EVALUATION OF ANTIINFLAMMATORY EFFECT OF Capparis Aphylla Roth. USING EXPERIMENTAL ANIMAL MODELS

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ABOUT AUTHORS:
Kanzaria S. H. ,  Patel D. V., Patel K.V., Gandhi T. R.
Anand Pharmacy College.
Anand, Gujarat
*sainikak@yahoo.in

ABSTRACT:
Objective: To evaluate anti-inflammatory effect of Capparis aphylla Roth. (CA) using experimental animal models. Material and method: Female Wistar rats (200 to 250gm) were randomly allocated to six groups (n=6). Except group-I (normal control), arthritis was induced in animals of all groups by injection of 0.2 ml Complete Freund’s Adjuvant (6mg/ml) on day one. Additionally Group III (Std) and Group IV–VI (CA-1, CA-2 and CA-3) received Indomethacin (100mg/kg), CA (300 mg/kg ; 240 mg/kg and 190 mg/kg) from day 1 to day 21 orally. Paw volumes of both sides (measured by plethysmography) was recorded  on day 0, 7, 14, 21. On 7th, 14th, 21stdays the severity of the secondary lesions was evaluated by measuring body weight, arthritic index, Erythrocyte Sedimentation Rate(ESR), Rheumatoid factor(RF), C- Reactive Protein(CRP), Albumin/Globulin(A/G) ratio, X-ray, histopathology of synovial joints. Result: Capparis Aphylla Roth. significantly prevented the freund’s adjuvant induced changes in body weight, serum A/G ratio , arthritic Index, paw edema, ESR, RF, CRP. the results were comparable with the standard drug indomethacin. Conclusion: Antiinflammatory activity of Capparis aphylla Roth. can beattributed to COX-II inhibition leading to decrease in inflammatory mediator.


REFERENCE ID: PHARMATUTOR-ART-1774

INTRODUCTION
Rheumatoid arthritis (RA) is a common chronic inflammatory autoimmune disease of unknown origin. It affects ∼1% of the population and causes the irreversible functional and anatomical joint damage. The have key factors of inflammation and destruction in the rheumatoid joint namely intracellular signaling and proliferation, adhesion, infammation, matrix degradation and angiogenesis that result in a persisting vicious circle. In addition to proinflammatory cytokines, acute phase response (like CRP), abnormal immunoglobulin levels and high rheumatoid factors (RFs) are also considered to be a modulating event during infammatory and immunogenic reaction. . [CHEMICO BIOLOGICAL INTERATION]

Bone loss is a common feature of various inflammatory arthritis Bone erosions results from the activation of an inflammatory response that increases the number and activity of osteoblasts. The complete freund”s adjuvant induced arthritis model represents a systemic inflammatory disease with bone and cartilage changes similar to those observed in RA. The common pathological features of adjuvant arthritis in rats and RA inhuman are joint swelling associated with cellular and pannus invasion of the joint space and bone resorption. Strong bone loss after intense arthritis is induced when adjuvant is injected into the foot pad. The major site of irreversible tissue damage originates from synovium lining  the joint capsule with cartilage and bone, which is often termed the “pannus” and this  is a characteristic feature of RA. [IJPD, CURATIVE EFFECT]

Capparis aphylla Roth.  is commonly known as Caper berry,  Karira, Kerada, Karer, Kair ,  belonging to the family Capparidaceae. Capparis aphylla Roth. has been used traditionally as an anti-inflammatory agent for enlarged cervical glands, sciatica, rheumatoid arthritis, swellings, skin eruptions, ringworm. [C.P. KHAR] Secondly Capparis aphylla Roth. root bark contains spermidine alkaloids which is known for its use in inflammations, asthma and gout.[C.P. KHAR] The different parts of C.aphylla  are  considered to  be  analgesic, diaphoretic, alexeteric, laxative, antihelminthes,  antibacterial, antifungal, antiviral,  and  good in cough,  asthma, ulcers, boils, vomiting, piles and all inflammations.[ANTIHYPERGLYCEMIC, ANTIOXIDENT]

Consequently, the present study has been undertaken to illustrate the beneficial outcome of the drug capparis aphylla Roth.in adjuvant induced arthritic rat model.


MATERIALS & METHODS.

·        Preparation of extract of Capparis aphylla Roth.
3.2 kg root bark of capparis deciduas were percolated three times with ethyl alcohol at room temperature for 2-3 weeks. The solvent from the combined yellowish brown percolate was removed under reduced pressure. 38 g of dark brown viscous residue was taken up in 1N hydrochloric acid and taken out exhudatively with ethyl acetate to remove neutral substances. The aqeous acidic solution was basified with ammonia, yielding yellow precipitates. These yellow precipitates was then extracted with chloroform which on evaporation gave the alkaloid concentrate. The column chromatography of the chloroform extract was carried out in neutral alumina oxide and eluted with a mixture of ethyl acetate:methanol(4:1) and thus isolated the yellow coloured . The yellow compound so obtained contained traces of impurities.

The dragendroffs reagent, methanol, alumina oxide, ethyl acetate, Na azide, Na citrate, succnic acid, bromocresol, brij-95, formaline, formic acid, freund’s adjuvant, carrageenan were obtained from Astron chemical, ahmedabad,Gujarat, India. The total protein kit, RF latex kit, CRP kit were obtained from Coral clinical system, Gujarat,India.

* Animals
Female Wistar albino rats weighing 200–250 g were used. The animals were fed ad libitum and housed in a room with a controlled ambient temperature (22 ±2 °C), humidity (50% ± 10%), and a 12-h light/dark cycle. Animals were acclimated to the housing conditions and handled for 3–4 days before experiments. All experimental procedures were conducted according to the OECD  Guidelines for the Care and Use of Laboratory Animals . The experimental protocols were approved by the Ethical  Committee on Animal Care.

* Preparation of drug solution:
Accurately weighed quantity of dried extract was suspended in distilled water  to prepare the appropriate stock solution of the drug i.e. 190mg/kg, 240mg/kg, 300mg/kg. Indomethacin 100mg/kg was also suspended in distilled water. The doses were administered by selecting theappropriate concentration of the stock solution by orally.

* Animal Grouping
Rats were divided into 6 equal groups (6 rats for each). Group 1 Normal Control group: rats received pure drinking water and regular food ; Group 2 Model  Control group: Freund’s adjuvant; Group 3 Standard group : CFA+ Indomethacin 100mg/kg; Group 4 test group: CFA+MECA 190mg/kg; Group 5 test group: CFA+ MECA 240mg/kg; Group 6 test dose: CFA+ MECA 300mg/kg. Animals were treated with drugs for 21 days.

* Induction of Complete Freund’s Adjuvant Arthritis

- Experimental procedure [Schorlemmer HU,et.al, 1999, 474.]
Female Wistar rats (180 to 250 g ) was be  randomly allocated to seven groups containing six animals each. Animals in Group I(normal control) received distilled water for the entire 21 day study period. Arthritis was be induced in animals of Group III- VII on day one  by injection of 0.2 ml complete Freund’s adjuvant (It consisted of 6 mg Mycobacterium butyricum  being suspended in heavy paraffin oil  to give a concentration of 6 mg/ml) in sub plantar region of the left hind paw. Group III(Std),Group IV(test drug 190 mg/kg), Group V(test drug 240 mg/kg), Group VI (test drug 300mg/kg ). Paw volumes of both sides and body weight was be recorded on the day of injection, whereby paw volume was be measured by plethysmography. On day 7,14,21 the volume of the injected paw was be  measured indicating the primary lesion and the influence of therapeutic agents on this phase. The severity of the induced adjuvant disease was be followed by measurement of the non injected paw (secondary lesions) with a plethysmometer. At the end of study ,on 21 days following parameters was be evaluated.

1.      Body weight[Lin, H-S Hu C-Y,2007, 150, 862–872]
Body weight of each animal was be measured on the day of CFA administration, and later, on 7th, 14th and 21st days. The mean percentage reduction in body weights with respect to that on day of CFA administration was be calculated for each drug treated group and compared with that of disease control group.

2.       Arthritic Index [Arthur Yin Fana, 121,2009 366–371.
On day 21, the severity of the secondary lesions was be evaluated visually and graded all to    following scheme.:

ORGANS

INDICATIONS

SCORING

EARS

Absence of nodules& redness

0


Presence of nodules& redness

1

NOSE

No swelling of connective tissue

0


Intensive swelling of connective tissue

1

TAIL

Absence of nodules

0


Presence of nodules

1

FORE   PAWS

Absence of inflammation

0


Inflammation of at least one jointq

1

HIND PAWS

Absence of inflammation

0


Slight inflammation

1


Moderate inflammation

2


Marked inflammation

3

arthritic score

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