PHYSICOCHEMICAL EVALUATION OF DIFFERENT BRANDS OF SULFADOXINEPYRIMETHAMINE IMPORTED INTO THE SIERRA LEONEAN MARKET

COLLEGE OF MEDICINE AND ALLIED HEALTH SCIENCES UNIVERSITY OF SIERRA LEONE

DEPARTMENT OF PHARMACEUTICS, FACULTY OF PHARMACEUTICAL SCIENCES

TRACEY E.C.JONES AUGUST 2012

1.

TABLE OF CONTENTS
1.1 1.2 INTRODUCTION ______________________________________________________________ 4 QUALITY CONTROL OF PHARMACEUTICAL TABLETS _________________________________ 5

1.2.1 Friability __________________________________________________________________________ 5 1.2.2 Uniformity of dosage units ____________________________________________________________ 5 1.2.2.1 Weight Variation _______________________________________________________________ 5 1.2.2.2 Content Uniformity _____________________________________________________________ 6 1.2.3 Chemical Assay ____________________________________________________________________ 6 1.2.4 Dissolution ________________________________________________________________________ 6 1.2.4.1 Dissolution Apparatus ___________________________________________________________ 6 1.2.4.1.1 Dissolution apparatus type I (Basket apparatus) ____________________________________ 6 1.2.4.1.2 Dissolution apparatus type II (Paddle apparatus) ___________________________________ 7 1.2.4.2 Factors affecting determination of dissolution rate_____________________________________ 8 1.2.4.2.1 Instrumental factors __________________________________________________________ 8 1.2.4.2.2 Temperature ________________________________________________________________ 9 1.2.4.2.3 Dissolution media ____________________________________________________________ 9

1.3

DRUG QUALITY _______________________________________________________________ 9

1.3.1 Generic versus brand products and drug quality __________________________________________ 10 1.3.1.1 Bioavailability _________________________________________________________________ 10 1.3.1.2 Issues beyond bioavailability _____________________________________________________ 10 1.3.2 Private Pharmacy and Drug Quality ____________________________________________________ 11

1.4 1.5

JUSTIFICATION OF STUDIES ____________________________________________________ 11 OBJECTIVES_________________________________________________________________ 12

General Objective __________________________________________________________________ 12 Specific Objectives _________________________________________________________________ 12

1.5.1 1.5.2

2. REVIEW OF RELATED LITERATURE __________________________________________________ 13

2.1 Malaria ______________________________________________________________________________ 13 2.1.1Life cycle ____________________________________________________________________________ 13 2.1.2: Recurrent malaria ___________________________________________________________________ 13 2.1.3: Signs and symptoms _________________________________________________________________ 14 2.1.4: Complications _______________________________________________________________________ 14 2.1.5: Medications ________________________________________________________________________ 14 2.1.6: Treatment __________________________________________________________________________ 15 (58) 2.2Sulfadoxine-Pyrimethamine ___________________________________________________________ 16 2.2.1Definition ___________________________________________________________________________ 16 2.2.2: Pharmacology _______________________________________________________________________ 16 2.2.3: Indications and usage ________________________________________________________________ 17 2.2.4: Microbiology _______________________________________________________________________ 17 2.2.4.1Pharmacodynamics _______________________________________________________________ 17 2.2.4.2: Activity in-vitro __________________________________________________________________ 17 2.2.4.3: Drug resistance __________________________________________________________________ 17 2.2.5: Dosage and Administration __________________________________________________________ 17 2.2.5.1: Malaria prophylaxis ______________________________________________________________ 17 2.2.5.2: Malaria treatment _______________________________________________________________ 18

2.2.6Storage/ Stability _____________________________________________________________________ 18 2.2.7: Drug Interactions ____________________________________________________________________ 18 2.2.8: Adverse Reactions ___________________________________________________________________ 18 2.2.9: Precautions_________________________________________________________________________ 18 2.2.10: Over dosage _______________________________________________________________________ 19 (58) 2.2.11: Pharmacokinetics ________________________________________________________________ 19 2.2.11.1: Absorption ____________________________________________________________________ 19 2.2.11.2: Distribution ____________________________________________________________________ 19 2.2.11.3 Metabolism ____________________________________________________________________ 19 2.2.11.4 Elimination _____________________________________________________________________ 19 2.2.12 Characteristics in Patients_____________________________________________________________ 20

3.1

MATERIALS USED IN THIS STUDY _______________________________________________ 21

Instruments _______________________________________________________________________ 21 Chemicals and reagents used _________________________________________________________ 21

3.1.1 3.1.2

3.2

METHODOLOGY _____________________________________________________________ 21
Location of study___________________________________________________________________ 21 Data collection and sample size _______________________________________________________ 21

3.2.1 3.2.2

3.3 3.4

CONDITIONS OF STORAGE OF DRUG SAMPLES COLLECTED ___________________________ 24 PHYSICO-CHEMICAL TESTS ____________________________________________________ 26

3.4.1 Assay ____________________________________________________________________________ 26 3.4.1.1 Apparatus ____________________________________________________________________ 26 3.4.1.2 Method description ____________________________________________________________ 26 3.4.2 Uniformity of weight________________________________________________________________ 27 3.4.2.1 Apparatus ____________________________________________________________________ 27 3.4.2.2 Method description ____________________________________________________________ 27 3.4.3 Dissolution test ____________________________________________________________________ 28 3.4.3.1 Apparatus ____________________________________________________________________ 28 3.4.3.2 Preparation of phosphate buffer solution __________________________________________ 28 3.4.3.3 Measurement of the ph of phosphate buffer solution ________________________________ 28 3.4.3.4 Method description ___________________________________________________________ 29 3.4.4 Friability test ______________________________________________________________________ 29 3.4.4.1 Apparatus ____________________________________________________________________ 29 3.4.4.2 Method description ____________________________________________________________ 29

4.1 4.2

WEIGHT UNIFORMITY OF TABLETS OF EIGHT BRANDS OF SULFADOXINE-PYRIMETHAMINE 31 ASSAY (% CONTENT) OF EIGHT BRANDS OF SULFADOXINE- PYRIMETHAMINE ___________ 34
Calculation of the amount of powdered sulfadoxine used in assay ___________________________ 34 Calculation of the amount of powdered pyrimethamine used. ______________________________ 36

4.2.1 4.2.2

4.3

DISSOLUTION OF EIGHT BRANDS OF SULFADOXINE-PYRIMETHAMINE _________________ 41

Calculation of the concentration of Sulfadoxine __________________________________________ 41 Calculation of the percentage amount of sulfadoxine dissolved: _____________________________ 43 Calculation of the concentration of pyrimethamine ________________________________________ 45

4.3.1 4.3.1 4.3.2

4.4 4.5

FRIABILITY OF EIGHT BRANDS OF SULFADOXINE-PYRIMETHAMINE ____________________ 50 DESCRIPTION OF TABLETS AND PACKAGING MATERIALS ____________________________ 52
Malareich ________________________________________________________________________ 52 Malafan __________________________________________________________________________ 52

4.5.1 4.5.2

4.5.3 4.5.4 4.5.5 4.5.6

Maladar __________________________________________________________________________ 52 Spymal ___________________________________________________________________________ 52 Antim ____________________________________________________________________________ 53 Maldox___________________________________________________________________________ 53

4.6 5 6.1 6.2 6.3

INSPECTION OF CONTAINERS FOR LABELLINGS ____________________________________ 53 DISCUSSION-----------------------------------------------------------------------------------------------------------54 CONCLUSIONS ______________________________________________________________ 55 RECOMMENDATIONS ________________________________________________________ 55 RELEVANCE OF THE STUDY IN SIERRA LEONE ______________________________________ 55 APPENDIX-------------------------------------------------------------------------------------------------------------57 REFERENCES

CHAPTER ONE 1.1 Introduction

The increase in the number of generic drug products from multiple sources has placed people involved in the delivery of health care in a position of having to select one from among several seemingly equivalent products. Variable clinical responses to these drug products supplied by two or more drug manufacturers are documented (1). These variable responses may be due to formulation ingredients employed, methods of handling, packaging and storage and even other rigors of manufacturing process. Thus there is need to determine their pharmaceutical equivalence in order to ensure interchangeability (2). Many developing countries do not have an effective means of monitoring the quality of generic drug products in the market. This results in widespread distribution of substandard and/or counterfeit drug products. It was in view of this fact that the World Health Organisation (WHO) issued guidelines for global standards and requirements for the registration, assessment, marketing, authorisation and quality control of generic pharmaceutical products(2). Generic drug products must satisfy the same standards of quality, efficacy and safety as those applicable to the innovator products. Preliminary physiochemical assessment of the products is very important and in-vitro dissolution testing can be a valuable predictor of in-vivo bioavailability and bioequivalence of oral solid dosage forms (3). Sulfadoxine-pyrimethamine (500mg and 25mg respectively) combination tablets were used for the prophylaxis and suppression of chloroquine-resistant plasmodium falciparium malaria which is a cause of high mortality among children in tropical Africa. The action of the sulfadoxine component is due to its effects in potentiating the effect of pyrimethamine. This action is presumably due to sequential blockade of different stages in plasmodial synthesis of nucleotides (4) . Apart from Fansidar (Roche Pharmaceuticals, Lagos), which is an expensive innovator product, several less expensive generic antimalarials are marketed in Sierra Leone. Hence there is the need to assess the bioequivalence of these generic products as compared with the innovator product. In the present study, the equivalence of different brands of sulfadoxine-pyrimethamine tablets sourced from retail pharmacies in Freetown, Sierra Leone would be determined using in-vitro methods. This preliminary study is aimed at obtaining baseline data towards the establishment of bioequivalence of the tablets.

1.2 Quality Control of Pharmaceutical Tablets The increasing complexity of modern pharmaceutical manufacture arising from a variety of unique drugs and dosage forms imposes complex ethical, legal and economic responsibilities on those concerned with the manufacturing of modern pharmaceuticals. An awareness of these factors is the responsibilities of those involved in the development, manufacture, control and marketing of quality products. A systematic quality assurance programme takes into consideration potential raw materials, in-process, packaging of materials, labelling and finished product variables. The general appearance of a tablet, its visual identity and above all its elegance is essential for consumer acceptance, for control of lot-to-lot uniformity and general tablet-to-tablet uniformity and monitoring trouble-free manufacturing. The control of the general appearance of a tablet involves measurement of a number of attributes such as a tablet size, shape, colour, presence or absence of an odour, taste, surface texture, physical flaws and consistency and eligibility of any identifying marking (5).The following discussions present basic concepts of the quality requirements of tablet dosage forms. 1.2.1 Friability Resistance to abrasion or friability is an extremely important attribute. In fact, it takes precedence over hardness and must not be ignored. Like disintegration and dissolution, hardness is quicker and easier to measure but they are mutually dependent. In principle, tablets are subjected to tumbling which is consistent with the level and time encountered during manufacture, dedusting, coating, handling, transport and packaging and of course with the patient. Specifically tablets of known weight (Wo) are dusted and tumbled in a container for a fixed time (turns/cycles or falls) and weighed (W) again. The friability, F, is given by: 1.1 The most widely used commercial tester is the Roche Friabilator, where 6g or 20 tablets are dedusted and placed in a 12 inch diameter Perspex drum and rotated 100 times (4mins). A limit of < 1% friability is often set, but less than 0.1% is a realistic goal in development(6). 1.2.2 Uniformity of dosage units The uniformity of dosage units can be demonstrated by either of two methods: weight variation or content uniformity. Both are used to control the uniformity of active constituent of tablets. The uniformity of dosage unit is more comprised in tablets formulated to be given in divided doses; in this instance, both weight uniformity and in some instances content uniformity may be questionable (7) 1.2.2.1 Weight Variation There are some important differences in the way various compendia control the determination of tablet weight, although the basic attitude is the same. Tablet weight, assuming perfect mixing, is a control of drug dose and is determined simply and quickly

both in-process and as part of quality control (6).Weight control is based on a sample of 20 tablets, but the interpretation of results obtained is different and debatable. Weight uniformity, however still applies, in particular , where the product contains greater than 50mg active ingredients or where the drug constitutes greater than 50% of the compression weight [USPXXVII].

1.2.2.2 Content Uniformity Although weight rather than the drug content has been traditionally used to control dose, the increasing number of low dose and potent drugs has led to greater need for content uniformity requirements instead of relying on a bulk assay result which assumes perfect mixing and remains a universal pharmacopoeial requirement [USPXXVII]. Since content uniformity of dosage units has its own share in drug quality and therapeutic efficacy; some studies have shown that this parameter failed to meet pharmacopoeial specifications (8) (9)and the authors justified that particle size and mixing of powders could be responsible.

1.2.3 Chemical Assay Assay of drug content is a global requirement. Typically a pooled sample of tablets (n=20) is analysed according to the monograph method; and the limits are set in each monograph in percentages of the labelled amount of the active constituent. Where samples smaller than 20 are taken the limits are tightened (6)and progress to content uniformity is dependent upon complying with assay. 1.2.4 Dissolution Dissolution tests have become one of the most used tests in the characterisation of drugs and the quality control of solid dosage forms. Although dissolution tests are mainly used as quality control to ensure end products or batch-to-batch consistency and to identify good and bad formulations, dissolution data may also be correlated with in vivo activity. Dissolution tests become especially important if dissolution is the rate limiting steps in drug absorption (10). Dissolution tests are therefore used to confirm compliance with confidential specifications and are needed as part of product license application (11). The general principle of dissolution tests is that the powder or solid dosage form is tested under uniform agitation in an attempt to achieve dissolution. Agitation is accomplished by either using a stirrer inside the apparatus or rotating the container holding the dosage form. Two general methods are included currently in USP and BP, the paddle and the basket methods [USPXXVII and BP2001]. 1.2.4.1 Dissolution Apparatus

1.2.4.1.1 Dissolution apparatus type I (Basket apparatus) The assembly consists of the following: a covered vessel made of glass or other inert, transparent material, a motor, a metallic drive shaft and a cylindrical basket. The vessel is

partially immersed in a suitable water-bath of any convenient size or placed in a heating jacket. The water-bath or heating jacket permits holding the temperature inside the vessel at 37 0.5 during the test and keeping the bath fluid in constant, smooth motion. No part of the assembly, including the environment in which the assembly is placed, contributes significant motion, agitation or vibration beyond that due to the smoothly rotating stirring element. An apparatus that permits observation of the specimen and the stirring element during the test is preferable. The vessel is cylindrically, with a hemispherical bottom and with a nominal capacity of 1000ml [USPXXVII and BP 2001]

Figure 1.1: Schematic diagram of dissolution apparatus type 1 (Basket apparatus)

1.2.4.1.2 Dissolution apparatus type II (Paddle apparatus) The assembly is the same as that of the apparatus I except that a paddle formed from a blade and a shaft is used as the stirring element. The shaft is positioned so that its axis is not more than 2mm at any point from the vertical axis of the vessel and rotates smoothly without

significant wobble. The vertical centre line of the blade passes through the axis of the shaft so that the bottom of the blade is flushed with the bottom of the shaft [USPXXVII].

Figure 1.2: Schematic diagram of dissolution apparatus type II (Paddle apparatus).

1.2.4.2

Factors affecting determination of dissolution rate

1.2.4.2.1 Instrumental factors One of the instrumental factors that affect dissolution is vibration [USPXXVII]. The United Stated Pharmacopoeia (USP) states that no part of the assembly, including the environment in which the assembly is placed, should contribute significant motion, agitation or vibration beyond that caused by a smoothly rotating element. There is little doubt that excessive vibration of the dissolution apparatus considerably increases dissolution rates (12). The other important instrumental factors that affect dissolution are the shaft and vessels. Limits should be set on shaft eccentricity to reduce the problem associated with variation in dissolution rate that may rise due to misalignment. The use of a hemispherical flask allows

some variation in construction. Minor changes in vessel shape may considerably alter the dissolution rate determined by the paddle method. Glass containers with an inside bottom flatter than specified gave a high bias in dissolution rate and those with an inside bottom of steeper curvature gave a low bias [USPXXVII]. Agitation speed also affects rates of dissolution. The speed should be uniform throughout the test time. For example, the dissolution rate planar constant release tablets were found to increase with increase in paddle rotation speed and increase in distance from the centre of the vessel bottom (13). 1.2.4.2.2 Temperature The dissolution fluids should be maintained at 37 0.5 . Even slight temperature variations may have a significant effect on tablet dissolution. A 1.5 change in temperature resulted in a 15% variation in the dissolution rates of salicylic acid waivers (11) .It is essential, therefore, to control the temperature of the water-bath continuously. 1.2.4.2.3 Dissolution media Ideally a dissolution medium should be formulated as close as possible to those of the invivo fluids; dissolution media based in 0.1M HCl are used to mimic gastric pH. This occurs despite evidence that the gastric pH in the majority of the population is the average of 1 to 3. Simulated gastric fluid is similarly used. Many compendial dissolution fluids are at a pH near neutral despite the fact that tablets, when swallowed, will meet a lower gastric pH. The use of surfactants and enzymes may also be a coarse approximation of the intestinal fluids, although surfactants may be included to increase drug solubility by solubilisation into micelles [USPXXVII]. To simulate in-vivo conditions it has been suggested that different materials (e.g., bovine serum albumin or milk be included in dissolution media) (11). As a quality control test, dissolution test leads to predictive values for the changes of dissolution rate on storage and, therefore, predicts the potential storage time of a product by estimating problems caused by aging. Delayed or erratic dissolution has been observed in sugar coated tablets of chlorpromazine and propantheline bromide [USPXXVII]. Sulfamethoxasol, trimethoprim and quinine also showed poor dissolution after storage for six months in simulated tropical storage condition (14). Tolbutamide tablets originating from different sources are also recommended to be interchanged with care since evaluation of such tablets showed different dissolution rates (15). Data retrieval is essential, not only for tablets but for all dosage forms requiring dissolution tests. Comparison with previous batches and earlier data on the same batch confirming the absence of aging effects is essential [USPXXVII].

1.3 Drug Quality Drugs play important role in improving human health and promoting well-being. However, to produce the desired effect they have to be safe, efficacious and of acceptable quality and have to be used rationally. The use of ineffective and poor quality drugs will not only endanger therapeutic treatment, but also erode public confidence in the countries health programmes. For that reason the production, storage, distribution and use of drugs in each country need to be regulated by the government. During the past few decades,

many pharmaceutical industries and distribution channels have flourished throughout the world; leading to an increase in the number of products circulating in the national and international markets. At the same time, however the presence of counterfeit and substandard drugs in those markets has increased substantially as a result of ineffective regulation of the manufacture and trade of pharmaceutical products by both exporting and importing countries (16). 1.3.1 1.3.1.1 Generic versus brand products and drug quality Bioavailability

The published guidelines on registration requirements to establish interchangeability of multisource (generic) pharmaceutical products (17), state that multisource products must satisfy the same standards of quality, safety and efficacy as those applicable to the corresponding innovator product. They recommend that quality attributes of a multisource product should be tested against the innovator product for which interchange is intended. The innovator product is usually the most logical comparator product because its quality, safety and efficacy should have been well assessed in pre- and post-marketing studies and, in addition, the data on its safety and efficacy are usually linked to a pharmaceutical product with defined specifications for quality and performance (18). Since the reference product would have been approved on the bases of clinical trials where the safety and efficacy are considered unnecessary but instead, a surrogate measure whereby the concentration of the drug in serum or plasma of human subject is considered more appropriate. Comparative bioavailability studies between test and reference products are involved in the assessment of bioequivalence between a generic and innovator product. This might confirm the absence of a significant difference in the rate and extent to which the active ingredient or inactive moiety in pharmaceutically equivalent dosage forms become available at the site of action when administered at the same molar dose under similar conditions in an appropriately designed study in human subjects (19). 1.3.1.2 Issues beyond bioavailability

Indeed substitution of generic drugs for brand name products is highly controversial and is often met with suspicion by health care providers and patients (20). Meredith conducted a MEDLINE search published between 1973 and 2003 and highlighted a number of different categories and patient subpopulations for which generic substitutions can still prove to be problematic (20). With this review the author recommended that health care providers should continue to exercise caution in the consideration of generic drug substitution under certain circumstances. FDA also recognises circumstance where generic substitution may not be appropriate, even though the generic and brand name products are considered to be therapeutically equivalent. The orange book (a list containing FDA Approved Drug Products with Therapeutic Equivalence) states that in those circumstances where the characteristics of a specific product other than its active ingredient, are important in the therapy of a particular patient, the physicians specification of that product is appropriate. For example, some patients might be confused by difference in color or shape of tablets,

10

while others might reduce compliance if they dislike the flavour of the substituted medication. A patient who is supposed to take a partial dose may face difficulties in splitting the pill if the generic version is scored differently. Some patients may be allergic to the coating of the generic substitute but not the coating of the brand name product. Some individuals may store their medications under adverse conditions that would affect the stability of the generic more than its brand name version (21). The other important factors that should be considered in the interchangeability of brand and generic products are the attitudes of providers, consumers and professional organisations as well as pharmaceutical firms to their products. Provider groups generally do not oppose generic substitution by pharmacists, as long as the prescribing physician maintains the right to limit such substitution when he or she believes it is medically inappropriate. On the other hand, from the study done to assess pharmacists individual and organisational views on generic medications, the authors concluded that despite pharmacists general acceptance of generic substitution, importance of therapeutic interchange, is likely to remain matters of concern for pharmacists and their professional organisations (22). Generic drug use is also a history of conflict from a variety of perspectives; in which manufacturers of brand name products abrasively seek to protect their patients from a variety of groups that want access to less expensive medications (23). Another conflict is professional, especially for the members of the pharmacy profession who view drug product selection as an important opportunity for pharmacists to use their professional judgement. The brand manufacturers, most of the time, suggest that not all products are bioequivalent and of the same quality. The pharmacy profession, generic drug manufacturers, health care institutions and the FDA have opposed this position. As care costs continue to rise, consumers, providers and policy makers need to assess the best way to keep health care affordable without adversely affecting care quality. With prescription costs serving as a major contributor to recent cost increases, generic drugs offer an important tool for reducing the rate of growth in overall health expenditures (5). Some studies also show the economic significance of interchange of generic products to brand name products more strongly than the bioequivalence and therapeutic outcomes(24). 1.3.2 Private Pharmacy and Drug Quality

The private sector is the prominent actor in the provision of pharmaceuticals particularly in developing countries. Private provision of drugs has been associated with risks regarding availability, affordability, rational use and drug quality. Ensuring an effective regulatory framework is therefore a major challenge for governments, yet the capacity of regulatory authorities is often out stripped by private sector growth. 1.4 Justification of studies The backbone of any good health care delivery system depends greatly on the quality of the drugs used for treatment and the number of qualified and competent pharmacists and

11

other health care personnel. These professionals should ensure that the drug market is effectively regulated and controlled according to international standards, and as well, carry out physical and chemical tests to ensure that each and every drug product conforms to the pharmacopoeial standards of purity, safety, efficacy and strength. In Sierra Leone, the number of trained pharmacists and other health professionals required to execute these functions is small (25). Many drugs enter the country illegally and these drugs are not tested for their purity and efficacy. It will therefore not be surprising if counterfeit, substandard and spurious pharmaceutical products are present in the countrys drug market in large proportions. Such drugs are usually produced by pharmaceutical companies that do not follow the tenets of Good Manufacturing Practice since their main aim is to maximise profit. The climatic conditions in tropical countries, particularly Sierra Leone, favour the rapid degradation of most pharmaceuticals especially if they are not kept under proper storage conditions. A good number of the drugs sold by pharmacy proprietors and petty traders are hardly kept under the recommended storage conditions. Therefore, the need for frequent analysis of these products for their potency should be in place.

1.5 Objectives From the prevailing situation of counterfeit and substandard products that circulate in the world market and from the attitude and understanding of pharmacists and their organisations toward generic and brand products, it is of prime importance to study the physicochemical properties of marketed pharmaceutical products. Hence, the general and specific objectives of this study are as follows: 1.5.1 General Objective

To evaluate the quality as well as the physicochemical equivalence of different brands of the anti-malarial drug combination, sulfadoxine-pyrimethamine imported into the Sierra Leonean market. 1.5.2 Specific Objectives To evaluate the physical properties such as hardness, friability and disintegration time of sulfadoxine-pyrimethamine tablets in comparison to specifications and requirements stated by the official compendia. To assay the contents of each tablets using official methods and inferring the quality of sulfadoxine-pyrimethamine products.

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CHAPTER TWO

2.0 Review of related literature 2.1 Malaria

Malaria is a mosquito-borne infectious disease of humans and other animals caused by protists (a type of microorganism) of the genus Plasmodium (phylum Apicomplexa). In humans, malaria is caused by P. falciparum, P. malariae, P. ovale, P. vivax and P. knowlesi.[26][27] Among those infected, P. falciparum is the most common species identified (~75%) followed by P. vivax (~20%).[28] P. falciparum accounts for the majority of deaths.[29] P. vivax proportionally is more common outside of Africa.[30] There have been documented human infections with several species of Plasmodium from higher apes; however, with the exception of P. knowlesia zoonotic species that causes malaria in macaques[27]these are mostly of limited public health importance.[31]
2.1.1Life cycle

The definitive hosts for malaria parasites are female mosquitoes of the Anopheles genus, which act as transmission vectors to humans and other vertebrates, the secondary hosts. Young mosquitoes first ingest the malaria parasite by feeding on an infected vertebrate carrier and the infected Anopheles mosquitoes eventually carry Plasmodium sporozoites in their salivary glands. A mosquito becomes infected when it takes a blood meal from an infected vertebrate. Once ingested, the parasite gametocytes taken up in the blood will further differentiate into male or female gametes and then fuse in the mosquito's gut. This produces an ookinete that penetrates the gut lining and produces an oocyst in the gut wall. When the oocyst ruptures, it releases sporozoites that migrate through the mosquito's body to the salivary glands, where they are then ready to infect a new human host. The sporozoites are injected into the skin, alongside saliva, when the mosquito takes a subsequent blood meal. This type of transmission is occasionally referred to as anterior station transfer.[32] Only female mosquitoes feed on blood; male mosquitoes feed on plant nectar, and thus do not transmit the disease. The females of the Anopheles genus of mosquito prefer to feed at night. They usually start searching for a meal at dusk, and will continue throughout the night until taking a meal.[33] Malaria parasites can also be transmitted by blood transfusions, although this is rare.[34]
2.1.2: Recurrent malaria

Malaria recurs after treatment for three reasons. Recrudescence occurs when parasites are not cleared by treatment, whereas reinfection indicates complete clearance with new infection established from a separate infective mosquito bite; both can occur with any malaria parasite species. Relapse is specific to P. vivax and P. ovale and involves re-emergence of blood-stage parasites from latent parasites (hypnozoites) in the liver.[28] Describing a case of malaria as cured by observing the disappearance of parasites from the bloodstream can, therefore, be deceptive. The longest incubation period reported for a P. vivax infection is 30 years.[35] Approximately one in five of P. vivax malaria cases in temperate areas involve overwintering by hypnozoites, with relapses beginning the year after the mosquito bite.[36]

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2.1.3: Signs and symptoms

The signs and symptoms of malaria typically begin 825 days following infection.[37] However, symptoms may occur later in those who have taken antimalarial medications as prevention.[28] The presentation may include fever, shivering, arthralgia (joint pain), vomiting, haemolytic anaemia, jaundice, hemoglobinuria, retinal damage,[38] and convulsions. Approximately 30% of people however will no longer have a fever upon presenting to a health care facility.[28] The classic symptom of malaria is cyclical occurrence of sudden coldness followed by rigor and then fever and sweating lasting about two hours or more, occurring every two days in P. vivax and P. ovale infections, and every three days for P. malariae. P. falciparum infection can cause recurrent fever every 3648 hours or a less pronounced and almost continuous fever.[39] For reasons that are poorly understood, but that may be related to high intracranial pressure, children with malaria frequently exhibit abnormal posturing, a sign indicating severe brain damage.[40] Cerebral malaria (encephalopathy specifically related to P. falciparum infection) is associated with retinal whitening, which may be a useful clinical sign in distinguishing malaria from other causes of fever.[41] Severe malaria is usually caused by P. falciparum, and typically arises 614 days after infection.[35] Non-falciparum species have however been found to be the cause of ~14% of cases of severe malaria in some groups.[28] Consequences of severe malaria include coma and death if untreatedyoung children and pregnant women are especially vulnerable. Splenomegaly (enlarged spleen), severe headache, cerebral ischemia, hepatomegaly (enlarged liver), hypoglycaemia, and hemoglobinuria with renal failure may occur. Renal failure is a feature of blackwater fever, where haemoglobin from lysed red blood cells leaks into the urine.[35]
2.1.4: Complications

There are a number of serious complications of malaria. Among these is the development of respiratory distress which occurs in up to 25% of adults and 40% of children with falciparum malaria.[42] The causes of this problem are diverse and include respiratory compensation of metabolic acidosis, noncardiogenic pulmonary oedema, concomitant pneumonia and severe anaemia. Acute respiratory distress syndrome (ARDS) may develop in 525% in adults and up to 29% of pregnant women but is rare in young children.
2.1.5: Medications

Several drugs, most of which are used for treatment of malaria, can be taken to prevent contracting the disease during travel to endemic areas. Chloroquine may be used where the parasite is still sensitive.[43] However, due to resistance one of three medicationsmefloquine (Lariam), doxycycline (available generically), or the combination of atovaquone and proguanil hydrochloride (Malarone)is frequently needed.[43] Doxycycline and the atovaquone and proguanil combination are the best tolerated; mefloquine is associated with higher rates of neurological and psychiatric symptoms.[43] The prophylactic effect does not begin immediately upon starting the drugs, so people temporarily visiting malaria-endemic areas usually begin taking the drugs one to two weeks before arriving and should continue taking them for four weeks after leaving (with the exception of atovaquone proguanil that only needs to be started two days prior and continued for seven
14

days afterwards). Generally, these drugs are taken daily or weekly, at a lower dose than is used for treatment of a person who contracts the disease. Use of prophylactic drugs is seldom practical for full-time residents of malaria-endemic areas, and their use is usually restricted to short-term visitors and travellers to malarial regions. This is due to the cost of purchasing the drugs, negative adverse effects from long-term use, and because some effective anti-malarial drugs are difficult to obtain outside of wealthy nations.[44] The use of prophylactic drugs where malaria-bearing mosquitoes are present may encourage the development of partial immunity.[45]
2.1.6: Treatment

The treatment of malaria depends on the severity of the disease; whether people can take oral drugs or must be admitted depends on the assessment and the experience of the clinician. Uncomplicated malaria Uncomplicated malaria may be treated with oral medications. The most effective strategy for P. falciparum infection is the use of artemisinins in combination with other antimalarials (known as artemisinin-combination therapy).[46] This is done to reduce the risk of resistance against artemisinin.[46] These additional antimalarials include amodiaquine, lumefantrine, mefloquine or sulfadoxine-pyrimethamine.[47] Another recommended combination is dihydroartemisinin and piperaquine.[47] In the 2000s (decade), malaria with partial resistance to artemisinins emerged in Southeast Asia.[48][49] Severe malaria Severe malaria requires the parenteral administration of antimalarial drugs. Until the mid-2000s the most used treatment for severe malaria was quinine, but artesunate has been shown to be superior to quinine in both children [50] and adults.[51][52] Treatment of severe malaria also involves supportive measures that are optimally performed in a critical care unit, including management of high fevers (hyperpyrexia) and the subsequent seizures that may result from it, and monitoring for respiratory depression, hypoglycaemia, and hypokalaemia.[53] Infection with P. vivax, P. ovale or P. malariae is usually treated on an outpatient basis (while a person is at home). Treatment of P. vivax requires both treatment of blood stages (with chloroquine or ACT) as well as clearance of liver forms with primaquine.[54] Whilst there is little argument that artemisinin-based combination treatments (ACTs) such as artemether-lumefantrine are effective in treating malaria, they are a lot more expensive than the treatments currently being used. The advocates of policy change are lobbying hard for the global financing mechanisms that will likely be necessary in order to subsidize the world's supply of ACTs [55]. The fervour of the advocates is intense [56], but national policiesparticularly the introduction of interventions that are in short supply and cost more to produceare unlikely to be implemented at the speed that a clinician treating an individual patient would desire. It is more likely that countries will explore the variety of strategic options open to them, which include ACTs as first-line treatment, as well as non-artemisinin combination treatments, such as amodiaquine with SP, and will examine the potential benefits and costs (57).

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With ACTs, the potential public health impact of reducing transmission is a factor to include in the evaluation of benefits versus costs, but there is little research on this impact in areas that are highly endemic for malaria. The finding of Sutherland and colleagues that ACTs also greatly reduce infectiousness will contribute to appropriate and informed decision-making for sustainable changes in treatment policy at the country level.

2.2Sulfadoxine-Pyrimethamine (58) 2.2.1Definition

Sulfadoxine-pyrimethamine is a combination antimalarial agent containing the sulphonamide antibacterial compound, sulfadoxine, and the anti-parasitic agent, pyrimethamine. Each tablet contains 500mg N1-(5, 6-dimethoxy-4-pyrimidinyl) sulphanilamide (sulfadoxine) and 25mg 2, 4-diamino-5-(p-chlorophenyl)-6-ethylpyrimidine (pyrimethamine). [Each tablet also contains pharmaceutical excipients, which vary from manufacturer to manufacturer. Structure: Sulfadoxine
H3CO H2N O2 H S N N OCH3 N

N1-(5, 6-dimethoxy-4-pyrimidinyl) sulphanilamide Pyrimethamine

H3CH2C N N NH2 Cl NH2

2, 4-diamino-5-(P-Chlorophenyl)-6-ethylpyrimidine
2.2.2: Pharmacology

Sulfadoxine and pyrimethamine sequentially block two enzymes involved in the biosynthesis of folinic acid within the malaria parasites. Thus, they are antifolates. Either drug by itself is only moderately effective in treating malaria, because the parasite, Plasmodium falciparum may be able to use exogenous folic acid, that is, folic acid which is present in the parasites environment, while in combination, the two substances have a synergic effect which outbalances that ability. The combination is considered to be more effective in treating malaria caused by P. falciparum than that caused by P. vivax for which chloroquine is considered more effective, though in the absence of a species-specific diagnosis the sulfadoxine-pyrimethamine combination may be indicated. Due to side effects, however, it is no longer recommended as a routine preventative, but only to treat serious malaria infections or to prevent them in areas where other drugs may not be effective.

16

2.2.3: Indications and usage

The sulfadoxine-pyrimethamine medication is indicated in the treatment of acute, uncomplicated P. falciparium malaria for patients in whom chloroquine resistance is suspected; malaria prophylaxis for travellers to areas where chloroquine resistant P. falciparium malaria is endemic and sensitive to sulfadoxine-pyrimethamine and when alternative therapy is not available or is contraindicated. Malaria prophylaxis with sulfadoxine-pyrimethamine is not routinely recommended.
2.2.4: Microbiology 2.2.4.1Pharmacodynamics

Sulfadoxine and pyrimethamine are folic acid antagonists. Sulfadoxine inhibits the activity of dihydropteroate synthase whereas pyrimethamine inhibits dihydrofolate reductase. Pyrimethamine is a synthetic diaminopyridine that inhibits the dihydrofolate reductase (DHFR) of plasmodia and thereby blocks the synthesis resulting in cell death. Sulfadoxine is a longacting sulphonamide, and it inhibits dihydropteroate synthase (DHPS), an enzyme that utilizes para-aminobenzoic acid in the synthesis of dihydropteroic acid. This enzyme is also a component of the folate metabolic pathway and is upstream of DHFR, the enzyme targeted by pyrimethamine. The combination of pyrimethamine and sulfadoxine thus offers a two-step synergistic blockade of plasmodial division.
2.2.4.2: Activity in-vitro

Sulfadoxine and pyrimethamine are active against asexual erythrocytic stages of P. falciparium. Sulfadoxine and pyrimethamine may also be effective against strains of P. falciparium resistant to chloroquine.
2.2.4.3: Drug resistance

Strains of P.falciparium with decreased susceptibility to sulfadoxine and/or pyrimethamine can be selected in vitro or in vivo. P. falciparium malaria that is clinically resistance to sulfadoxinepyrimethamine occurs frequently in parts of South-East Asia and South America, and is also prevalent in East and Central Africa. Therefore, sulfadoxine-pyrimethamine should be used with caution in these areas. Likewise, sulfadoxine-pyrimethamine may not be effective for treatment of recrudescent malaria that develops after prior therapy (or prophylaxis) with sulfadoxinepyrimethamine.
2.2.5: Dosage and Administration 2.2.5.1: Malaria prophylaxis

The first dose is taken 1 or 2 days before arrival in an endemic area; continued during the stay and for 4-6 weeks after return. The dosage regimen for adults is one tablet once per week or 2 tablets once every 2 weeks .The dosage for children older than 2 months to 18 years of age is based on body weight: one and a half tablet for those more than 45kg, one tablet for those 3145kg, three-fourth of a tablet for those21-30kg, half of a tablet for those 11-20kg and one-fourth of a tablet for those 5-10kg;all of which should be taken once weekly. It is generally advised that prophylactic therapy should not be continued for more than 2 years [58].

17

2.2.5.2: Malaria treatment

The dosage regimen for adults is a single dose of 2 to 3 tablets. The dosage for children older than 2 months to 18 years of age is based on body weight: three tablets for those more than 45kg, two tablets for those 31-45kg, one and a half tablets for those 21-30kg one tablet for those 1120kg, half of a tablet for those 5-10kg; all of which should be taken once weekly.
2.2.6Storage/ Stability

Tablets of sulfadoxine-pyrimethamine should be stored at 68 to 77 F in a dry place.

2.2.7: Drug Interactions

Antifolic drugs (e.g. sulphonamides, trimethoprim-sulfamethoxazole) should not be taken by patients receiving sulfadoxine-pyrimethamine. The incidence and severity of adverse reactions may be increased with chloroquine, compared with taking sulfadoxine-pyrimethamine alone. Also methenamine may increase the potential for formation of insoluble precipitates in the urine and methotrexate levels may increase with sulfadoxine-pyrimethamine, therefore the dose should be adjusted as needed. Tretinoin may augment photo toxicity when administered with sulfadoxine-pyrimethamine.
2.2.8: Adverse Reactions

Adverse reactions which involve the central nervous system include opathy, ataxia, convulsions, fatigue, hallucinations, headache, insomnia, mental depression, muscle weakness, nervousness, peripheral neuritis, polyneuritis, tinnitus and vertigo. The dermatologic adverse reactions are erythema multiforme, exfoliative dermatitis, generalized skin eruptions, Lyell syndrome, photosensitisation, pruritus, slight hair loss, Stevens-Johnson syndrome, toxic epidermal necrolysis and urticaria. In the gastrointestinal tract, sulfadoxine-pyrimethamine may cause abdominal pain, diarrhoea, emesis, feeling of fullness, glossitis, nausea, pancreatic and stomatitis. Genitourinary adverse reactions such as bilirubin and serum creatinine elevation, crystalluria, interstitial nephritis, renal failure, toxic nephrosis with oliguria and anuria may occur. Hematologic lymphatic adverse reactions like agranulocytosis, aplastic anaemia, eosinophilia, haemolytic anaemia, hypoprothrombinemia, leukopenia, megaloblastic anaemia, methemoglobinemia, purpura, serum sickness, and thrombocytopenia occur with sulfadoxinepyrimethamine. Also, the hepatic disorders such as hepatitis, hepatocellular necrosis and transient rise of liver enzymes occur with sulfadoxine-pyrimethamine. The hypersensitivity reactions allergic myocarditis and allergic pericarditis, and arthralgia a musculoskeletal disorder are adverse reactions of sulfadoxine-pyrimethamine. Respiratory disorders such as pulmonary infiltrates resembling eosinophilic or allergic alveolitis and other reactions such as anaphylactoid reactions, chills, drug fever, lupus erythematosus phenomenon and periarteritis nodosa may also occur with sulfadoxine-pyrimethamine.
2.2.9: Precautions

Fatalities will occur because of severe reactions including Stevens-Johnson syndrome. Sulfadoxine-pyrimethamine prophylaxis should be discontinued at the first appearance of skin rash, if a significant reduction in the count of any formed blood elements is noted, or upon the occurrence of active bacterial or fungal infections. Regularly schedule liver enzyme test, analysis of urine for crystalluria should be monitored. Sulfadoxine-pyrimethamine is

18

contraindicated in pregnancy at term because it crosses the placenta, and in lactating women since it is excreted in breast milk causing kernicterus. Sulfadoxine-pyrimethamine should not be given to infants younger than two months of age because inadequate development of the glucuronide-forming enzyme system. In the elderly this drug should be used with caution because of the greater frequency of decreased hepatic, renal or cardiac function and concomitant diseases or other drug therapy. Sulfadoxine-pyrimethamine should also be used with caution in patients with renal or hepatic impairment, possible folate deficiency, severe allergy or bronchial asthma, or G-6-PD deficiency. Excess sun exposure should be avoided and if signs of folic acid deficiency occur, the treatment should be discontinued. When recovery of the depressed platelets or white blood cell counts is too slow, folinic acid may be administered. Fatalities will occur because of fulminant hepatic necrosis, agranulocytosis, aplastic anaemia and other blood dyscrasias.

2.2.10: Over dosage

In over dosage, symptoms such as anorexia, central nervous system stimulation (including convulsions), crystalluria, glossitis, headache, leukopenia, megaloblastic anaemia, nausea, thrombocytopenia and vomiting may occur.

2.2.11: Pharmacokinetics (58)

2.2.11.1: Absorption After administration of 1 tablet of sulfadoxine-pyrimethamine, peak plasma levels for pyrimethamine (approximately 0.2mg/L) and for sulfadoxine (approximately 60mg/L) are reached after about 4hrs. 2.2.11.2: Distribution The volume of distribution for sulfadoxine and pyrimethamine is 0.14L/Kg and 2.3 L/Kg, respectively. Patients taking 1 tablet a week (recommended adult dose for malaria prophylaxis) can be expected to have mean steady state plasma concentration of about 0.15mg/L for pyrimethamine after about four weeks and about 98mg/L for sulfadoxine after about seven weeks. Plasma protein binding is about 90% for both pyrimethamine and sulfadoxine. Both pyrimethamine and sulfadoxine cross the placental barrier and pass into breast milk.
2.2.11.3 Metabolism

About 5% of sulfadoxine appears in the plasma as acetylated metabolite, about 2% to 3% as the glucuronide. Pyrimethamine is formed to several unidentified metabolites.
2.2.11.4 Elimination

A relatively long elimination half-life is characteristic of both sulfadoxine and pyrimethamine. The mean values are about 100h for pyrimethamine and about 200h for sulfadoxine. Both pyrimethamine and sulfadoxine are eliminated mainly via the kidneys.

19

2.2.12 Characteristics in Patients

In malaria patients, single pharmacokinetic parameters may differ from those in healthy subjects, depending on the population concerned. In patient with renal insufficiency, delayed elimination of sulfadoxine and pyrimethamine must be anticipated.

20

CHAPTER THREE

EXPERIMENTAL

3.1 MATERIALS USED IN THIS STUDY

3.1.1 INSTRUMENTS The following instruments were used in this study: i) ii) iii) iv) Sartorius analytical balance (serial no. EB2245) Lambda 35 UV/VIS spectrometer (Perkin Elmer) pH Meter Orion SN 57019 [Move Plan (USA), Inc.] Dissolution apparatus DISS-51939 42 Bath Model 5100 SN 5150581 apparatus (DISTEK).

3.1.2 CHEMICALS AND REAGENTS USED i) ii) iii) iv) v) vi) Chloroform (LAB-SCAN, analytical science) Hydrochloric acid Distilled water Ethanol solution (96%)[RADCHEM] Potassium dihydrogen phosphate [RADCHEM] Sodium hydroxide (GN International, New Delhi)

3.2 METHODOLOGY 3.2.1 LOCATION OF STUDY This study was conducted in the Western Area of Freetown. The data and samples of drugs for this study were collected from different pharmacies in Freetown. The Quality Control and Pharmaceutical Laboratory at the Pharmacy Board of Sierra Leone, New England, Freetown was used for analysis of the drugs samples collected. 3.2.2 DATA COLLECTION AND SAMPLE SIZE Data on the labels of different brands of sulfadoxine pyrimethamine was collected from different pharmacies in the Western Area. Also, 72 tablets of each brand were procured. Table 1 shows a tabulated list of the different brands of sulfadoxine-pyrimethamine that were used in this study.

21

Table 3.2-1: List of Sulfadoxine- Pyrimethamine Tablets Procured No Brand or Trade Name Malareich
R

Dosage form Strength

and

Manufacturer and country of Origin

Batch No or Lot

Manufac- Expiry turing Date Date

Tablet: Sulfadoxine 500mg and pyrimethamine -25mg

MalafanT
M

MaladarT
M

Spymal

Tablet: Sulfadoxine 500mg and Pyrimethamine -25mg Tablet: Sulfadoxine500mg and Pyrimethamine -25mg Tablet: Sulfadoxine500mg and Pyrimethamine-25mg

Medreich limited 4/3 Avalahalli, Off Kanakapura Road, 690324 Bangalore-560069. India Kinapharma Limited North 11005 Industrial Area Accra, Ghana Ernest Chemists Limited P.O. Box 0107K 3345 Accra, Ghana JBM Pharmaceuticals G-17,Meghal Indl.Estate, Devidayal Rd, Mulund(W),Mumba e-400080 India Mihika Pharmaceuticals Mumbai-400080 India Bliss GVS Pharmaceutical Ltd 10 DewanUdyog Nagar Alijali Palybor India Emzor Pharmaceutical Industries Ltd. Plot 3C Block A Isolo Lagos Nigeria Kinapharma Ltd North Ind Area Accra Ghana

12/2009

12/2012

04/2011

04/2015

7/2010

7/2014

JM-64

01/2011

12/2013

Antim

Tablet: sulfadoxine500mg of pyrimethamine-25 mg Tablet: Sulfadoxine500mg and Pyrimethamine -25mg Tablet: Sulfadoxine500mg and Pyrimethamine -25mg Tablet: Sulfadoxine500mg and Pyrimethamine -25mg

MP-1101 10/2011

09/2014

Palodoxin eTM

BG-06

12/2011

11/2014

Maldox

3227Q

07/2011

07/2014

Malafan

11013

12/2011

12/2015

22

(a) Malareich

(b) Maldox

(b) Maladar

(d) Malafan

Palodoxine

(e) Antim *Free health care drug for malaria.

(f) Palodoxine*

Plate 3.1: Photographs of packages of some samples of sulfadoxine-pyrimethamine

23

3.3 CONDITIONS OF STORAGE OF DRUG SAMPLES COLLECTED All the drug samples collected were kept in a dry place and at a temperature not exceeding 25C. The drugs were also kept in an atmosphere free from sunlight. The essence of this proper storage conditions is to maintain the stability of the active ingredient present in the sulfadoxine pyrimethamine tablets.

(a) Dissolution Apparatus

(b) Syringe and needle

Separating funnel

Retort stand and clamp

(c) Filter paper, funnel and conical flask

(d) Separating funnel & Retort stand and clamp

24

(e) Sartorius analytical balance

(f) Beaker

(g) Separating funnel

(h) 1000ml volumetric flask

(i)100 ml volumetric flask

(j) Measuring cylinder

25

(k) Magnetic hot plate

(l) Lambda 35 UV/VIS Spectrometer

Plate 3.2: Pictures of Apparatus Used.

3.4 PHYSICO-CHEMICAL TESTS The quality of the various brands of sulfadoxine pyrimethamine tablets was determined by performing the following tests: 3.4.1 ASSAY Assay of the contents of sulfadoxine-pyrimethamine tablets of each brand were analyzed using the method described in the Laboratory In-house Method of the Quality Control and Pharmaceutical Laboratory at the Pharmacy Board of Sierra Leone. 3.4.1.1 APPARATUS

The Lambda 35 UV/VIS spectrometer [Plate 3.2(l)] from the Quality Control Pharmaceutical laboratory at the Pharmacy Board of Sierra Leone was used to measure the absorbance of the different brands of sulfadoxine-pyrimethamine tablets. 3.4.1.2 METHOD DESCRIPTION (59)

Twenty tablets of each brand of sulfadoxine-pyrimethamine was ground into a powder form using a mortar and pestle. An amount of the powdered tablets of sulfadoxinepyrimethamine equivalent to 0.0656lg of sulfadoxine was weighed in an electronic
26

balance and suspended in 40ml of 0.1M NaOH in a separating funnel. The solution was shaken well to dissolve the sulfadoxine-pyrimethamine and extracted with five portions of 30ml chloroform. 10ml of 0.1M NaOH was poured into another separating funnel which was used for each chloroform extract. After addition of 30ml chloroform, the mixture was shaken well and on allowing to stand, two layers were formed. The upper layer being the aqueous layer (NaOH) which contains sulfadoxine (milky layer) and the bottom layer is the chloroform layer in which pyrimethamine is soluble (colourless layer). The chloroform layer was separated from the aqueous layer into a beaker and was evaporated to dryness. The aqueous layer was poured into an l00ml volumetric flask and diluted to volume with 0.01M NaOH. 7ml of the resulting solution was pipetted into another 100ml volumetric flask and diluted to volume with 0.01M NaOH. 40ml of 0.1M HCl was added to the beaker from which the chloroform was evaporated and heated for 5mins to dissolve the pyrimethamine. This solution was transferred into a 100ml volumetric flask and diluted to volume with 0.1M HCl. 25 ml of the resulting solution was pipetted into a 50ml volumetric flask and diluted to volume with 0.1 M HCl. The separated samples were assayed for sulfadoxine and pyrimethamine by UV/VIS spectrometric technique at 272nm using 0.0lM NaOH as blank. The amount of sulfadoxine dissolved in 30mins was calculated using 762 as absorbance [A (lcm,1%)] and that of pyrimethamine dissolved in 30 mins was calculated using 320 as absorbance [A (lcm,1%)]. 3.4.2 UNIFORMITY OF WEIGHT This test is used as a simple means of estimating the content of active ingredients per tablet. It is based upon the assumption that the active ingredients are homogenously dispersed throughout the contents of a batch of tablets. 3.4.2.1 APPARATUS

The uniformity of weight test was carried out using the Sartorius analytical balance, serial no. EB2245. 3.4.2.2 METHOD DESCRIPTION

The uniformity of weight test for sulfadoxine-pyrimethamine tablets was carried out by weighing a total of 20 tablets from each batch and then weighing them individually. The average weight of the 20 tablets was calculated and hence the limit range was determined followed by the percentage deviation (60).

27

Limit Range Lower limit Upper limit = =

Percentage deviation Low limit =

Upper limit

3.4.3 DISSOLUTION TEST This test determines whether the sulfadoxine-pyrimethamine tablets dissolve or go into solution within the prescribed time (30 mins) (59) when placed in a liquid medium (phosphate buffer) under the prescribed experimental conditions. 3.4.3.1 APPARATUS

The dissolution test was performed using the dissolution apparatus DISS-51939-42, Bath Model 5100 SN 5150582 see [Plate 3.2(a)]. 3.4.3.2 PREPARATION OF PHOSPHATE BUFFER SOLUTION

8.4g of sodium hydroxide and 54.44g of potassium hydrogen phosphate were weighed. The sodium hydroxide was dissolved in distilled water in a 200ml beaker, poured into a 1000ml volumetric flask and diluted to volume with distilled water to form a stock solution of sodium hydroxide. The stock solution of potassium hydrogen phosphate was prepared by dissolving the potassium hydrogen phosphate in distilled water, transferring it into a 1000ml volumetric flask and diluting to volume with distilled water. The resulting solution was poured into a bottle and further diluted to 2000ml. The buffer solution (6000ml) of pH 6.8 was prepared by mixing 672 ml of sodium hydroxide stock solution, 1500ml of potassium hydrogen phosphate stock solution and 3828ml of water. The pH of the buffer solution was then measured after its preparation. 3.4.3.3 MEASUREMENT OF THE pH OF PHOSPHATE BUFFER SOLUTION

The pH of the phosphate buffer solution was measured using a pH Meter Orion SN: 57019. The pH meter was first calibrated using standard solutions of pH 4.01, 7.01 and 10.01 by dipping the electrode of the pH meter into these solutions and swirling it. After calibration, the electrode of the pH meter was placed into the buffer solution to measure

28

the pH of the solution. In cases wherein the pH of the buffer solution was below 6.8, few drops of sodium hydroxide were added to increase the pH to 6.8. However there were also cases in which the pH was above 6.8 and few drops of phosphoric acid were added to decrease the pH to 6.8. 3.4.3.4 METHOD DESCRIPTION (59)

900ml of the phosphate buffer solution was poured into each of the six vessels of the dissolution apparatus II and allowed to equilibrate to 37.0C. After equilibrating, a weighed tablet was placed into each vessel of the apparatus and was allowed to dissolve for 30 minutes at 75 rpm. The temperature of each vessel was recorded before, during and after the test. At the end of the dissolution process some amount of the solution was filtered into a conical flask using a filter funnel with a filter paper. 25ml of the filtrate was pipetted into a separating funnel containing 20ml of 0.1M sodium hydroxide and shaken well to dissolve. The resulting solution was extracted with five portions of 30ml chloroform and the combined chloroform extracts was washed with 10ml of 0.1M NaOH. The combined chloroform layer was transferred into a beaker and evaporated to dryness. 40ml of 0.1M HCl was added to the beaker and heated for 5mins to dissolve the pyrimethamine. The solution was transferred into a 100ml volumetric flask and diluted to volume with 0.1M HCl. The absorbance was measured at 272nm using 0.1M HCl as blank and the amount of pyrimethamine dissolved in 30mins was calculated using 320 as absorbance [A (1cm,1%)]. The combined aqueous layer was transferred into a 100ml volumetric flask and diluted to volume with 0.1M NaOH. 1ml of the resulting solution was pipetted into another 100ml volumetric flask and diluted to volume with 0.1M NaOH. the absorbance was measured at 272nm using 0.1M NaOH as blank and the amount of sulfadoxine dissolved in 30mins was calculated using 762 as absorbance [A (1cm,1%)].

3.4.4 FRIABILITY TEST The friability of a tablet is an expression of how easily the tablet breaks up or how easily it crumbles. It is expressed as a percentage loss in weight for the number of tablets tumbling in a short cylinder for a specified number of rotations. 3.4.4.1 APPARATUS

Friability measurements were carried out by the use of the Vankel Friabilator with a serial number 4-1310-0798. 3.4.4.2 METHOD DESCRIPTION

29

Twenty tablets of each brand of sulfadoxine-pyrimethamine were weighed by the use of an electronic balance and placed into the drum of the Vankel Friabilator. This weight was designated as the initial weight of the tablet. The drum was allowed to rotate 100 times for 4mins, after which the tablets were removed. The tablets were cleared from lose dusts and broken particles. The tablets were reweighed and this weight was designated as the final weight of the tablets. The percentage weight loss was calculated using equation 1.1, Friability (% weight loss) =

Where:

= =

30

CHAPTER FOUR

RESULTS

4.1 Weight Uniformity of tablets of eight brands of Sulfadoxine-pyrimethamine

Weight of 20 tablets Average weight of one tablet Table 4.1: Weight of 20 individual tablets of eight brands of Sulfadoxine-pyrimethamine
Tablet

Malareich 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 0.6568 0.6856 0.6718 0.6646 0.6896 0.6820 0.6729 0.6781 0.6631 0.6769 0.6797 0.6701 0.6776 0.6720 0.6759 0.6770 0.6721 0.6772 0.6836 0.6776

Maladar 0.6316 0.6358 0.6465 0.6487 0.6354 0.6306 0.6276 0.6348 0.6294 0.6390 0.6359 0.6332 0.6323 0.6654 0.6256 0.6366 0.6307 0.6381 0.6447 0.6333

Brand/ Weight of Individual Tablets (g) Spymal Malafan Malafan Antim BN:11005 BN:11013 0.6205 0.5929 0.5995 0.5913 0.6156 0.5983 0.5949 0.6281 0.6058 0.6039 0.6076 0.5716 0.6105 0.6079 0.6001 0.6047 0.6231 0.6046 0.6022 0.5762 0.6020 0.6025 0.6017 0.5993 0.5999 0.6109 0.5974 0.5928 0.6294 0.6010 0.5957 0.5854 0.6215 0.5959 0.5979 0.6012 0.6378 0.6032 0.5925 0.6040 0.5984 0.6083 0.6217 0.5828 0.5958 0.5962 0.6028 0.6063 0.6285 0.5941 0.6052 0.5508 0.6256 0.5881 0.6100 0.5931 0.6261 0.5908 0.5978 0.6014 0.6122 0.6120 0.5955 0.5838 0.5982 0.5957 0.5857 0.5957 0.6228 0.5952 0.5942 0.5918 0.6310 0.6000 0.6067 0.5974 0.6128 0.6062 0.6118 0.6031

Palodoxine 0.6192 0.6193 0.6192 0.6169 0.6232 0.6100 0.6163 0.6165 0.6165 0.6098 0.6228 0.6233 0.6198 0.6292 0.6128 0.6262 0.6180 0.6243 0.6153 0.6329

Maldox 0.6327 0.6376 0.6208 0.6371 0.6293 0.6177 0.6177 0.6162 0.6067 0.6127 0.6089 0.6346 0.6285 0.6263 0.6215 0.6133 0.6154 0.6423 0.6340 0.6204

Result of the weight of 20 individual tablets: [

31

Finding the limit range

( )

Let average weight of tablet

0.95xg

g Limit range [ ]

Calculation of percentage deviation Percentage deviation Lower limit

Upper limit

32

Percentage deviation

*(

)+

Table 4.2: Weight Uniformity test results for eight brands of Sulfadoxine-pyrimethamine No Sample MalareichR MalafanTM BN:11005 MalafanTM BN:11013 Spymal PaladoxineTM Antim Maladar TM Maladox Average weight of 20 tablets (g) 0.675105 0.600905 0.601785 0.616215 0.61975 0.593105 0.636925 0.62402 Percentage Deviation (%) negative positive

1. 2. 3. 4. 5. 6. 7. 8.

Table 4.2 shows the weight uniformity test results for 8 different brands of sulfadoxinepyrimethamine tablets. Seven out of eight brands have a percentage deviation at Only the Antim brand had a percentage deviation at

7.50 7.00 6.50 6.00 5.50 5.00 4.50 4.00 3.50 3.00 2.50 2.00 1.50 1.00 0.50 0.00 -0.50 -1.00 -1.50 -2.00 -2.50 -3.00 -3.50 -4.00 -4.50 -5.00 -5.50 -6.00 -6.50 -7.00 -7.50

5.9 4.47 3.5 2.15 1.85 2.12 % Deviation(-ve) % Deviation(+ve) -2.71 -2.13 -1.78 -3.31 -1.61 -1.54 -2.78 3.31 2.92

-7.13

Figure 4.1 Percentage weight deviation bar chart of eight brands of sulfadoxine-pyrimethamine

33

4.2 Assay (% content) of eight brands of sulfadoxine- pyrimethamine 4.2.1 Calculation of the amount of powdered sulfadoxine used in assay

From the procedure, 0.06561g equivalent of powdered sulfadoxine was weighed. Weight of powdered sulfadoxine-pyrimethamine

The actual weight of powdered tablet (T) used was Three samples of each brand of sulfadoxine-pyrimethamine were assayed: S1, S2 and S3. S1 S2 S3

Average absorbance of the three samples: S1 S2 S3 Calculation of the content of sulfadoxine in 1 tablet [ ( Sample 1 ( ) Average absorbance ( )

)]

34

Sample 2 ( ) Average absorbance ( )

Sample 3 ( ) Average absorbance ( )

Calculation of percentage content of sulfadoxine in 1 tablet: ( But, final nominal concentration (FNC) is: 0.06561g of sulfadoxine was diluted to 100ml with 0.01MNaOH Thus the concentration of sulfadoxine is 0.06561 1ml of 0.06561 The dilution factor was diluted to 100ml with 0.01M NaOH 100 )

Therefore the final nominal concentration

From equation 4.6, percentage content is : Sample 1 ( ) Content of sample 1= FNC

35

Sample 2 ( ) Content of sample 2 FNC

Sample 3 ( ) Content of sample 3 FNC

=
limit for in-house method
4.2.2

Calculation of the amount of powdered pyrimethamine used.

Weight of powdered sulfadoxine-pyrimethamine used Amount of pyrimethamine in 1 tablet

36

Average weight of tablet

The actual amount of pyrimethamine used was Three samples of pyrimethamine were assayed, Average absorbance of the three samples: and .

content is: Sample 1 ( ) Average absorbance ( )

Sample 2 ( ) Average absorbance ( )

Sample 3 ( ) Average absorbance ( )

37

Calculation of percentage content of Pyrimethamine in 1 tablet: From equation 4.6, % content is: But, final nominal concentration (FNC) is: Pg. of pyrimethamine was diluted to 100ml with 0.01M HCL Thus the concentration of sulfadoxine was P 25ml of P was diluted to 100ml with 0.01M HCL

The dilution factor

Therefore the final nominal concentration

FNC

Percentage content

Sample 1 ( ) Content of sample 1 FNC

Sample 2 ( ) Content of sample 1 FNC

38

Sample 3 ( ) Content of sample 1 FNC

39

Table 4.3: Assay Results (% Content) No Brand %Sulfadoxine 1. 2. 3. 4. 5. 6. 7. 8. MalareichR MalafanTM BN:11005 MalafanTM BN:11013 Spymal PalodoxineTM Antim MaladarTM Maldox 97.98 100.37 101.56 97.82 99.32 98.06 96.98 97.42 Percentage Content %Pyrimethamine 101.97 98.78 97.48 105.26 95.99 95.57 97.87 91.83

From the above table, all of the samples of sulfadoxine-pyrimethamine passed assay test according to the percentage content limit of the in-house method [limit 90%-110.0%].

110 105 100 95 90 85

% Content of Sulfadoxine % Content of Pyrimethamine

Figure 4.3: Percentage content of sulfadoxine and pyrimethamine in eight brands of sulfadoxinepyrimethamine

40

4.3 Dissolution of eight brands of sulfadoxine-pyrimethamine 4.3.1 Calculation of the concentration of Sulfadoxine of sulfadoxine was dissolved in buffer of 900ml of sulfadoxine was dissolved in 100ml of buffer

100ml of 25ml of buffer contains 0.05555555556 buffer contains

Six vessels were used and three absorbance were obtained for each vessel Calculation of the average absorbance of sulfadoxine: G H I J K L

Calculation of the amount of sulfadoxine dissolved: [ ( )]

41

Vessel ( ) Average absorbance of sample ( ) Amount dissolved Vessel ( ) Average absorbance of sample ( ) Amount dissolved Vessel ( ) Average absorbance of sample ( ) Amount dissolved Vessel ( ) Average absorbance of sample ( ) Amount dissolved Vessel ( ) Average absorbance of sample ( ) Amount dissolved Vessel ( )

Average absorbance of sample ( ) Amount dissolved

42

4.3.1

Calculation of the percentage amount of sulfadoxine dissolved:

But final FNC is: 1ml of 0.01388888889% was diluted to 100ml with 0.1MHCl

Percentage amount dissolved: Vessel ( ) Amount dissolved

Vessel (

Amount dissolved

Vessel (

Amount dissolved

43

Vessel (

Amount dissolved

Vessel (

Amount dissolved

Vessel (

Amount dissolved

44

[ NLT

4.3.2

Calculation of the concentration of pyrimethamine

of pyrimethamine was dissolved in 900ml of of sulfadoxine was dissolved in 100ml of buffer

buffer

100ml of 25ml of buffer contains buffer contains

Six vessels were used and three absorbance were obtained for each vessel Calculation of the average absorbance of pyrimethamine:

Q R

45

Calculation of the amount of pyrimethamine dissolved: [ ( )]

Vessel ( ) Average absorbance of sample ( ) Amount dissolved Vessel ( ) Average absorbance of sample ( ) Amount dissolved Vessel ( ) Average absorbance of sample ( ) Amount dissolved

Vessel ( ) Average absorbance of sample ( ) Amount dissolved

Vessel ( ) Average absorbance of sample ( ) Amount dissolved

46

Vessel ( ) Average absorbance of sample ( ) Amount dissolved

Calculation of the percentage amount of Pyrimethamine dissolved:

But final FNC is: 0.00069444444% Percentage amount dissolved: Vessel ( )

Amount dissolved

Vessel (

Amount dissolved

47

Vessel (

Amount dissolved

Vessel (

Amount dissolved

Vessel (

Amount dissolved

48

Vessel (

Amount dissolved

( [ NLT ]

Table 4.4: Dissolution (% amount dissolved) of eight brands of sulfadoxine-pyrimethamine

No

Brand %Sulfadoxine MalareichR MalafanTM BN:11005 MalafanTM BN:11013 Spymal PalodoxineTM Antim MaladarTM Maldox

Percentage Content %Pyrimethamine 86.88 86.13 90.78 16.08 31.44 52.69 141.05 36.69

1. 2. 3. 4. 5. 6. 7. 8.

107.56 95.07 98.93 30.98 81.50 69.95 99.50 47.06

49

From the above table, 4 out of 8 samples of the sulfadoxine-pyrimethamine tablet passed the dissolution test according to the percentage amount dissolved limit of the in-house method [NLT 65%]. For 2 of the 8 samples (Palodoxine and Antim), only the sulfadoxine active ingredient passed the test whilst the pyrimethamine failed. Maldox and Spymal failed the test for both sulfadoxine and pyrimethamine.

160 140 120 100 80 60 40 20 0 % Amount of Sulfadoxine Dissolved % Amount of Pyrimethamine Dissolved

Figure4.3: Percentage amount dissolved of sulfadoxine and pyrimethamine in eight brands of

sulfadoxine-pyrimethamine

4.4 Friability of eight brands of sulfadoxine-pyrimethamine Weight of 20 tablets before friability (W1) Weight of 20 tablets after friability (W2) ( )

50

Table 4.5: Percent friability of eight brands of Sulfadoxine-pyrimethamine* No Brand Initial weight of Sample (g) 13.5217 12.0713 11.9715 11.0402 12.4380 11.6145 12.7697 Final Weight of sample (g) 13.4976 11.9945 11.8936 11.0057 12.3843 11.5499 12.7135 12.5603 % loss

1. 2. 3. 4. 5. 6. 7.

Malareich Malafan BN:11005 Malafan BN:11013 Spymal Palodoxine Antim Maladar

0.18 0.64 0.65 0.31 0.43 0.56 0.44 0.56

Maladox 12.6304 8. * Number of tablets = 20

From the above table the percentage friability test results of the different samples of sulfadoxinepyrimethamine is below 1.0%

Friabilty (% loss)
0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 Friabilty (% loss)

Figure 4.4 Friability of eight brands of sulfadoxine-pyrimethamine

51

4.5 DESCRIPTION OF TABLETS AND PACKAGING MATERIALS 4.5.1 Malareich Tablet: Malareich is a white circular uncoated tablet with the inscription malareich on one side and the logo M on the other side. Primary packaging: This is a grey aluminium stripped pack containing tablets. It has the Malareich and the logo on the front face and a diagram of mosquitoes, the code: SB102M, manufacturing date, expiry date and batch number on the other side. Secondary packaging: It is a white, blue and pink rectangular folding box made of card board with the inscription malareich and the logo on both surfaces. It also has other information on the drug written on it. There is a leaflet in the box that gives information about the tablet. 4.5.2 MALAFAN Tablet: Malafan is a white circular uncoated tablet scored on one side and the inscription malafan on the other side. Primary Packaging: This is an aluminium strip pack with diagrams of mosquitoes and malafan tablet content inscribed on both sides. Secondary Packaging: It is a white rectangular paper packet with the name of the tablet and diagram of a mosquito inscribed on both sides. 4.5.3 MALADAR Tablet: It is a white circular tablet scored on one side with the inscription Maladar on the scored side and ECL on the other side. Primary Packaging: This is an aluminium strip pack with mosquito diagrams and the inscription Maladar on one side, and Maladar and the tablet content inscribed on the other side. Secondary Packaging: It is a brown and white rectangular folding box made of card board with the inscription MaladarTM on both sides. There is a leaflet inside the box which contains information on the tablet. 4.5.4 SPYMAL Tablet: Spymal is a round circular uncoated tablet which is white on one side and pink on the other side. It is scored on the pink side. Primary Packaging: It is a thermoform blister pack containing three tablets. Secondary Packaging: This is a green and white rectangle folding box made of card board. The name of the tablet and a diagram of a mosquito is inscribed on one side and information on the table on the other side.
52

4.5.5 ANTIM Tablet: Antim is a white circular uncoated table which is scored on one side.

Primary Packaging: This is a thermoformed blister pack containing three (3) tablets. Secondary Packaging: It is a mauve and white rectangular folding box made of cardboard bearing the name of the product, logo, the diagram of a mosquito and other information on both sides. Inside the box there is a white paper leaflet bearing information on the product. 4.5.6 MALDOX Tablet: Maldox is a white circular uncoated tablet which is scored on one side. It has the inscription MALDOX on the scored side and EMZOR on the side. Primary Packaging: It is a thermoform blister pack with an aluminium foil containing the name of the product, diagrams of mosquitoes and other information on the product. Secondary Packaging: This is a green rectangular folding box made of card board bearing the name of the product, a mosquito diagram, tablet content and other information on both sides. Inside the box, theres a paper leaflet which contains information on the product. 4.6 INSPECTION OF CONTAINERS FOR LABELLINGS Each of the containers of the 8 brands of sulfadoxine-pyrimethamine was inspected for correct labelling. The specification in the BP for labelling was strictly followed. The specifications (61) include: a) Name of the drug product. b) A list of active ingredient showing the amount of each present. c) The batch number assigned by the manufacturer. d) The manufacturing and expiry date. e) Any special storage conditions or handling precautions that may be necessary. f) Directions for use, and warnings and precautions that may be necessary . g) Name and address of the manufacturer or the company or person responsible for placing the product on the market.

53

DISCUSSION

The aim of this project was to evaluate the quality as well as the physicochemical equivalence of eight brands of the anti-malarial drug combination, sulfadoxinepyrimethamine imported into the Sierra Leonean market. In the weight uniformity test, the various percentages of deviation are given in Table 4.2. The BP(61) states that for sulfadoxine-pyrimethamine tablets with a dose of 250mg or more, not more than two of the individual tablet weight should deviate from the average weight by more than and none by more than . An examination of table 4.2 revealed that none of the brands of sulfadoxine-pyrimethamine exceeded the specified limit of the B.P The result obtained for the chemical assay tests of eight brands of sulfadoxinepyrimethamine tablets (Table 4.3) revealed that all of the eight brands conformed to the requirement as specified in the In-house method. According to the In-house method (59) each tablet of sulfadoxine-pyrimethamine should contain not less than 90.0% and not more than 110.0% of the stated amount of active ingredients (sulfadoxine and pyrimethamine) present. None of the eight brands deviated outside the limit of 90.0% to 110.0%. This shows that the samples are within In-house standard in relation to the active ingredient present. The dissolution test result (Table 4.4) indicated that four out of the eight brands of sulfadoxine-pyrimethamine tablets tested conformed to the requirement specified in the In-house standard. According to the in-house method (59), each tablet of sulfadoxinepyrimethamine dissolved should be not less than 65% of the stated amount of active ingredients (sulfadoxine and pyrimethamine). None of the four brands deviated out of the limit of not less than 65%. This shows that these brands are within pharmacopoeial standard in relation to the dissolved active ingredient. However, the other four brands after repeated tests (repeated twice) failed to comply to the pharmacopoeial requirements of active ingredients of sulfadoxine-pyrimethamine tablets. This shows that the four brands are substandard in terms of the dissolution of the active ingredients present in each. According to Table 4.4 the eight different brands of sulfadoxine pyrimethamine tablet had friability (%loss) values less than 1.0%. According to the pharmacopoeial forum (62) , tablets with friability of less than 1.0% should disintegrate within 15 mins. This shows that the tablets will disintegrate within the stipulated time for tablets. All of the eight brands of sulfadoxine-pyrimethamine tablets inspected were appropriately labelled. The labelling on each packaging of the drug product inspected strictly followed the B.P (61) specifications of labelling of finished drug products.

54

CONCLUSIONS AND RECOMMENDATIONS

6.1 CONCLUSIONS From the data of the various physical and chemical tests carried out for the eight brands of sulfadoxine-pyrimethamine tablets, the following conclusions were drawn: All of the eight brands of sulfadoxine-pyrimethamine tablets tested conformed to the pharmacopoeial standards on weight uniformity. All of the eight brands of sulfadoxine-pyrimethamine tablets tested conformed to the pharmacopoeial standard on the percentage content (assay). All of the eight brands of sulfadoxine-pyrimethamine tablets tested conform to the pharmacopoeial standards of friability Four out of the eight brands of sulfadoxine-pyrimethamine tablets tested conformed to the pharmacopoeial standard of the percentage amount dissolved (dissolution test). The other four brands of sulfadoxine-pyrimethamine tablets tested were considered substandard as they failed the dissolution test. The information given on the labels on the packaging of the eight brands of sulfadoxinepyrimethamine conform to the pharmacopoeial standard of labelling as specified in the B.P.

6.2 Recommendations The results of this investigation will serve as a base line for broader investigations and also define or determine future procedures that would be used in further research projects. The following recommendations can be considered: The coverage of study could be extended (to the provinces instead of Freetown alone) so as to get a larger number of brands and samples of sulfadoxine-pyrimethamine tablets to be investigated. The use of the USP assay method of HPLC in view of its better specificity and accuracy. 6.3 Relevance of the study in Sierra Leone This finding will give a vivid picture as to the importance of Drug Quality Control in the Health Care Delivery System of Sierra Leone. Poor quality medicines are not only a health hazard, but a waste of money for both the government and individual consumers. Such drugs can damage health and destroy lives if not promptly identified and removed from the distribution channel. In this regard, relentless effort should be made by pharmacists or drug inspectors to ensure that all drugs produced or

55

imported into this country pass through the channels of inspection and quality control. It is only through Quality Control that quality drugs can be obtained, and such drugs can help in improving the health status of the country. Also, different brands of the same drug may vary in purity, efficacy and crushing strength. Therefore, the testing of such drugs for their purity, efficacy and strength is invaluable in determining which brand is the best. This will give the individual patient the opportunity of purchasing the best brand for a particular drug in curing a particular disease condition.

56

APPENDIX

57

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