Preparation of Neomytilan, a Bioglycan from the Mussel Crenomytilus grayanus , and its Immunostimulating Activity
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PREPARATION OF NEOMYTILAN, A BIOGLYCAN FROM THE MUSSEL Crenomytilus grayanus, AND ITS IMMUNOSTIMULATING ACTIVITY
I. V. Chikalovets,1,2* V. I. Molchanova,1 O. V. Chernikov,1 and P. A. Lukcyanov1
UDC 577.114:592:594.124
Bioglycans isolated from the Far-East mussel Crenomytilus grayanus were compared analytically. It was found that neomytilan, a bioglycan with reduced protein content, and the previously isolated mytilan stimulated phagocytosis of murine peritoneal macrophages in vitro and influenced the synthesis of pro-inflammatory cytokines TNF-D, IL-6, and IFN-J. Keywords: bioglycans, mussel Crenomytilus grayanus, cytokines, immunostimulating activity. Marine biological resources have recently been used more and more often as sources for preparing new physiologically active compounds. We isolated earlier from the mantle of the mussel Crenomytilus grayanus a bioglycan that was called mytilan, a high-molecular-weight (>2 MDa) carbohydrate-protein complex, the components of which were bonded through a strong non-covalent bond. The carbohydrate component (90–95%) of mytilan was a glycogen-like 1,6;1,4-D-D-glucan with a high degree of branching in the carbohydrate chain and a small number of 1,2- and 1,3-bound D-glucose residues at the branching points [1]. Much literature has been published on the physiological activity of mytilan in animal experiments [2–6]. It has been used for a long time as a component of cosmetic creams that are manufactured by several Russian enterprises. However, a drawback of this preparation was the comparatively high (3–8%) content in it of potentially allergenic proteins. Therefore, we isolated the new preparation neomytilan with reduced protein content. The goal of the present work was to compare the immunomodulating properties of these bioglycans. Two bioglycans were isolated from broth obtained by heat treatment of Far-East mussels using a combination of ultrafiltration, precipitation, and ion-exchange chromatography. These were the previously known mytilan and the new neomytilan. The carbohydrate and protein contents in neomytilan were 99 and 0.62%, respectively, whereas mytilan contained 8–10% protein that was present as a D-galactose-binding lectin [7]. The identification of the carbohydrate components of these bioglycans was confirmed by GC, gel-filtration over a column of Sepharose CL-4B, 13C NMR, and optical rotation. According to results of gel-permeation chromatography over Sepharose CL-4B, neomytilan was similar to mytilan and was a high-molecular-weight polysaccharide of molecular weight >2 MDa (data not presented). The only monosaccharide in neomytilan hydrolysate that was identified by GC was D-glucose, as was shown previously for mytilan. 13C NMR data showed that these bioglycans were identical to a D-glucan (glycogen) with D-1,6- and 1,4-D-glucoside bonds. Resonances in the spectrum with chemical shifts 100.3, 77.7, and 61.04 ppm were assigned to atoms C-1, C-4, and C-6 of D-1,4-bound monosaccharide residues; those at 100.3 and 69.9 ppm, to C-1 and C-6 atoms of monosacchari
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