Aldose Reductase Inhibitors from Stellera chamaejasme.

Abstract

Six biflavonoids (chamaejasmin, 7-methoxyneochamaejasmin, 7-methoxychamaejasmin, chamaejasmenin B, chamaechromone, wikstrol A) from Stellera chamaejasme. L. have been assayed to show good aldose reductase inhibiting activity. These compounds may be useful to improve diabetic complications such as neuropathy, cataract formation, neophropathy, and retinopathy.

Keywords:

• Aldose reductase inhibitors
• biflavonoids
• diabetic complications
• Stellera chamaejasme. L

Introduction

Stellera chamaejasme. L. (Thymelaeaceae) is widespread in the north of China. It has traditionally been used as an herbal remedy for scabies and tinea. It has been found to possess obvious antitumor and antiviral, especially anti-HIV, activities (Feng et al., 1995; Ikekawa & Ikekawa, 1996; Yoshida et al., 1996; Endo et al., 1998). During our previous study, several biflavonoids have been isolated from this plant (Feng et al., 200120022003).

In order to study further the bioactivities of these compounds, inhibiting action against aldose reductase, which plays an important role in the etiology of the complications of diabetes, has been assayed. Six biflavonoids, chamaejasmin (1), 7-methoxyneochamaejasmin (2), 7-methoxychamaejasmin (3), chamaejasmenin B (4), chamaechromone (5) and wikstrol A (6) (Fig. 1), showed good inhibiting activity. It has been suggested that diabetic complications in experimental animals can be improved by the inhibitors of aldose reductase (Beyer-Measr & Crue, 1985), as well as in clinical trails (Handelsman & Turtle, 1981). Thus, these compounds may be useful to improve diabetic complications such as neuropathy, cataract formation, neophropathy, and retinopathy.

Figure 1. Structures of biflavonoids from S. chamaejasme. L.

Materials and Methods

General experimental procedures

The centrifuge used was a Hettich EBR 12R. Incubation was done in a DNP-9052 incubator (Shanghai Jinghong Experimental Apparatus Co. Ltd., Shanghai, China). The fluorescence was measured on a Type 650-10 spectrofluorometer (Hitachi, Tokyo, Japan).

Plant material, extraction, and isolation

For plant material, extraction, and isolation, please refer to the literature (Feng et al., 200120022003).

Assay of aldose reductase inhibiting activity

Lenses of Wistar rats were homogenized in 135 mM Na, K-phosphate buffer (pH 7.0) containing 10 mM 2-mercaptoethanol and centrifuged at 100,000 × g for 30 min. The supernatant fluid was used as the enzyme fraction. The incubation mixture contained 135 mM Na, K-phosphate buffer (pH 7.0), 100 mM Li₂SO₄, 0.03 mM NADPH, 1 mM L-glyceraldehyde as a substrate, and 100 µl enzyme fraction, with or without 25 µl of sample solution in a total volume of 0.5 ml. Test samples were dissolved in DMSO. The reaction was initiated by the addition of NADPH at 30°C. After 30 min of incubation, the reaction was stopped by the addition of 150 µl of 0.5 M HCl. Then, 0.5 ml of 6 M NaOH containing 10 mM imidazole was added, and the solution was heated at 60°C for 20 min to convert NADP to a fluorescent product. The fluorescence was measured at room temperature with an excitation wavelength of 360 nm and an emission wavelength of 460 nm.

Results and Discussion

Six biflavonoids showed good inhibiting action against aldose reductase. From the IC₅₀ values in Table 1, it seemed that biflavanones 1, 2, 3, and 4 had better activity compared with biflavonoids 5 and 6, which belong to chromone derivatives and flavan biflavonoids respectively. We concluded from these data that the more OH these four biflavanones (1, 2, 3, 4) possess, the better activity they show.

Table 1.. Inhibiting activity against aldose reductase of biflavonoids from S. chamaejasme. L.

Name and no.		IC50 (µM)
Chamaejasmin	1	1.8
7-methoxyneochamaejasmin	2	2.9
7-methoxychamaejasmin	3	4.1
chamaejasmenin B	4	5.9
chamaechromone	5	7.4
wikstrol A	6	27.7

Further animal experiments need to be done to prove whether these compounds could improve diabetic complications in vivo..