Intestinal alpha-glucosidase inhibitory activity and toxicological evaluation of Nymphaea stellata flowers extract.
OBJECTIVE
Nymphaea stellata willd. flowers (NSF) are used as a traditional medicine in India and Nepal to treat diabetic disease. Different works have demonstrated that NSF extract showed antihyperglycemic effect on alloxan-induced diabetic rats. In the present work we evaluated in vitro intestinal alpha-glucosidase inhibition as the possible mode of action of NSF extract on suppressing postprandial hyperglycemia for curing diabetic mellitus. In addition, NSF extract was studied to assess its possible acute oral toxicity and genotoxicity.


METHODS
Rat intestinal crude enzyme preparation and Caco-2 monolayer were used to evaluate alpha-glucosidase inhibitory activity of NSF extract. The main alpha-glucosidase inhibitors were detected by HPLC. For acute toxicity test, NSF extract was administered at doses of 2000, 5000 and 10,000 mg/kg body to three groups of 10 ICR mice each, and then clinical symptoms including mortality, clinical sign and gross findings were observed once a day for 14 days. In Ames test, histidine-dependent auxotrophic mutants of Salmonella typhimurium (strains TA97, TA98, TA100, TA102 and TA1535) were used and incubated in the presence and absence of S9 metabolic activation using NSF extract with concentrations of 150-5000 microg/plate. The chromosome aberration test was conducted with Chinese hamster lung (CHL) cells treated with NSF extract at doses of 150-5000 microg/ml in the presence and absence of S9 metabolic activation. In the in vivo mouse micronucleus assay, 9-week-old male and female ICR mice (n=90, 25-30 g) were administered daily by oral gavage at doses of 2.5, 5.0 and 10.0 g/kg body for 1 or 2 days. Bone marrow smears were prepared from each treatment group 24h after last administration and then polychromatic erythrocytes (PCEs) and normochromatic erythrocytes (NCEs) were identified.


RESULTS
NSF extract showed potent rat intestinal alpha-glucosidase inhibitory activity for maltose hydrolysis with ED(50) value of 0.1 mg/ml. In Caco-2 monolayer, alpha-glucosidase activity for the maltose hydrolysis was down-regulated by NSF extract at a concentration of 0.05 mg/well level, showing 74% inhibition compared to the saline treated control. NSF was rich in phenol contents and the main alpha-glucosidase inhibitor, 1,2,3,4,6-penta-O-galloyl-beta-D-glucose, was identified together with two phenolic compounds of gallic acid and corilagin. In acute toxicity test, NSF extract did not produce any toxic signs or deaths and the LD(50) value of this extract could be greater than 10,000 mg/kg body weight. These results of genotoxicity assessment showed that NSF extract did not cause genotoxic effects in Ames test, in the in vitro chromosomal aberration assay and in the in vivo micronucleus assay.


CONCLUSIONS
The current study shows that the extract from Nymphaea stellata flowers exhibits significant intestinal alpha-glucosidase inhibitory activity, without showing any acute toxicity or genotoxicity, which may be useful in suppressing postprandial hyperglycemia in diabetics. The results presented here suggest that the use of NSF in folk medicine as a natural antidiabetic treatment could be safe as well as beneficial.
Huang
Yi-Na
YN
Department of Public Health, Hua Xi Medicinal Center of Sichuan University, Chengdu, PR China.
Zhao
Ying-Lan
YL
Gao
Xiao-Ling
XL
Zhao
Zhi-Feng
ZF
Jing
Zan
Z
Zeng
Wei-Cai
WC
Yang
Rong
R
Peng
Rong
R
Tong
Tao
T
Wang
Long-Feng
LF
Cen
Jia-Qi
JQ
Gao
Hong
H
D000818:Animals; D018938:Caco-2 Cells; D002460:Cell Line; D003106:Colon / Q000187:drug effects* / Q000201:enzymology; D006224:Cricetinae; D003412:Cricetulus; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology* / Q000633:toxicity; D005260:Female; D035264:Flowers; D065089:Glycoside Hydrolase Inhibitors; D006801:Humans; D006868:Hydrolysis; D007928:Lethal Dose 50; D008168:Lung / Q000187:drug effects; D008297:Male; D008320:Maltose / Q000378:metabolism; D051379:Mice; D008813:Mice, Inbred ICR; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology; D030721:Nymphaea / Q000737:chemistry* / Q000633:toxicity; D010636:Phenols / Q000494:pharmacology* / Q000633:toxicity; D010936:Plant Extracts / Q000494:pharmacology* / Q000633:toxicity; D051381:Rats
D004791:Enzyme Inhibitors; D065089:Glycoside Hydrolase Inhibitors; D010636:Phenols; D010936:Plant Extracts; D008320:Maltose