Analytical Study of Selected Anti-Inflammatory Drugs
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Analytical Study of Selected Anti-Inflammatory Drugs Thesis Presented by Enas Taha Abdelhamed M.Sc. in Pharmaceutical Sciences Pharmaceutical Chemistry Faculty of Pharmacy - Cairo University 2012 Submitted for The Degree of Doctor of Philosophy In Pharmaceutical Sciences (Pharmaceutical Chemistry) Under the supervision of Prof. Dr. Sonia Talat Hassib Professor of Pharmaceutical Chemistry Faculty of Pharmacy - Cairo University Prof. Dr. Ghaneya Sayed Hassan Professor of Pharmaceutical Chemistry Faculty of Pharmacy - Cairo University Prof. Dr. Asmaa Ahmed El-Zaher Professor of Pharmaceutical Chemistry Faculty of Pharmacy - Cairo University Dr. Marwa Ahmed Fouad Associate Professor of Pharmaceutical Chemistry Faculty of Pharmacy - Cairo University Faculty of Pharmacy Cairo University 2018 Abstract Four simple, accurate, sensitive and economic Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopic (ATR-FTIR) methods have been developed for the quantitative estimation of some non-steroidal anti- inflammatory drugs alone or in presence of related substances. The first method involves the determination of etodolac by direct measurement of the absorbance at 1716 cm-1. In the second method, the second derivative of the IR spectra of tolfenamic acid and its imperity (2-chlorobenzoic acid) was used and the amplitudes were measured at 1084.27 cm-1 and 1058.02 cm-1 for tolfenamic acid and 2-chlorobenzoic acid, respectively. The third method used the first derivative of the IR spectra of bumadizone and its reported degradation product, N,N-diphenylhydrazine and the amplitudes were measured at 2874.98 cm-1 and 2160.32 cm-1 for bumadizone and N,N-diphenylhydrazine, respectively. The fourth method depends on measuring the amplitude of diacerein at 1059.18 cm-1 and of rhein, its reported degradation product, at 1079.32 cm-1 in their first derivative spectra. The four methods were successfully applied on the pharmaceutical formulations by extracting the active constituent in chloroform and the extract was directly measured in liquid phase mode using a specific cell. Moreover, validation of these methods was carried out following International Conference of Harmonisation (ICH) guidelines. Synthesis of new derivatives through an esterification reaction between the carboxylic acid functional group in NSAIDs and a naturally occurring phenolic antioxidant, thymol, to give anew prodrug. This is one of possible approaches to solve GIT irritation problem. The suggested prodrugs to be prepared are Etodolac-thymol and Tolfenamic acid-thymol prodrugs. The new prodrugs structure should be confirmed by FT-IR, 1H NMR, 13C NMR, mass spectroscopy and elemental analysis. After synthesis and structure confirmation, the prodrugs will be evaluated for their chemical stability in 2 different temperatures and pHs. Moreover, a kinetic study in 20% liver homogenate and 10% buffered plasma would be carried based on RP- HPLC method for the separation and determination of the 2 new prodrugs and their related bioactive products. Key words: ATR-FTIR, NSAIDs, Etodolac, Diacerein, Bumadizone, Tolfenamic acid, Prodrugs, RP- HPLC. 3 Aim of the Work NSAIDs represent one of the most important classes used for the treatment of many inflammatory diseases and possessing analgesic and antipyretic activity. The aim of this work is to develop simple, accurate, precise and, most importantly, sensitive methods for the determination of some NSAIDs either alone or in combination with their related substances. The work involves the analysis of these drugs in their pure form or in commercially used pharmaceutical dosage forms using ATR-FTIR spectroscopic technique. Drugs cited in this part are etodolac, diacerein, bumadizone and tolfenamic acid. The pharmacopoeial methods described for the analysis of etodolac and tolfenamic acid depend on titrimetric method in pure form. In literature, numerous analytical methods have been reported for the determination of etodolac and diacerein, while few methods were reported for bumadizone and tolfenamic acid. Till date, no reported ATR-FT-IR method appears to be available for the determination of the cited drugs. Derivative IR-spectroscopy has been suggested for resolving spectral overlap displayed by Diacerein and its degradation product rhein, bumadizone and its degradation product, N,N-diphenylhydrazine, finally tolfenamic acid and its impurity, 2-chlorobenzoic acid. In addition, method validation and statistical analysis was included to compare the obtained results of the proposed methods with those of reference or pharmacopoeial methods. Gastric irritation and ulcerogenic effect of the acidic NSAIDs are one of the most challenging problems in designing novel anti-inflammatory agents. 4 Therefore, prodrug approach can give an opportunity in medicinal chemistry to improve some undesirable properties hindering the clinical usefulness of a drug. The work was aimed to include synthesis of new derivatives through an esterification reaction between the carboxylic acid functional group in NSAIDs and a naturally occurring phenolic compound, thymol, to give a new prodrug. Thymol was selected to get a safer promoiety, a target which is always challenging in designing prodrugs. These types of promoieties were traditionally in use for their medicinal as well as flavoring properties with well documented safety profiles, thus do not involve the risk of unwanted effects after they are hydrolyzed. This is one of possible approaches to solve GIT irritation problem. The suggested prodrugs prepared were etodolac-thymol tolfenamic acid- thymol prodrugs. The new prodrugs structure were confirmed by FT-IR, 1H NMR, 13C NMR, mass spectroscopy and elemental analysis. After synthesis and structure confirmation, the prodrugs were evaluated for their chemical stability in different temperatures and pHs. Moreover, a kinetic study in 20% liver homogenate and 10% buffered plasma were carried out based on a new developed RP-HPLC method for the separation and determination of the new prodrugs and their related bioactive products. 5 Non-Steroidal Anti-Inflammatory Drugs (NSAIDS). The term NSAID indicates a class of drugs known as non-steroidal anti- inflammatory drugs. This is an important therapeutic class of drugs which in addition to their anti-inflammatory effects, they may possess both analgesic and antipyretic activities. (1) The mechanism of action of NSAIDs involves inhibition of cyclooxygenases (COX), enzymes that initiate the formation of prostanoids.(2) There are 3 subtypes of COX enzymes: COX-1(constitutive), COX-2 (inducible in inflammatory processes), and isozyme COX-3.(3) To be an effective competitive inhibitor of arachidonic acid that binds to COX, (Figure- 1), a drug must possess both high lipophilic and acidic properties to mimic the natural substrate. Some NSAIDs inhibit the lipoxygenase pathway, which may itself result in the production of algogenic metabolites.(4) Figure-1: Arachidonic acid cascade.(4) 6 Classification of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs). NSAIDs can be classified based on their chemical structure into: 1. Salicylate and salicylic acid derivatives: This group of drugs was among the first introduced into medicine, and aspirin is the prototype of this group. (3) Most of these drugs are either marketed as salts of salicylic acid, ester or amide derivatives. Salicylate derivatives may inhibit both forms of COX by reversible or irreversible mechanism (5) (Figure- 2a). Diflunisal is a moderately potent inhibitor of prostaglandin biosynthesis; it does not have appreciable effect on platelet aggregation and does not significantly produce gastric or intestinal bleeding (6) (Figure-2b). O O a. Aspirin O HO HO O HO F b. Diflunisal F Figure-2: Salicylate and salicylic acid derivatives. 2. Acetic acid derivatives: i) Phenylacetic acid derivative Diclofenac is used to treat painful conditions such as arthritis, sprains and strains, gout, migraine, dental pain, and pain after surgical operations. It decreases pain and inflammation (7) (Figure-3). Diclofenac potassium salt was developed to increase absorption rate with rapid onset of pain relief. (5) 7 HO O Cl NH Cl Figure-3: Chemical structure of diclofenac. ii) Aryl and heteroaryl carboxylic acid derivatives Indomethacin is more potent COX inhibitor than aspirin, but patient intolerance generally limits its use to short-term dosing (5) (Figure-4a). Ketorolac, (Figure-4b) is a non-steroidal agent with potent analgesic and moderate anti-inflammatory activity. It is administered orally, intramuscularly, intravenously, and as topical ophthalmic solution.(8) Sulindac, (Figure-4c) is a prodrug which is metabolized to a pharmacologically active sulfide derivative. This metabolite potently inhibits COX. Sulindac is also metabolized to the inactive sulfone derivative. (9) Etodolac, (Figure-4d) is an acetic acid derivative, which is primarily used in the treatment of rheumatic diseases and postoperative pain. It is rapidly metabolized in the liver, followed by renal elimination as the primary route of excretion. (10) O O OH a. Indomethacin N Cl O O O b. Ketorolac N OH 8 F O HO S c. Sulindac O HO H N d. Etodolac O O Figure-4: Aryl- and heteroaryl carboxylic acid derivatives. 3. Propionic Acid derivatives: Propionic acid derivatives are nonselective COX inhibitors, although there are considerable variations in their potency as COX inhibitors, they are approved for use in the symptomatic treatment of rheumatoid arthritis and osteoarthritis. Some are also approved for pain, ankylosing