Hypoglycaemic Activity of Four Plants Used inChilean Popular Medicine

I. Lemus,1 R. Garcı́a,2* E. Delvillar and G. Knop 3

1Department of Pharmacological and Toxicological Chemistry, Faculty of Chemical and Pharmaceutical Sciences, University of Chile, POBox 233, Santiago-10, Santiago, Chile2Faculty of Medicine, University of Chile, Av. Independencia 1027, Santiago-10, Chile3Laboratorio Homeopa´tico Knop, Av. Industrial 1198, Quilpue´, Chile

The hypoglycaemic activity of a 20% dried leaf infusion ofBauhinia candicansBenth. (Leguminosae),Galega officinalisL. (Leguminosae),Morus alba L. (Moraceae) and Rubus ulmifolius Schott. (Rosaceae),used for diabetes in Chilean popular medicine, was evaluated in alloxan and streptozotocin induced hy-perglycaemic rats.

In normal rats the different infusions did not modify significantly the glycaemia in the period studied, butin diabetic rats different results were observed, depending on the diabetogenic drug used.B. candicansandR. ulmifolius infusions elicited remarkable hypoglycaemic effects in both experimental models.B. candicanspresented a greater decrease of glycaemia in alloxan diabetic rats (39%) andR. ulmifolius showed a similaractivity in both alloxan and streptozotocin diabetic rats (28% and 29%).

Activity-guided fractionation of R. ulmifolius showed that petroleum ether extracts elicited a markedhypoglycaemic effect (35%) in the streptozotocin induced model. Copyright# 1999 John Wiley & Sons,Ltd.

Keywords: Bauhinia candicans; Rubus ulmifolius; Galega officinalis; Morus alba; hypoglycaemic screening; alloxan andstreptozotocin diabetogenic drugs.

INTRODUCTION

The use of plants as therapeutic tools, especially thoseused to relieve chronic pathologies, have had a remark-able role in the popular medicine of different countries.Diabetes mellitus, which does not have a widelysatisfactory therapy and whose incidence is difficult toknow because of the various diagnostic criteria applied, isa disease that probably ranges from 1% to 2% ofthe population, surpassing 120000 patients in Chile(Valiente, 1978).

The present study was carried out to evaluate thehypoglycaemic activity of the main plants used inChilean folk medicine for the treatment of diabetes, intwo models of diabetes induced in rats (Mun˜oz et al.,1981).

MATERIAL AND METHODS

Plant material. The plant material was collected duringthe spring in the Metropolitan Region (Quinta Normaland Quebrada de Macul) and V Region (Quilpue´),keeping a voucher specimen of each plant in theHerbarium of the Chemical and Pharmacy School ofthe University of Chile (Table 1). The leaves of eachplant were shade-dried, and ground before being tested.

Animals. 60-day-old male and female Wistar ratsweighing between 170 and 240 g were used. The ratswere normally fed during the experiment with wateradlibitum.

Experimental design. A group of rats was kept as anormal control group, the rest of the animals were madediabetic by two different models.

(i) Alloxan-induced diabetic model. Rats were anaes-thetized for 2 min with ethyl ether, and then receivedalloxan monohydrate (ALX, Sigma), in a single intrave-nous injection (50 mg/kg), dissolved in physiologicalserum(Lazarow and Palay, 1946).

(ii) Streptozotocin-induced diabetic model.The othergroup of rats received a single intraperitoneal injection(65 mg/kg) of streptozotocin (STZ, Sigma) dissolved in

Table 1. Plant species used as antidiabetic in Chilean folkmedicine

Plant species Popular name Registera Family

Bauhinia candicansBenth.

Pata de vaca 21031 Leguminosae

Galega of®cinalis L. Galega 19784 LeguminosaeMorus alba L. Morera 19783 MoraceaeRubus ulmifolius

Schott.Zarzamora 21025 Rosaceae

a Register number of voucher specimen in the Herbarium ofChemical and Pharmacy School (SQF).

PHYTOTHERAPY RESEARCHPhytother. Res.13, 91–94 (1999)

CCC 0951–418X/99/020091–04 $17.50Copyright# 1999 John Wiley & Sons, Ltd.

* Correspondence to: R. Garcia, Department of Pharmacological andToxicological Chemistry, Faculty of Chemical and Pharmaceutical Sciences,University of Chile, PO Box 233, Santiago-10, Santiago, Chile.Contract/grant sponsor: Dr German Knop.

Accepted 2 June 1998

citrate buffer 0.05M, pH 4, according to Junodet al.,(1969).

The glucose level was determined 72 h post-injection.Rats with a blood glucose level over 300 mg/mL wereincluded in the experiment. The induced diabetic rats(ALX, STZ) were separated into four groups of eight ratsper group. Normal and diabetic rats received in a 24 hperiod, three oral doses of 2 mL each of the 20% infusionof the respective plant.

Blood samples were drawn from the tail before

treatment (zero time), and at 1, 2 and 3 h after the thirddose. The total blood sugar was estimated by theenzymatic method of the glucose oxidase (Bergmeyerand Bernt, 1974).

Statistical analysis.Students’t-test was used to analysethe results according to Snedecor and Cochran (1976),consideringp< 0.05 as a significant difference.

Extract preparation. The plant with greater hypogly-

Figure 1. Hypoglycaemic activity of different plant infusions in experimentalstreptozotocin diabetic rats. 20% infusions were administered orally in threedoses of 2mL in a 24 h period. Streptozotocin was administered i.p. in a doseof 65 mg/kg in citrate buffer 0.5 M, pH 4.5. Percent reduction in plasma glucosecompared with diabetic control group.

Figure 2. Hypoglycaemic activity of different plant infusions in experimentalalloxan diabetic rats. 20% infusions were administered orally in three doses of2 mL in a 24 h period. Alloxan monohydrate was administered i.p. in a single doseof 50 mg/kg dissolved in physiological serum. Percent reduction in plasma glucose

92 I. LEMUS ET AL.

Copyright# 1999 John Wiley & Sons, Ltd. Phytother. Res.13, 91–94 (1999)

caemic activity was extracted at room temperature bysuccessive maceration with petroleum ether and metha-nol solvents. The infusions were preparedin situ at aratio of 200 g of plant powder/L of boiling water. Theextracts were evaporated to dryness with a rotaryevaporator in a water vacuum pump and at a temperaturebelow 60°C. The dried extracts were dissolved orsuspended in distilled water, and were administered todiabetic rats at doses equivalent to an infusion of 0.4 g ofdry plant per 2 mL of dose.

RESULTS

Intravenous injections of normal rats with alloxan(50 mg/kg) or streptozotocin (65 mg/kg), induced asimilar and statistically significant (p< 0.001) increaseof the serum levels of glucose (over 400% of basalglycaemia) 72 h post-injection, compared with thenormal group (Table 2).

On comparing the effects of the 20% plant infusionson the plasma glucose of streptozotocin- and alloxan-induced diabetic rats (Table 3, Fig. 1, 2), a differentresponse could be observed in experimentally diabeticrats to the various plants under study.B. candicansandR. ulmifolius (infusions) were the only species whichreduced significantly the serum glucose (p< 0.001) inboth models of diabetic rats.

DISCUSSION

In spite of presenting certain differences in metabolicand physiopathologic alterations observed in the dia-betes mellitus (Smithet al., 1980), these compoundscause damage to theb cells of the pancreas (Agarwal,1980). Streptozotocin presents a greater selectivity ontheb cells and a lower toxicity than alloxan, the animalsrequiring insulin for their survival (Junodet al., 1969).The diabetes produced by this substance is very similarto juvenile diabetes in the human. Although alloxanusually causes generalized toxicity and a high percen-tage of mortality in experimental animals, it is lessspecific to theb cells of the pancreas, allowing theorganism to respond to a stimulus, and reduce levels ofglycaemia (Lazarov and Palay, 1946).

In streptozocin diabetic rats,R. ulmifoliuspresentedgreater hypoglycaemic activity, with a gradual decreaseof glycaemia until reaching a maximum at 3 h (29%)B.candicansshowed a maximum activity at 2 h (22%),reaching a minimum value of 8% at 3 h. The other two T

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Table 2. Basal glycaemia determination in normal controlrats and in experimentally induced diabetic rats(ALX and STZ) 72 h post-injection

Type of animal Basal glycaemia (mg %)

Normal rat 120.05� 5.78STZ-diabetic rat 510.42� 9.91ALX-diabetic rat 506.33� 9.13

STZ, streptozotocin; ALX, alloxan.Each value corresponds to the average SEM for at least 6measurements. p< 0.001.

HYPOGLYCAEMIC ACTIVITY OF CHILEAN PLANTS 93

Copyright# 1999 John Wiley & Sons, Ltd. Phytother. Res.13, 91–94 (1999)

plantsG. officinalisandM. albadid not exhibit activity inthis model.

In alloxan diabetic rats, all the plant species presenteda marked decrease of glycaemia (p< 0.05), with theintensity of biological activity decreasing fromB.candicans, throughG. officinalis, R. ulmifoliusand M.alba, with a fall in plasma sugar of 39%, 30%, 28% and20%, respectively.

The different activity of the infusion ofR. ulmifoliusinrelation to the other plants, when both experimentalmodels are compared, led us to study extracts of thisplant, in agreement with the previously developedmethod forB. candicans(Lemuset al., 1986). This studyshowed that the ether extract ofR. ulmifoliuscaused a

greater decrease of serum glucose in the STZ diabetic ratsand similar decreases to those obtained for the infusion inthe ALX experimental model, as shown in Table 4.

From the results obtained, we conclude that the fourplants under study were active in the alloxan-inducedhyperglycaemia rat model, but in the streptozotocindiabetic rat model, the plants have diverse activities onplasma glucoseR. ulmifoliusbeing the more effective.

Acknowledgements

The authors are grateful to Dr German Knop for financing this researchproject.

REFERENCES

Agarwal, M. (1980). Streptozotocin: mechanisms of action.FEBS. Lett. 120 (1), 3.

Bergmeyer, H. U., and Bernt, E. (1974). Methods of EnzymaticAnalysis, 2nd edn, pp. 1205±1212.

Junod, A., Lambert, A., Stauffacher, W., and Renold, A.(1969). Diabetogenic action of streptozotocin: Relation-ship of dose to metabolic response. J. Clin. Invest. 48,2129±2139.

Lazarov, W., and Palay, S. (1946). The production and courseof alloxan diabetes in the rat. J. Lab. Clin. Med. 31, 11±17.

Lemus, I., GarcõÂa, R., Jabsa, Z., and Erazo, S. (1986). Actionhypoglyce miante de l'extrait de Bauhinia candicansBentham. Plantes Me dicinales et Phytotha rapie XX, (1),8±17.

MunÄ oz, M., Barrera, E., and Meza, L. (1981). El Uso Medicinaly Alimenticio de Plantas Nativas y Naturalizadas en Chile,p. 150. Publicacio n ocasional No 33. Museo Nacional deHistoria Natural, Santiago.

Smith, S. L., Novotny, M., Moore, S. A., and Felten, D. L.(1980). Studies of hyperlipidemia in drug-induced dia-betic rats by high-performance liquid chromatography. J.Chromatogr. 221, 19±26.

Snedecor, G. W., and Cochran, W. G. (1967). StatisticalMethods. Iowa State University Press, Ames, Iowa.

Valiente, S. (1978). Diabetes Como Problema NutricionalColectivo. Apartado Docente No 111.INTA.

Table 4. Effect of orally administered R. ulmifolius petroleum ether extract (0.61% g/mL), on glycaemia in streptozotocin(STZ) and alloxan (ALX)-induced diabetic rats

Diabetic induced rat Basal glycaemia (mg%) 1 h (mg%) (%) 2 h (mg%) (%) 3 h (mg%) (%)

STZ 510.42� 9.91 350.65� 42.26 (31) 348.58� 34.87 (32) 334.09� 30.10 (35)ALX 506.33� 9.13 419.79� 25.25 (17) 396.40� 33.56 (24) 382.23� 36.44 (26)

(%) Percentage of plasma sugar reduction compared with diabetic control group for streptozotocin (STZ) and alloxan (ALX)-induced diabetic rats.The values are statistically signi®cant (p< 0.001).These values represent the average SEM for at least 6 cases.

94 I. LEMUS ET AL.

Copyright# 1999 John Wiley & Sons, Ltd. Phytother. Res.13, 91–94 (1999)