SHORT COM M UNiCA TlON

Hypotensive Activity of Thymus orospedanus Alcoholic Extract

J. Jimenez, A. Zarzuelo,? and M. E. Crespo Departamento de Farmacologia, Facultad de Farmacia, Universidad de Granada, 18001 Spain

The hypotensive effects of the alcoholic extract of Thymus orospedanus were investigated in both in vivo and in vitro assays. Our results indicate that the extract reduced rat blood pressure and modified the dose-response curve to epinephrine in the rat vas deferens and the rabbit artery, indicative of a competitive antagonism.

Keywords: Thymus orospedanus; alcoholic extract; hypotensive action; epinephrine inhibition.

INTRODUCTION

Thymus orospedanus H . del Villar, an abundant species native to southern Spain, has been the object of several previous studies in our laboratory involving chemical analysis as well as pharmacological effects (Crespo et al., 1986; Cab0 et al., 1987).

A wide ranging phytochemical analysis has also been carried out on a series of extracts (hexane, ethanol and aqueous) of the flowering apex. The alcoholic extract contained very high levels of polyphenolic compounds. These results led to the study of the pharmacological effects of extracts of T. orospedanus, including its diuretic and hypotensive actions (Fos et al., 1979; Monea and Racz-Kotilla, 1978; Barberan, 1986). In this communication we have investigated the mechanism by which the extract modifies the dose-response curve of epinephrine on isolated organs.

MATERIALS AND METHODS

Plant extracts. Thymus orospedanus was collected on the slopes of the Sierra Elvira (province of Granada) during the 1985 flowering season. The flowering apices were dried in a warm air oven at 20-25 “C. Following dessication a series of hexane and ethanol extractions were prepared by Soxhlet extraction. The ethanol extract was used for pharmacological assays. After vacuum evaporation of the solvent at a maximum temperature of 40 “C the resulting dry residue was redissolved in distilled water preparatory to use.

Pharmacological methods. In vivo experiments. The doses of dry residue given were 150 and 300mg/kg which was below the lethal dose 50% (LD50) of 1500 f 275 mg/kg.

Blood pressure activity. Male Wistar rats weighing 250 f 50 g were divided into five groups of ten animals

t Author to whom correspondence should be addressed.

each. Arterial pressure was measured in the tail after warming the animals to 40°C under an IR lamp. Arterial pulse was determined with a piezoelectric transducer and the amplified signal was recorded using a Letica LE 500 digital system. After a ten day period of adaptation, animals with significant variation in blood pressure throughout the day were eliminated from the study. Arterial pressure was measured before administration of the substance (basal) and 1, 3, 5 and 24h after oral administration of 150 and 300 mg/kg of dry residue in 2 mL of distilled water.

In vitro experiments. Isolated tissue preparations were carried out according to the technique of Magnus (1948).

Rat duodenum. Male albino Wistar rats (200f 50 g) were killed by decapitation. Tubular segments (2-3 cm long) were cut from the duodenum and each piece was suspended in a 20 mL organ bath containing Tyrode solution.

Rat vas deferens. Male albino Wistar rats weighing about 200 f 50 g were used. The animals were killed by decapitation. The abdomens were opened by midline incision, the pair of vas deferens was removed and the tissue was immediately placed in a 10mL organ bath containing Krebs solution.

Rabbit artery. Female New Zealand rabbits weigh- ing about 1.5 f 0.2 kg were killed by decapitation. The thoracic cavity was opened and a portion of the aorta was removed. From this segment a spiral segment approximately 3 cm long was taken and placed in a organ bath containing 10mL Krebs- Henseleit’s solution.

Drugs. Epinephrine tartrate [LCl, acetylcholine chlorhydrate [Merck] and barium chloride [Merck] were used as agonists.

Concentration-response curves. The accumulative concentration-response curves for the agonists were recorded. The assay was repeated following the

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152 PHYTOTHERAPY RESEARCH, VOL. 2, NO. 3, 1988 0 Heyden & Son Limited, 1988

HYPOTENSIVE ACTIVITY OF T. OROSPEDANCJS ALCOHOLIC EXTRACT

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-14

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Figure 1. Influence of T. orospedanus extract on blood pressure in rat. Each point indicates mean + SEM. (1) controls (distilled water). (2) T. orospedanus 150mg/kg, (3) T. orospedanus 300 mg/kg. *** p < 0.001, ** p < 0.01, * p < 0.05.

addition of 0.03, 0.06 and 0.12mg/mL T. orospeda- nus extract. The following equipment was used in the experiments: an isolated organ bath (Panlab), an isometric transducer (TRI 010 Letica) and a Dynograph (R-411 Beckman).

RESULTS AND DISCUSSION

The alcoholic extract of T. oropedanus exhibited no significant action regarding an increase in urinary volume or on sodium levels in urine, but did produce a significant decrease in arterial blood pressure at a dose of 150mg/kg during the first hour following administration, whilst the 300 ma/kg dose continued to show hypotensive effects up to 5 h (Fig. 1).

Table 1 shows a significant increase in epinephrine Ed5, produced by 0.03, 0.06 and 0.12mg/mL of T. orospedanus alcoholic extract in both the rat vas deferens and the rabbit artery, whereas the sustance failed to have significant effects on the ED5, of the other agonists such as acethylcholine and barium chloride. The changes induced by the extract on the ED,, of epinephrine, in the absence of any noticeable decrease in its maximal effect, suggests that the extract was acting as an epinephrine antagonist competitively at the doses used in the present study. The results of the in vivo and in vitro tests suggest a possible hypotensive activity by the alcoholic extract of T. orospedanus apparently acting as an antagonist to epinephrine.

Acknowledgement

The authors wish to thank Ms Karen Shashok for her assistance in translating this manuscript into English.

Table 1. Influence of alcoholic extracts obtained from T. orospedunus on EDSo and E,, after addition of different agonists in tissue preparation

ED50 T. orospedanus Agonist hngImL1 (PM) P Em,,(%) P

Rat duodenum Acetylchlorine - 1.34 f 0.36 100 (23 X 10-'M- 0.03 1.47 f 0.37 NS 97.16 f 2.46 NS

14.61 X M) 0.06 1.98 f 0.47 NS 93.89 f 4.88 NS 0.12 2.07 f 0.59 NS 91.22 f 6.06 NS

(16 x 1 0 - ~ M- 0.03 0.77 f 0.1 5 NS 100 NS 10 x 1 0 - ~ M) 0.06 1.46 f 0.45 NS 100 NS

0.12 1.73 f 0.76 NS 100 NS

(0.5 X M- 0.03 5.1 f 1.22 0.05 100 f 10.819 NS 42 X M) 0.06 7.31 f 0.77 0.005 90.80 f 8.08 NS

0.12 13.17 f 1.70 0.001 100.61 i 14.24 NS

(5 X l o - * M- 0.06 1.68 f 0.50 0.05 92.5 f 5.30 NS 85 X M) 0.12 8.6 f 4.1 0.05 90 f 7.07 NS

Rat duodenum BaCI, - 1.13f0.37 100

Rat vas deferens Epinephrine - 2.14 f 0.42 100

Rabbit artery Epinephrine - 0.67 f 0.17 100

Mean i SEM. Number of experiments = 8 (Control) or 4 (with extract). p = values in comparison to the experiments without extract. Em,, is expressed as a percentage of the maximum effect obtained in the presence of the extract by comparing with that of experiments performed without extract.

REFERENCES

Barberan, F. A. T. (1986). The flavonoid compounds from the Labiatae. Fitoterapia 57, 67.

Cabo, J., Crespo, M. E., Jimenez, J., Navarro, C., and Zarzuelo, A. (1987). Pharmacodynamic study of the Thymus orospedanus essential oil. Fitoterapia 58, 33.

Crespo, M. E.. Cabo, J., Jimenez, J., Navarro, C., and Zarzuelo, A. (1986). Composition of the essential oil in Thymus orospedanus. J. Nat. Prod. 49, 558.

Fos, D., Giradlez, J., and Renedo, M. J. (1979). Diuretic activity

of squill Urginea maritima Bak. components. Cienc. Ind. Farm. 11, 141.

Magnus, R. (1948). In Farmacologia Experimental, ed. by F. Guerra, p. 114. Utecha, Mexico.

Monea, N., and Racz-Kotilla, E. (1978) Hypotensive effect of some Centaurea species (Family Asteracea). Rev. Med. 24, 166.

Received 27 October 1987; accepted (revised) 18 January 1988

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