In order to identify the specific antigens for pine wood nematode (PWN), we confirmed that one of the genes commonly found in the transcriptome, proteome and secretory proteins of PWN belonged to the Aldose Reductase (AR) family protein. 36.5 kDa PWN-AR1 was expressed and purified using Baculovirus Expression System. Total 1,546 hybridoma fusion library was generated and screened for specificity to PWN-AR1 by Enzyme Linked Immunosorbent assay (ELISA). Nine clones showed strong immunoreactivity to PWN-AR1 were limited-diluted. Total 864 limited-diluted clones were further screened using PWN-AR1 by ELISA and 34 monoclonal antibody (Mab) clones were selected. 34 Mab clones were further screened using PWN extracts and a standard PWN-infected pine tree extract by ELISA. Finally nine clones were selected and their immunoreactivities to 4 different nematodes were examined by ELISA. Seven clones pecifically recognized PWN while two clones recognized 4 nematodes. Our data suggested that PWN-AR1 is a PWN secretory enzyme while PWN is invading pine trees, Thus, PWN-AR1-Mabs could be used to develop diagnosis tools for PWN and its infected pine trees. (This work was supported by National Institute of Forest Science)

To evaluate therapeutic efficacy of aldose reductase (AR) inhibition, the leaves of Aster yomena were tested for inhibition of AR. The stepwise polarity fractions of A. yomena were tested. Among them, the n-BuOH fraction showed highest activities against AR inhibition. Our results demonstrated that A. yomena could be a worthy natural source source for curing against diabetic complications.

To evaluate therapeutic efficacy of aldose reductase (AR) inhibition, the seeds of Perilla frutescens var. japonica (PF) were tested for inhibition of AR. The stepwise polarity fractions of PF were tested. The CH2Cl2 and EtOAc fractions showed highest activities (IC50 5.47 and 3.63 μg/mL, respectively). Compounds 1-5 were isolated from the CH2Cl2 and EtOAc fractions by silica-gel and LH-20 Sephadex. Their structures were elucidated as β-sitosterol (1), diosmetin (2), kaempferol (3), luteolin (4), and apigenin (5). Compounds 1-5 were exhibited AR inhibitory. Among them, luteolin (4) and apigenin (5) exhibited AR inhibitory activities (IC50 0.40 and 1.10 μg/mL, respectively). Our results demonstrated that PF could be a worthy natural source for curing against diabetic complications.

We evaluated the inhibitory effects of extracts and components of Geranium thunbergii on aldose reductase (AR) and galactitol formation in rat lenses with high levels of galactose as a part of our ongoing search of natural sources for therapeutic and preventive agents for diabetic complications. The inhibitory effects of water, methanol and ethanol extracts of G. thunbergii on rat lens AR (RLAR) were determined. Comparing inhibitory effects of various solvent extracts, ethanol extract showed RLAR inhibitory activity (IC50 values, 5.24 and 6.39μg/ml, respectively). The ethanol extract was fractionated to chloroform, ethyl acetate and water. Of these, the ethyl acetate fraction from ethanol extract of G. thunbergii exhibited RLAR inhibitory activity (IC50 value, 2.64μg/ml). In order to identify the bioactive components of ethyl acetate soluble fraction of ethanol extract from G. thunbergii, eight compounds, namely gallic acid (1), protocatechuic acid (2), p-hydroxybenzoic acid (3), brevifolin carboxylic acid (4), geraniin (5), ellagic acid (6), kaempferol-3-O-arabinofuranosyl-7-O-rhamnopyranoside (7), kaempferitrin (8) were isolated. The isolates were subjected to in vitro bioassays to evaluate their inhibitory activity on RLAR and galactitol formation in rat lenses. The ellagic tannins (5, 6) and flavonoid (7) exhibited strong inhibitory effects on RLAR. Also, these three compounds (5, 6 and 7) suppressed galactitol accumulation in rat lens under high galactose conditions, demonstrating the potential to prevent galactitol accumulation exo vivo. These results suggest that the extracts and components of G. thunbergii are a promising agent in the prevention or treatment of diabetic complications.

Taxifolin-3-O-β-D-xylopyranoside and quercetin-3-O-α-L-rhamnopyranoside were isolated from an EtOAc-soluble extract of the leaves of Chamaecyparis obtuse. Quercetin-3-O-α-L-rhamnopyranoside was found to possess a potent inhibitory activity of human recombinant aldose reductase in vitro, its IC50 value being 11.5 μM. Kinetic analysis showed that quercetin-3-O-α-L-rhamnopyranoside exhibited uncompetitive inhibition against DL-glyceraldehyde. Also, quercetin-3-O-α-L-rhamnopyranoside suppresses sorbitol accumulation in rat lens under high glucose conditions, demonstrating the potential to prevent sorbitol accumulation in vivo. These results suggest that this compound may be a promising agent in the prevention or treatment of diabetic complications.