenzoic acid contained oligomeric arylamide.

Decomposition of sodium hypochlorite in an aqueous solution in the presence of 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) nitroxyl radical as a catalyst

Nitroxyl radical TEMPO is found to catalyze the decomposition of sodium hypochlorite in an aqueous-alkali medium. The mechanism of NaOCl decomposition to form ClO· radical is proposed which involves protonated TEMPO, oxoammonium salt TEMPO+, and hydroxyl radical.

Synthesis of conjugates of hyaluronic and nicotinic acids

Conjugates with nicotinic acid of hyaluronic acid carboxylic and hydroxyl groups were synthesized and exhibited polyampholyte properties.