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BACKGROUND & LITERATURE REVIEW
Many medicines and remedies have been prepared from plants and herbs for centuries. It has been known for long that the edible plants have constituents which have medicinal value besides their nutritional benefits. The edible Abelmoschus escuclentus, commonly called okra, has variety of traditional uses such as in sore throat, syphilis, gonorrhea, dysuria, leucorrhea, GI ulcers etc.
Abelmoschus escuclentus traces its beginning from Abyssinia, spreading through to Eastern Mediterranean, India, Africa, North America, South America and the Caribbean. It is a soft-stemmed annual plant of the malvaceae family that is commonly used because of its nourishing components. It has vitamin A, vitamin C, folic acid, Vitamin B6, thiamin, riboflavin, niacin, calcium,1,2 magnesium, iron, zinc, fatty acids and unique pectic polysaccharides.
T. Huynh Ngoc evaluated okra for antihyperlipidemic property and concluded that it can lower triglycerides and cholesterol in hyperlipedimic mice.3 Similarly; Liu IM worked on its antihyperglycemic effects.4 Okra is also considered best vegetable for those feeling weak, fatigued, and suffering from depression. It has also been used as diuretic agent, for treatment of dental diseases and to reduce gastric irritations.5 Abelmoschus escuclentus has many components that have pain relieving properties, for example, magnesium, thiamine8, riboflavin9 and zinc11. Studies have shown that these components either act individually as an antinociceptive agent or aid in antinociception by enhancing the analgesic character of other compounds.
Various animal models have been designed to determine the effects of analgesics against acute and persistent pain since analgesic effects in animals are comparable with the therapeutic effects in man. A hot plate method has been widely used for years to elucidate the antinociceptive character of various compounds and plant extracts against the acute pain.8 This method is used for determining the centrally acting analgesics. Formalin test is used for determining the antinociceptive effects for persistent pain models.9, 10 It is a reliable method with two phases of nociceptive response. The first or early phase is due to its action on nociceptors while the second or late phase is due to inflammatory response involving prostaglandins. This can help in differentiating the newer drugs for their nociceptive activity against inflammatory and non inflammatory pain.12
There are different mechanisms of nociception involving peripheral or central mediators and various receptors for them. Such mediators can be inflammatory or non-inflammatory. Cyclooxygenase mediated inflammatory pain and opioid receptor mediated antinociception are among those. COX inhibitor assay kit is used for the analysis of Cyclooxygenase inhibiting activity. This assay utilized enzyme assay technique measuring the synthesis of prostaglandins.13, 14
RATIONALE OF STUDY
For century, various analgesics have been produced that are effective in treating different types of pain. Still, in nature many plants are available that can be processed to formulate newer drugs. Abelmoschus has its folk lore use for the management of joint pains and other aching conditions. Work has been done on its anti inflammatory5 and antioxidant6 properties while little data is available on its antinociceptive activity. Recently, a study conducted in India, has shown that extract of Abelmoschus escuclentus was effective in reducing pain in animal model.7 It is necessary to evaluate the behavior of Abelmoschus escuclentus against acute and chronic pain and to find the possible mechanism of this activity.
This would provide scientific basis for its use in folklore medicine and, also, for the preparation of safe, effective and inexpensive herbal medication for relieving pain. It will also aid in elucidating that whether Abelmoschus escuclentus works by central or peripheral antinociceptive mechanism and is effective against inflammatory or non-inflammatory pain or both.
Based on the traditional use, dried Abelmoschus escuclentus fruits are taken in powder form to treat certain pain conditions like backache, muscle ache. Its oil is available with hakim to be rubbed at pain full knees. However, no scientific data is available that shows conclusively whether it has such analgesic property or not. Therefore, this study is designed to demonstrate the probable antinociceptive action of Abelmoschus escuclentus as depicted in its folklore use. It will also be elucidated preliminarily regarding the possible mechanism of this antinociceptive activity.
The objective of the study is to determine the antinociceptive activity of extract of Abelmoschus escuclentus fruit in acute and persistent/inflammatory pain animal models. It will also partially elucidate the possible mechanism of this antinociceptive effect and compare that effect with a standard analgesic.
Extraction of plant material
After collection, the okra fruits will be shade dried for 15 days and crushed into for further extraction. The extract will be prepared by hot continues soxhlet extraction using methanol: water-70:30.15
Preliminary phytochemical screening of the powdered material will be carried out.
Albino rats, 150-200g,7 will be used for this purpose.
Dose of extract
Different doses of the Abelmoschus used in experiments to study pharmacological activities ranges from 100mg/kg to 800 mg/kg.15, 16 For the purpose of this experiment, doses of 300 mg/kg will be used.
Control group for hotplate method: Normal saline will be given to rats
Extract (300 mg/kg) will be given to rats for hotplate method
Opioid antagonist (naloxone Dose) with 300 mg/kg extract for hot plate method
Extract (300 mg/kg) in formalin induced pain model for measuring response in 1st phase (0-15 min)
Extract (300 mg/kg) in formalin induced pain model for measuring response in 2nd phase (20-50 min)
Control group for formalin induced pain model
Indomethacin 10 mg/kg in formalin induced pain model for measuring response in 2nd phase (20-50 min)
Route of administration
Measurement of antinociceptive activity in acute pain model
Hot plate analgesiometer will be used to determine the analgesic activity for an acute pain model. The rats shall be placed on the hot plate at 55Â° to 56Â°C and time to lick the paws due to thermal pain before and after the administration of the extract for 180 minutes will be determined.
Measurement of antinociceptive activity in persistent/inflammatory pain model
Formalin induced pain in rat paw will be used to assess the antinociceptive activity of Abelmoschus escuclentus extract. The response will be measured in different groups against the early and late nociception by formalin. Response will be measured every 5 minutes with observation time of 60 seconds for each measurement. The response is measured as the number of flinches per observation period and/or licking/ biting of the injected paw, recorded as total licking time (s) per observation period.
This will be compared with the analgesic response produced by the standard drug indomethacin.
Determination of mechanism of antinociception
Two possible mechanism of antinociceptive action will be determined.
Measurement of antinociceptive effect of extract in the presence of an opioid antagonist, naloxone, will be done by hotplate analgesiometer.
In vitro analysis of cyclooxygenase inhibitor activity by COX inhibitor assay using enzyme assay technique will be conducted.
Student t test to compare mean response time
Analysis of variance (ANOVA) for multiple comparisons against control with suitable post hoc test.
P value < 0.05 will be considered statistically significant.