Pharmacological Studies on Some Synthetic Dimethoxy Flavones

Jayshree, N (2011) Pharmacological Studies on Some Synthetic Dimethoxy Flavones. Doctoral thesis, The Tamilnadu Dr. M.G.R. Medical University, Chennai.


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The quest for novel therapeutic compounds to treat existing diseases as well as for the new diseases is never ending. Though the advent of molecular biology and combinatorial chemistry brought in the concept of rational drug design, the interest in nature as a source of potential therapeutic agent and in herbal based products continues. The plethora of phytochemicals present in plants may be exploited directly or they may provide a drug lead or pharmacophore. The flavonoids form one of the most numerous and ubiquitous group of phytochemicals. They are known to be potent antioxidants and have exhibited several activities. The anti-inflammatory, hepatoprotective, antiatherogenic and anticancer activities are some of the few activities that have been strongly associated with flavonoids. The underlying common mechanism in all these diseases is the free radical mediated cell damage. Flavonoids by their free radical scavenging abilities are of use in treating these disorders. There are reports of the analgesic activity of flavonoids too. Many of these reports are on the natural flavonoids. The synthetic flavonoid derivatives have also been shown to possess many of the activities of the natural flavonoids. With this background, it was decided to evaluate the pharmacological activities of some synthetic dimethoxy flavones. The compounds chosen for this study are 3,6-dimethoxy flavones, 6,2'-dimethoxy flavone and 6,3'-dimethoxy flavone. There are no reports of any studies with these compounds. Flavonoids are a group of compounds with individual agents having multiple activities. Hence, in this study, a broad based pharmacological evaluation of the three dimethoxy flavones was planned to be carried out. The compounds were subjected to antioxidant, analgesic, anti-inflammatory, hepatoprotective, in vitro anticancer and antimicrobial activities. Since all the three compounds were being tested for the first time, acute toxicity test was carried out. The test revealed that even in the dose of 2000mg/kg, the compounds did not produce mortality nor were there any gross behavioural changes. This indicates the safe nature of the compounds and the compounds were taken up for further studies. The compounds were tested for their antioxidant capability using in vitro methods. The DPPH and the hydroxyl radical scavenging ability were evaluated. All the three compounds exhibited dose dependant scavenging ability. The IC50 value for 6,3'-DMF was found to be lower than that of the standard drug Vitamin E for scavenging DPPH radical. In the case of hydroxyl radical scavenging, 3,6-DMF and 6,2'-DMF were able to bring about almost 95% radical scavenging at the highest concentration of 800μg/ml. In the case of 6,3'-DMF a much lower concentration of 30μg/ml produced a similar effect. The three dimethoxy flavones were then evaluated for analgesic and anti-inflammatory activities. To explore the analgesic ability, three different, well established models of nociception were used in order to obtain a complete picture of the types of pain for which these compounds would be useful. In the acetic acid model, 3,6-DMF showed maximum inhibition of nociception which was 87%. The other two compounds produced a maximum inhibition of 70% at the dose of 100mg/kg. In the formalin model too, during the latter phase 3,6-DMF produced a maximum inhibition of nociception at a dose of 100mg/kg and this was found to be 83.96% The other two compounds produced about 75% inhibition of nociception. In this model, during the earlier phase, the extent of inhibition of nociception was less ranging from about 40% to 48% with 3,6-DMF showing the least activity and 6,3'-DMF showing the maximum activity. In the tail immersion model too, the inhibition of nociception ranged from 53 to 71% with 3,6-DMF showing the least activity and 6,3'-DMF showing the maximum activity. The results from these studies indicate that all the three compounds possess characteristic analgesic activity. They were active against pain of inflammatory origin and neurogenic pain. 3,6-DMF seems to show a preferential activity against inflammatory pain whereas 6,2'-DMF and 6,3'-DMF are more selective for neurogenic pain. The in vitro membrane stabilition study is an indicator for anti-inflammatory activity of any compound. Hence this test was also performed. All the three compounds induced membrane stabilisation. The stabilisation produced by 100μg/ml of the compounds was comparable to that of the standard drug Diclofenac 50μg/ml. The in vivo anti-inflammatory ability of the compounds was tested using the carrageenan induced hind paw oedema in rats. All the three compounds produced a significant decrease in inflammation. 6,2'-DMF (100mg/kg) produced a maximum inhibition of 78.61% at the fifth hour evaluation, 6,3'-DMF produced 68.18% inhibition and 3,6-DMF produced 61.75% inhibition. Diclofenac sodium at a dose of 100mg/kg produced 81.32% inhibition at the fifth hour evaluation. The study on the possible mechanisms of analgesia and anti-inflammation produced by these compounds indicated that the opioid receptors played a definite role in the induction of analgesia. This was shown by the ability of Naloxone to block the analgesia produced by the compounds. GABAergic system also seems to contribute to the analgesic potential of these compounds. This was confirmed by pretreating the animals with Bicuculline which is a GABAA antagonist. Pretreatment with Bicuculline attenuated the analgesic activity of these compounds. There was no apparent role of the adrenergic system or the ATP sensitive potassium channels in the analgesic activity of these compounds. An in vitro study on the effect of the compound on mediators of pain and inflammation indicates the role of COX inhibition as well as reduction of TNF-α and IL-1β levels. Since the opioid mechanism was shown to be involved in the analgesic activity of the compound, the development of tolerance to the analgesic effect of these compounds was studied by repeatedly injecting the compounds at short intervals of 4h and checking for their analgesic activity. Morphine showed development of tolerance whereas there was no development of tolerance with the dimethoxy flavones. Interestingly 3,6-DMF showed an increase in the analgesic effect as result of repeated injections. This may indicate an accumulation of this compound in the body or it may point to the long biological half life of this compound. The other two compounds showed only a marginal increase in analgesic activity. A combination of an analgesic drug with another drug which is either analgesic or not an analgesic is commonly employed to improve the analgesic profile as well as to reduce unwarranted side effects. The three dimethoxy compounds were seen to potentiate the analgesic effect of sub-maximal doses of both Morphine and Diclofenac sodium. Most of the non steroidal analgesic anti-inflammatory drugs as well as the anti-inflammatory corticosteroids show gastric irritation and ulcer development as a side effect. Hence, the dimethoxy flavones were also tested for their ulcerogenic potential. The compounds did not show any signs of ulceration. They were also able to protect the gastric mucosal damage induced by Aspirin. Compounds with antioxidant property are generally known to be efficient hepatoprotective agents and one of the commonly used hepatoprotectants is silymarin which is a flavanolignan. The hepatoprotective activity of these compounds was assessed by using the Paracetamol induced model of hepatotoxicity. The Paracetamol treated group of animals showed an increase in the hepatic enzyme levels such as AST, ALT, ALP and GGT. The level of bilirubin was also more while the protein levels were decreased. The antioxidant enzyme levels such as SOD, CAT, GSH-Px, and GST were all decreased while the level of lipid peroxidation products was increased. Silymain could reverse these effects when given at a dose of 50mg/kg. The dimethoxy flavones also were able to reverse the effect of Paracetamol induced damage at the dose levels of 100mg/kg indicating the hepatoprotective nature of these compounds. Epidimeological studies indicate that increased intake of flavonoids is able to decrease the incidence of cancer. The antioxidant capability of flavonoids is seen to play a major role in their anticancer abilities. Hence, in the present study too, the in vitro anticancer ability of the dimethoxy flavones against HepG2 cell lines was studied. The viability of cells was analysed using the MTT assay method. All the three compounds showed a decrease in the member of viable cells in a time and dose dependant manner. Studies on cell cycle analysis revealed that these dimethoxy flavones could bring about accumulation of cells in G1 phase cell cycle indicating that this could be one more mechanism for the antiproliferative ability of the compounds. Further studies also showed a tendency for induction of apoptosis with the three dimethoxy flavones. This was evaluated by the propidium iodide staining method as well as the agarose gel electrophoresis techniques. The antimicrobial potency of the compounds was also assessed. The three compounds showed almost a similar pattern of antibacterial activity. They seemed to have greater activity against gram negative organisms as compared to gram positive microorganisms as was seen with their MIC values. In the antimycobacterial assay, only 6,3'-DMF showed inhibition of the Mycobacterium tuberculosis H37Rv at a concentration of 100μg/ml. The antifungal studies also revealed all the three compounds were able to inhibit the tested pathogenic fungi at concentrations of 5μg/ml and above. The antiviral activity against Chikungunya virus as well as the HSV-1 and 2 virus was seen at a concentration of 100μg/ml for all the three dimethoxy flavones when the virus was incubated with 3,6-DMF and 6,3'-DMF at least for a period of one hour and 6,2'-DMF showed an activity when it was incubated for at least half an hour. In conclusion, this study on the three synthetic dimethoxy flavones – 3,6-dimethoxy flavone, 6,2'-dimethoxy flavone and 6,3'-dimethoxy flavone has revealed the pharmacological potential of these compounds. 6,3'-DMF showed excellent antioxidant activity in the in vitro studies. All the three compounds possess good analgesic and anti-inflammatory activities. Unlike many of the analgesic and anti-inflammatory compounds currently available, they did not show ulcerogenic activity. 3,6-DMF was also able to offer protection against the Aspirin induced ulcers. Hence these compounds may be useful as analgesic and anti-inflammatory drugs either alone or in conjugation with the existing drugs. The three flavones have also shown good hepatoprotective ability especially at a higher dose. Their antifungal activity has been found to be excellent. The in vitro studies have revealed the anticancer potential of these compounds. Studies using animal models of cancer could confirm this fact. Pharmacokinetic and clinical studies are suggested to substantiate the findings of this study in future. The present study has shown that 3,6-dimethoxy flavone, 6,2'-dimethoxy flavone and 6,3'-dimethoxy flavone hold promise for ushering in a new vista of drugs for the benefit of future generations.

Item Type: Thesis (Doctoral)
Additional Information: Reg.No.21201
Uncontrolled Keywords: Synthetic Dimethoxy Flavones, Pharmacological studies.
Subjects: PHARMACY > Pharmaceutics
Depositing User: Subramani R
Date Deposited: 31 Jul 2020 08:46
Last Modified: 24 Oct 2022 13:19

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