Most people are not very interested in the history of a particular medicine. Quinine, however, is rather different and occupies a special place. For 300 years this was the only specific treatment for malaria. It has now been used for 360 years. It was being used long before Ehrlich wrote down the principles of chemotherapy. The story of its discovery, the important part which quinine has played in the colonisation of the tropics, its role in both World Wars and during the Vietnam war, and the present come-back of this product all make it unique. At present quinine and related products are used in the treatment of P. falciparum malaria, as an antiarrhythmic, as a muscle relaxant and as a flavouring (Schweppes!). There are also some minor applications such as the treatment of babesiosis.

The bark of a certain tree in Peru has been used in traditional medicine since time immemorial. According to the legend in 1638 in Lima, in what is now Peru, Anna del Chinchon, the second wife of Don Luiz Fernandez de Cabrera Bobadilla y Mendoza, count of Chinchon, the Spanish Viceroy of Peru, was successfully treated with this bark for a "malaria attack". Historians very much doubt the authenticity of this story. According to the notes of a certain Augustinian priest, Antonio de la Calancha (1633) the "fever bark" had come from Loxa, 700 km further north, in present-day Ecuador. The cinnamon coloured bark was finely ground and then given to the patient with hot water or warm wine. There were also reports of other acts of healing, such as that of a certain Don Juan Lopez de Canizares in 1630. The Jesuits brought the bark of this plant with them to Rome (Father Bartolomé Tafur) and Spain (Father Alonso Messias Venegas). The first load of bark was sent to Europe in 1641, to Seville, the only port which could then accept goods from the New World. In Rome the effect of the bark was tested under the supervision of Cardinal Juan de Lugo and Gabriella Fonseca, the personal physician to Pope Innocent X. In 1654 the Peruvian bark was introduced into England, but the British protestants objected to testing a "Catholic potion". Oliver Cromwell refused to use it, and is said to have died of malaria in 1658. A few years later the British doctor Robert Talbor healed King Charles II with his "secret drink". In 1678 he was sent by the same English King to France where he successfully treated the son of the French king Louis XIV (1638-1715). He revealed his formula to the French (quinine with opium in wine) in exchange for 3000 golden crowns and a pension for life. The formula was published after his death. In 1677 the tree bark was included in the London Pharmacopoeia as "Cortex Peruanus". In 1692 the Chinese emperor K’ang Hsi of the Manchu dynasty was successfully treated with it, due to which the new Westerners were looked upon favourably in his country. This had far-reaching historical consequences for western influence in China, and for Christianity in particular.

It is possible that malaria was a disease imported into the New World by the Westerners. It would therefore be strange if the Indians had already used the bark for this new illness. Nowhere is it reported that the Indians used quinine bark against fever. Although intermittent fevers (ague) were then well known, the difference between malaria and other causes of fever did not really become clear until the germ theory became generally accepted at the end of the 19^th century. No one identified "malaria" as such, no one knew what Cinchona alkaloids were, let alone quinine. It is a fact that Indians who worked in the cold highlands, drank warm quinine tea so that they shivered less. Quinine suppresses striated muscular tissue in two ways: directly by prolonging the refractory period after muscle contraction and indirectly by heightening the threshold at the neuromuscular junction. This is illustrated by the fact that small amounts of quinine increase the severity of myasthenia gravis symptoms. Quinine can also be used in the treatment of myotonia congenita and leg cramps at night. It was probably due to the observation that quinine reduces shivers, and that malaria is often accompanied by fever, that this was administered. It may have been pure chance that precisely this agent also killed the parasite and brought healing.

In the first half of the 18^th century (1735-1738) France organised a scientific expedition in order to determine the form of the earth by measuring the length of an arc of a meridian and thus establishing the flattening of our globe towards the poles. On this expedition, which was led by Charles-Marie de La Condamine, there was a botanist, Joseph de Jussieu. In 1735 he described the tree as a member of the Rubiaceae, the same plant family as coffee. Initially he gave it the name Quinquina condamine. This caused confusion since another tree, the Quina-quina (Myroxylon balsamum), the source of peru balsam, could also be used for medical purposes. He brought specimens back to Europe. In memory of the noble lady mentioned above, the quinine tree was named Cinchona after her by Linnaeus (the misspelling was due to Linnaeus in 1739 and was included in his book "Genera Plantarum"). In 1820 the Frenchmen Pierre Pelletier and Joseph Caventou isolated pure quinine from the bark (etymological origin of "quinine": from "Chinchon" or from the Inca "Kinia" = bark?). They received 10,000 francs for this from the Paris Institute of Sciences. Quinine could not yet be synthesised, however, and so its supply remained dependent on continuous importation of the bark from South America. A large proportion of the harvest was transported to the port of Paita to be shipped north from there to Panama. It was then taken overland to Portobello, to again be shipped westwards to Havana and then to Spain. Part of the load regularly rotted and became worthless. Some of the bark was smuggled. Often the cargo was mixed with useless bark from other trees. It was thought that the bitter taste of the quinine bark was important to its healing power. All kinds of bitter tree barks were therefore sold as authentic quinine bark. Massive harvesting of the wild trees endangered further production. In 1795 the explorer Willem Alexander von Humboldt (1769-1859) recorded that in that year alone near Loxa 25,000 wild trees perished. Exhaustion of the natural supplies was partially counteracted when it was discovered that after pollarding new branches formed, which after 6 years had bark containing a higher concentration of the active product. An attempt was made to set up plantations. The export value of Cinchona bark was very high. Bolivia, Peru, Ecuador and Colombia tried to benefit from their monopoly positions. However, this threatened the British and Dutch overseas interests. In 1852 Justus Hasskarl, director of a Dutch botanical garden on Java, organised a secret mission to obtain seeds from the trees in South America. The next year he returned with a bag of seeds, for which he was knighted. The quinine content in the bark proved to be disappointingly low, however. Apparently the different varieties of Cinchona trees also differed in the amount of alkaloids which they produced. It proved a difficult genus for botanists and taxonomists. There were more than 40 species and each had several varieties. With the help of a Bolivian Aymara Indian, Manuel Incra Mamani, in 1865 the Australian Charles Ledger smuggled seeds from the quinine tree (Cinchona ledgeriana) out of Bolivia. After the British government refused to buy the seeds (they did not trust Ledger), he sold them to the Netherlands for 20 dollars a pound. This was the best investment ever. A huge series of plantations was set up with these seeds on Java. Some plantations were started in the Nilgiri hills in India. After selective breeding of the plant it was possible to produce large quantities of quinine. The amount of quinine in the bark rose from 0.3% to 8%, sometimes in exceptional cases to 13% and even 18%. In 1930 the Dutch plantations produced 22 million pounds of bark on Java, which covered 97% of the world-wide need for quinine. Only four of approximately 40 species of the Cinchona, are of direct practical importance in the production of quinine. Cinchona ledgeriana, C. officinalis, C. calisaya, C. succirubra and to a lesser extent C. pubescens, are the chief sources. Traditionally the barks are classified according to colour: yellow Cinchona is obtained from C. ledgeriana and C. calisaya, red Cinchona from C. succirubra and brown Cinchona from C. officinalis.

The plants grow between 10°N and 22°S, with the exception of plantations in India, where they are cultivated up to 17°N. Cinchona is generally cultivated at heights above sea level of between 1000 and 2000 metres. High humidity and a continuous supply of water throughout the entire year are important. As with many plants the trees are short and squat at the outer limits of their growth zone. Below 1000 metres they quickly become diseased. In the wild they can grow to 20 metres high and live for 80 years under optimum conditions. In plantations they rarely reach more than 6 metres. Approximately 1500 trees are planted per hectare. They are propagated via seed or using vegetative methods (grafting). The plant is a cross pollinator and there is self sterility. Maintaining genetic purity is therefore a problem. Trees propagated from seed from a tree with a high yield produce a heterogeneous population with a generally low yield. In Indonesia C. ledgeriana is usually grafted onto C. succirubra. This became the basis of the commercial procedure for obtaining clonal material. Infections can have catastrophic results in monocultures (cf. potato blight and the famine in Ireland). Phytophthora palmivora, Pythium vexans and Pellicularia salmonicolor are important pathogens for quinine trees. Recently Phytophthora cinnamoni has become a major problem in Rwanda and Kenya. The control of such pathogens in plantations is therefore strict. Recognition of this genetic weakness has meant that the question of maintaining the largest possible natural genetic reservoir is taken seriously by protecting the trees living in the wild.

Quinine is obtained from the bark of the tree. The highest concentrations are found in the outer layers, the epidermis and periderm of the branches and trunk. The root bark contains lower concentrations. If careful the outermost bark layer can be scraped off without touching the cambium. The trees will then renew their bark. It was noticed early on that Cinchona-alkaloids develop a red colour after they have been in direct sunlight for a time. It was also observed that the bark on the shady side was richer in alkaloids than that on the sunny side. This led to the technique of "mossing". The bark was protected by local application of moss to the trunk. This produced a considerably higher yield of alkaloids. By stripping the bark longitudinally followed by mossing, trees could be harvested continuously. However, this requires a great amount of manual work and is not economic. Nowadays the trees are felled. As an alternative, after heavy pruning two or three new shoots can be left to grow and these can be harvested seven years later, or pruned again. The bark is removed and dried in the shade. After being ground, the powder is mixed with lime and water and then follows extraction, e.g. with hot toluene. The alkaloids can then be isolated with diluted sulphuric acid. The quinine is purified by means of crystallisation. Nowadays in-vitro callus and suspension cultures can be produced from the plants, but commercial in-vitro culture is not economically attractive.

As well as quinine the bark also contains variable amounts of related alkaloids such as cinchonine, another product with anti-malaria activity. Indeed, 36 different alkaloids have been isolated, but most are present only in trace amounts. Due to the presence of asymmetric carbon atoms (in particular carbons 8 and 9) in the molecule, various enantiomers can be distinguished: quinine (8S, 9R), quinidine (8R, 9S), epiquinine (8S, 9S) and epiquinidine (8R, 9R). Below are the percentages of the alkaloids obtained via the present standard extraction and via boiling Peruvian Cinchona calisaya bark in either water or 70% alcohol. It has been found that as a percentage of the total number of alkaloids, quinine is more or less constant at approximately 85%:

Much was changed by the second World War. In 1940 the German army seized the entire European reserve supplies of quinine when they took Amsterdam. For the West, obtaining Cinchona bark from the plantations in Southeast Asia became impossible when in 1942 the Japanese occupied Indonesia. There was a small Cinchona plantation in the Philippines, but a few weeks later this also fell into Japanese hands. The last allied aeroplane to leave the Philippines, however, had 4 million Cinchona seeds on board. These were sent to Costa Rica to be planted. Plantations were also set up later in other countries: Kenya, Rwanda, Congo, and so on. However, it would take several years before there could be any harvest. The American government sent the botanist Raymond Fosberg from the Smithsonian Institute to South America to secure a new supply of bark, originating from trees living in the wild. He succeeded in part, but he was never able to find Cinchona ledgeriana. In the meantime alternatives had to be found, for malaria took a huge toll during the conflict in the Pacific. The stereochemistry proved very important to the activity of the product. In 1908 Rabe was able to determine the chemical structure of quinine. It had 4 chiral centres. Synthetic production of quinine proved to be very difficult, but was eventually achieved in 1944 by Woodward and Doering of Harvard University. However, large-scale production was not economically viable. This led to investigation of alternatives. One alternative to quinine was mepacrine but this product had various unfortunate side effects and the first cases of resistance had already been observed in New Guinea.

The synthetic preparation of primaquine was perfected after the war. The British war programme led to the development of proguanil, which itself served as a model for the development of pyrimethamine. Pyrimethamine in combination with sulphadoxine was introduced in 1970 under the name Fansidar. After World War II it was hoped that malaria would be definitively eradicated. The use of chloroquine and the world-wide campaign to eradicate malaria (WHO [World Health Organisation]), led initially to a considerable reduction in malaria infections all over the world. After the anti-malaria campaign diminished due to various circumstances, the resistance of Anopheles to various insecticides and the development of chloroquine-resistant and multiresistant P. falciparum, malaria once more became one of the major problems.

Due to the great initial success of chloroquine, in the late ’50s there was no longer so much need for quinine as an anti-malaria agent. The Cinchona plantations would have gone bankrupt and cultivation would have stopped, had it not been that quinidine, the stereo-isomer of quinine, was discovered in cardiology as an anti-arrhythmic agent. It proved difficult to synthesise quinidine chemically. Quinidine occurs in the tree bark, but in small amounts. It was possible to convert quinine chemically to quinidine (via the intermediary quininone and its epimer quinidinone). For this reason the plantations were kept going. By the time quinidine fell out of use due to the development of other anti-arrhythmics, quinine was once more in demand for malaria treatment.

*

Whereas World War II led to the discovery of some new anti-malaria agents, the Vietnam war stimulated a huge programme for the discovery of new drugs. The Walter Reed Army Institute of Research of the United States army investigated thousands of constituents. This research resulted in mefloquine (Lariam®) and halofantrine (Halfan®). Research in China produced artemisinin, pyronaridine and benflumetol. In 1976 Trager and Jensen at the Rockefeller University in New York, developed a method of culturing P. falciparum in vitro. This brought research into the molecular biology of the parasite, the resistance patterns and new drugs into the fast lane. In 1997 Claudia Golenda, Jun Li and Ronald Rosenberg of the Walter Reed Army Institute of Research (Washington) and the NIH [National Institutes of Health, Bethesda] described a continuous in vitro culture method for P. vivax. This technique used a high concentration of Duffy-positive reticulocytes in the culture medium. The expected increase in chloroquine-resistant P. vivax could therefore now be studied in the laboratory. No single treatment regimen nowadays gives a 100 % guarantee of cure. More and more combination chemotherapy will come into use in the near future (quinine + doxycycline, quinine + clindamycin, quinine + Fansidar, atovaquone + proguanil, mefloquine + artesunate, artemether + lumefantrine, and so on).

• 17^th century: quinine (actually a mixture of various Cinchona-alkaloids administered in warm wine)
• 1925: Plasmoquine = Pamaquine, Praequine
• 1932: Atebrine = Mepacrine, Quinacrine
• 1934: Chloroquine (Germany, Sontochin)
• 1944: Proguanil = Paludrine®
• 1950: Pyrimethamine = Daraprim®
• 1952: Primaquine
• 1955: Amodiaquine = Flavoquine®, Camoquine®
• 1969: Artemisinin (only in China)
• 1971: Sulphadoxine/pyrimethamine = Fansidar®
• 1986: Mefloquine = Lariam®
• 1987: Artemether and Artesunate (also outside China)
• 1988: Halofantrine = Halfan®
• 1992: Atovaquone (experimental)
• 1997: Etaquine – Tafenoquine (experimental)
• 1999: Co-artemether (artemether + lumefantrine)
• 1999: Atovaquone / proguanil = Malarone® commercialised

Broadly speaking, anti-malaria drugs can be divided into four major classes

• Blood schizonticides
• Antifolates
• Antimitochondrials
• Redox process-based agents

When the malaria parasite leaves the liver and penetrates an erythrocyte, it can at last begin a haemoglobin diet. However, it cannot use the iron-containing haem group. What is more, released ferriprotoporfyrin IX (syn. = haemin) is toxic for the parasite. It contains trivalent iron (ferric = Fe³⁺). Normally the parasite polymerises haemin to non-toxic malaria pigment. Chloroquine, quinine, mefloquine and halofantrine interfere with the detoxification of haemin in the digestive vacuole of the parasite. The drugs prevent this detoxification so that haemin can generate free radicals and membrane damage follows. It is therefore logical that the drugs are not active against the parasitic stages which precede the blood forms (sporozoites, liver forms) and which do not consume haemoglobin.

Folic acid is an important metabolic factor. Humans obtain this vitamin from the food they eat. The malaria parasite, on the other hand, must produce it for itself. Para-aminobenzoic acid (PABA) is used at an early stage of the biosynthesis of folic acid by the enzyme dihydropteroate synthetase. This step is inhibited by structural analogues of PABA, such as sulphonamides and sulphones, e.g. sulphanilamide, sulphadoxine and dapsone. The next synthesis step is catalysed by dihydrofolate reductase. This step is prevented by pyrimethamine, trimethoprim and cycloguanil (prodrug = proguanil), to such an extent that tetrahydrofolate – the end product – is not formed. The combination of these two sequential inhibitors forms the basis of Fansidar® (similar to cotrimoxazole). Resistance to both antifolates is easily developed, however (a specific point mutation in each gene is sufficient).

Although artemisinin derivatives and 8-aminoquinolines cause mitochondrial swelling, this organelle is not their chief target. Some antibiotics such as tetracycline and clindamycin prevent protein synthesis by mitochondrial ribosomes (these are similar to the ribosomes found in bacteria). They are slow-acting. Atovaquone is a naphthoquinone which specifically destroys the electron transport chains of Apicomplexa. The molecule is rather similar to ubiquinone (coenzyme Q) which plays a role in the energy transfer between cytochrome B and C1. The enzymes of Plasmodium falciparum are 1000 times more sensitive to atovaquone than the corresponding enzymes in humans. Resistance can easily develop if it is used in monotherapy.

Primaquine and etaquine exercise their action via redox-active quinone metabolites. They are selectively toxic for the pre-erythrocytic stages and are the only medicaments which kill hypnozoites. Etaquine has in addition a pronounced blood schizonticidal action.

This is a powerful product, which acts upon the schizonts of the parasites in the blood (it is a schizonticide). It thus acts chiefly in the second half of the maturation cycle: on the parasites which are sequestered in the small blood vessels (not on the young ring forms in the peripheral circulation). Quinine also possesses gametocytocidal activity against P. vivax, P. malariae and P. ovale (but not against gametocytes of P. falciparum). Although it was earlier claimed that quinine had a weak antipyretic action (used for example in Latepyrine-quinine®), this is questioned nowadays. Besides treatment of P. falciparum malaria, quinine is also used to reduce muscle cramps. Quinine is available in several forms. It is a substance consisting of 2 parts: a base (the active part) and another chemical group (important for solubility). The 2 components together are called the salt. Some confusion may arise as regards the dosage of quinine, depending on whether this is expressed in mg of the base or in mg of the salt (generally the salt is used).

Quinine base is 82 % of the total substance if the salt is the dihydrochloride or sulphate, but is only 59 % if the salt is the bisulphate. Quinimax® is a product frequently used in Africa. Some 100 mg tablets contain only 59 mg of quinine base, however, and a very small amount of other substances. This may lead to under-dosage.

Quinine sulphate is administered orally. It is absorbed well in the intestines. An obsolete product is Arsiquinoforme®. This combination drug contains 62% quinine in the form of quinine formiate and quinine acetarsolate. Each 225 mg tablet contains 18 mg of pentavalent arsenic. Quinine bihydrochloride is injected, preferably by slow IV (infusion with glucose because of the risk of hypoglycaemia). IM injections may lead to sterile abscesses, but can be used where necessary if there are no alternatives available. For IM injection, it is best to use a diluted solution (60 to 100 mg/ml) in place of the concentrated solution (300 mg/ml). Quinine administered via IM injection is absorbed well even in severe malaria. Treatment with quinine is unpleasant (bitter taste, cinchonism) and poor compliance after the acute phase is common.

The basic regimen is 10 mg salt/kg, every 8 hours, orally or slow IV. This should be continued for at least 4 days, preferably 7 to 10 days. This is an unpleasant treatment. Because there is still a risk of relapse if quinine is used in monotherapy, another product is generally combined with it, e.g. tetracycline or vibramycin. Sometimes treatment with Fansidar® is given after a few days, which shortens the treatment period. In the case of cerebral malaria it is best to give a loading dose (first administration = double dose or quinine bichlorhydride 7 mg salt/kg IV over 30 minutes; or once 20 mg/kg IV over 4 to 8 hours). P.S. Please note that the bichlorhydride is given, and not the monochlorhydride. The latter is less water-soluble. Sometimes there is a brief increase in the parasitaemia after beginning therapy. This does not directly mean that the therapy has failed.

If a patient vomits within an hour after swallowing the medication, the whole dose should be repeated. If vomiting occurs longer than one hour after ingestion, no new dose is necessary. In hepatic or renal insufficiency the normal dose of quinine is administered for the first 48 hours. The dose is then reduced to half or one third (administration of quinine is then every 24 hours). There is controversy over this, however, (probably much less of a reduction is necessary). ECG monitoring is recommended during quinine therapy in patients with existing kidney failure (watch in particular for QTc-prolongation and arrhythmia). If haemodialysis is required, quinine should be administered after (rather than before) although little or no quinine is removed by this procedure. Dose adjustment during haemodialysis is not necessary. Classic haemodialysis cannot be used therapeutically in quinine intoxication.

Overdosage of quinine may lead to very severe situations such as deafness, delirium, bradycardia, hypotension, respiratory arrest or death (lethal dose approximately 8 gram). Overdosage may also lead to blindness via a direct toxic effect on the retina and possibly also due to spasms of the retinal blood vessels and subsequent retinal ischaemia. The half-life of quinine in the blood is short (12 hours). Most of the quinine in blood is bound to proteins. The bound fraction increases from 80% to 90% in acute malaria (increase in acute phase proteins). Overdosage is thus less dangerous in active malaria. The total quinine blood level is of much less importance than the blood level of free quinine, which is much more difficult to measure. 95% of the quinine is converted in the liver and then eliminated via the kidneys.

Unlike the majority of other bitter products which occur naturally, the bitter taste of quinine is short-acting with no annoying after-taste. It is therefore used as a flavouring to produce tonic water. The British colonialists in India often drank gin and tonic. The present-day tonic water contains approximately 15 mg per litre, however, only enough to give a bitter taste. Copious drinking of gin and tonic in order to prevent malaria, is thus only an excuse for drinking gin.

Why is there almost no quinine resistance? The product has been used for 360 years. This is in stark contrast to the resistance to other malaria drugs or antibiotic resistance in bacteria, where the "useful life" of a product is measured in years or a few decades. The concept of a standard dose was only developed in the twentieth century. Earlier the duration of treatment and the dosage were left to the discretion of the doctor. This, together with the fact that the concentrations of alkaloids varied greatly from plant to plant and that quinine was never pure, meant that malaria was treated with a therapy which must have produced the most varied blood levels. Yet no wide spread quinine resistance has been reported. The answer to the question why there is virtually no quinine resistance, could be very important. Is the target molecule of quinine so special that mutation is not possible? It would then be very helpful to know this target. It could also be that there is quinine resistance, but that it was not, and has not been recognised. However, this is doubtful. Is it that the present recommended dose is much higher than that which was formerly necessary? Is it the fact that "quinine" is actually a mixture of various active products, which prevents resistance developing? Resistance to combined therapy requires multiple, simultaneous mutations which is less readily achieved than that to single products. It is, however, possible that quinine has not previously been used at levels which create sufficient evolutionary pressure. The majority of malaria cases in Europe and America were P. vivax infections. Even in British India, P. vivax represented the lion’s share of infections. In P. falciparum endemic regions, only a few fortunate people were able to take quinine and then only when they had to (because of unpleasant side-effects). Few used quinine as a prophylactic agent (compared to the indigenous population). What is more, quinine has a short half-life, so that the parasite was only exposed to subtherapeutic concentrations for a short time. Probably its limited use is the reason for the absence of resistance, and with continuous use on a large scale, quinine resistance may yet become a reality in years to come.

The trophozoite in the red blood cell breaks down haemoglobin using lysosomal enzymes. In this digestive process ferriprotoporphyrin IX (haemin) is released from the haemoglobin protein chains. This porphyrin is toxic to the parasite and is usually polymerised to non-toxic malaria pigment. The weak base chloroquine accumulates in the acid lysosome and binds to ferriprotoporphyrin IX. In this way detoxicification of the latter is prevented and the parasite is killed. Since the liver parasite does not feed on haemoglobin and the effect of chloroquine is to prevent detoxification of the haem ring, this drug is not active at the pre-erythrocytic stages of Plasmodium sp. Outside the cell, chloroquine as well as hydroxychloroquine are present mainly in a protonated form, that, due to their positive charge, cannot cross the plasma membrane. However, the non-protonated form can enter the cell. Inside the cell, the molecule gains a proton (H⁺) in a manner inverserly proportional to the pH, i.e. the lower the pH, the more chloroquine will bind an extra proton. Chloroquine will be concentrated in acidic cell organelles, such as Golgi vesicles and lysosomes, where due to the low pH, most chloroquine molecules will be positively charged. Since chloroquine is a weak base, the pH will rise. By increasing this pH, several enzymes such as acid hydrolases can be inhibited. Post-translational modification of newly synthesised proteins can be disturbed.

Chloroquine is available in tablet form as chloroquine sulphate (Nivaquine®) and as chloroquine diphosphate (Resochine®). Other brand names are Daramal®, Anochlor®, Promal®, Avlochlor®. Hydroxychloroquine sulphate (Plaquenil®) is different and is used in rheumatoid arthritis. It is used in psoriasis patients (at the same dosage). The injectable form is chloroquine dihydrochloride. There is a combination tablet of chloroquine 100mg + proguanil 200 mg (Savarine®), which makes compliance easier for prophylactic use (1 tablet daily). The dose is always expressed as base (not as salt). This allows easier comparison between the different products. Nowadays most Nivaquine® tablets contain 100 mg. It is also highly advisable to check the current dose per tablet in the region where you are working, so as not to cause accidental overdosage or underdosage.

• Chloroquine diphosphate 250 mg tablet = 150 mg chloroquine base
• Chloroquine sulphate 200 mg tablet = 150 mg chloroquine base
• Chloroquine dihydrochloride 50 mg/ml = 40 mg base/ml

Chloroquine is a powerful schizonticide. It has strong affinity for various tissues and organs. It is fast-acting and remains in the blood for many days. A brief treatment is therefore possible. The excretion of chloroquine and its metabolites occurs mainly via the kidneys and is slightly improved by acidification of the urine (500 mg vitamin C every 4 hours). Chloroquine may be given orally, SC, IM or SLOWLY IV (infusion). Never inject an ampoule of chloroquine IV rapidly as a bolus. It is essential that rapid infusion is avoided. Chloroquine bihydrochloride IM is well absorbed (>80% even in severe malaria). The injections are not painful.

There are several different treatment regimens. Orally 25 mg/kg is given spread over three days. The following is practical for an adult weighing 60 kg: first, 6 tablets of 100 mg base, followed after 6 hours by another 3 tablets, after 24 hours by another 3 tablets and after 48 hours by another 3 tablets: 6-3-3-3. If parenteral therapy is required due to coma caused by chloroquine-sensitive P. falciparum, chloroquine bihydrochloride may be given IM or IV. The total dose should be 25 mg base/kg. There are several regimens, e.g. 10 mg base/kg over 8 hours, followed by 15 mg base/kg over 24 hours. Another frequently used regimen is 3.5 mg base/kg SC or IM every 6 hours. Parenteral administration should be discontinued as soon as oral administration is possible. In moderate renal impairment (creatinine clearance > 10 ml/min) no adjustment of the dose is necessary.

Chloroquine is cheap and not very toxic in normal use. Some people are allergic (pruritus, rash) or suffer nausea. People with psoriasis are more at risk of side effects. A reversible precipitation of chloroquine in the cornea may occur, resulting in small opacities. This may result in seeing haloes around objects, blurred vision or photophobia. This form of keratopathy may become manifest quite rapidly (a few weeks after beginning treatment). After discontinuing the medication it is completely reversible. Chloroquine accumulates in melanin-containing tissues. Chronic use may lead to abnormalities of the choroid and retina (chorioretinitis). This toxic retinopathy is not reversible. The abnormalities are always bilateral and symmetrical. Often there is maculopathy (bull’s eye) with central and paracentral scotomata, but constriction of the peripheral field of vision may also occur. The total cumulative dose before such problems occur is generally 100 gram chloroquine or more. Hydroxychloroquine has rather lower retinal toxicity. Tinnitus, hearing loss and neural deafness have been reported as possible rare side effects. Sometimes a proximal "chloroquine myopathy" occurs. The muscular weakness in myasthenia gravis is exacerbated by chloroquine and this disorder is a formal contra-indication for the use of this product. Breast feeding may be continued without change while taking chloroquine.

Chloroquine has a narrow safety margin (just 30 mg/kg may be fatal). In case of overdosage myocardial depression, hypotension, severe arrhythmias and tissue hypoxia may occur. ST-segment abnormalities and T-wave inversion occur. Broadening of the QRS complex (>0.12") and ventricular arrhythmias have a poor prognosis. The patient may become comatose, vomit and aspire the stomach contents. In acute intoxication diazepam is given (Valium® 1 mg/kg) and adrenalin (= epinephrine) or dopamine if these are available. First protect the airways and correct any existing cardiovascular disorders and then consider gastric lavage if the overdose was recent. Introducing activated carbon into the stomach is also beneficial. Acidification of the urine, osmotic diuresis, haemoperfusion and haemodialysis are of little use. After oral ingestion of an overdose the effect is evident approximately 1 hour later. Death follows after a further 2 to 3 hours.

The individuals within a population of parasites are not identical. Drugs do not have an equal effect on each variant. The larger the population and the shorter the life cycle of an organism, the faster mutations will become manifest. Of course Anopheles and Plasmodium have changed more in recent decades than have humans (evolutionary pressure due to insecticides and drugs). The first signs of chloroquine-resistant P. falciparum infections occurred in the ’60s, more or less simultaneously in Colombia and Thailand. This resistance spread progressively and is now a significant problem in many countries (see map with zones A, B and C). There are three grades of cloroquine resistance (RI, RII, RIII). In RI the parasitaemia after therapy is so low that it falls below the detection threshold, to rise above it again within 28 days. In RII the parasitaemia is reduced by at least 75 %, but the parasites remain detectabable in the peripheral blood. In RIII chloroquine has no effect on the parasitaemia. In spite of the presence of this resistance, chloroquine still has a place in treatment. It is still active against other plasmodium species. Only some falciparum parasites are resistant and only some have type RIII resistance. In non-urgent situations, therefore, this product may still be used. Recently chloroquine-resistant P. vivax strains have been discovered (Papua New Guinea, Indonesia, India, Brazil, Guyana). To date P. ovale is still 100% susceptible to nivaquine. The first chloroquine-resistant P. malariae has been reported (Malaysia, 2002).

As well as for malaria, chloroquine is also used for a number of other disorders: rheumatoid arthritis, polymorphous light eruption, discoid lupus erythematosus, cutaneous sarcoidosis, hepatic amoebiasis, Q fever, cutaneous porphyria tarda etc. Chloroquine also exerts direct antiviral effects by inhibiting pH-dependent steps of the replication of several coronaviruses, Borna virus, Mayaro virus and retroviruses (there is even a very modest anti-HIV activity). Chloroquine inhibits the uncoating of hepatitis A virus. Certain flaviviruses will be inhibited by affecting the normal proteolytic processing of the flavivirus prM protein (precursor of membrane protein). Chloroquine has immunomodulatory properties, such as suppressing the production and release of TNF-alpha and interleukin-6.

The porphyrias are a group of disorders characterised by disturbed production of haem. Most are autosomal dominant and inherited, and there is quite variable expression of the disease. Porphyria cutanea tarda can also be acquired, however, (oestrogens and excessive alcohol play an important role). The disorder generally becomes evident around the age of 40 to 50 and is characterised by hepatic abnormalities and significant skin fragility (chiefly the hands, legs and face. Blisters, erosions and scabs appear on skin exposed to sunlight. A very dark skin (hyperpigmentation) with sclerodermoid changes may occur. It is important for the patient to stop drinking alcohol, and to avoid oestrogen and iron preparations. Hepatic siderosis is generally present and repeated phlebotomies are indicated (400-500 ml every 1 or 2 weeks, until Hb < 11 g%). Generally 4-10 litres of blood are removed before a therapeutic effect is achieved. Chloroquine is used in the treatment of porphyria cutanea tarda. A low dose is used (125-250 mg twice a week). N.B.: daily administration of chloroquine causes acute liver toxicity and massive porphyrinuria. Chloroquine complexes with uroporphyrin and increases excretion from the liver. A clinical effect can be expected within 4-6 months (biochemical remission is slower). The risk of hepatoma is probably reduced by the treatment.

This is similar to chloroquine. Amodiaquine is converted to an active metabolite (desethyl-amodiaquine). 200 mg of amodiaquine base is equivalent to 260 mg amodiaquine hydrochloride. The product has no taste (unlike chloroquine which is bitter). Long-term use causes grey skin pigmentation in white people. Sometimes there are severe side effects (agranulocytosis in approximately 1/2000, liver toxicity in approximately 1/15,000). Amodiaquine (Camoquine®, Flavoquine®, Malarid®) are therefore still only used rarely. There is less resistance to amodiaquine than to chloroquine. Since the product is eliminated slowly, a single dose of 600 mg was (and is) sufficient. Amopyroquine (Propoquine®) is an amodiaquine analogue, but is little used and then only IM. Its safety during pregnancy is uncertain.

• Maloprim® or Deltaprim® The combination of pyrimethamine (12.5 mg) with dapsone (100 mg) was and still is used as prophylaxis in some regions, including Zimbabwe and South Africa. It is only slightly effective and the indications diminish from year to year. Dapsone is removed from the body quite rapidly (t½ = 28 hours). It cannot be used as a curative drug.

• Fansidar® is a combination product of pyrimethamine 25 mg and sulphadoxine 500 mg per tablet (Mekalfin® is another commercial name). Sulphadoxine is a long-acting sulphonamide (t½ = 8 days) which in case of allergy may cause severe skin lesions (erythema multiforme and Stevens-Johnson syndrome). For this reason it is best not used for prevention. Plasmodium falciparum has developed resistance to this product in some parts of the world. The curative dose for an adult of 70 kg is 3 tablets taken as one dose. The action is slow. The idea behind the combination of pyrimethamine with a sulphonamide is the sequential inhibition of tetrahydrofolic acid formation which is necessary for purine and pyrimidine (DNA precursors) synthesis. Theoretically there is a risk of icterus (and kernicterus) if pregnant women use it, but this risk is very limited.

This is a long-acting product (after 2 to 3 weeks half of the dose is still present). There is also a combination of mefloquine + pyrimethamine + sulphadoxine (Fansimef®). Mefloquine has a rather slow onset of action.

The curative dose of mefloquine for an adult is 500 mg orally, to be repeated after 12 hours and after 24 hours (sometimes 750-500-250 mg is also given). In Europe the dose is expressed as the base (250 mg base = 274 mg mefloquine hydrochloride) while in the USA it is expressed as the salt (250 mg salt = 228 mg base). Therefore, a 500 mg dose in the USA gives approximately 10% less active drug than in Europe. There is no injectable form. The action of mefloquine upon the myocardium is the same as that of quinine. There must be an interval of 12 hours between the last dose of quinine and taking mefloquine (quinine is removed rapidly from the body). During treatment of an attack mefloquine may be given if there is renal insufficiency (a common problem in severe malaria). At curative doses there are quite often unpleasant side effects, such as nausea, insomnia, tremor, anxiety and more rarely convulsions or psychotic episodes.

Mefloquine plays an important role in prophylaxis. The product is 98% bound to plasma proteins. Breakdown to an inactive metabolite occurs in the liver. Excretion is mainly via the liver and biliary route, and to a very limited extent via the kidneys. The plasma half-life is 2 to 3 weeks. Ingestion of 1 tablet per week produces stable blood levels after 7 weeks. This may be drastically shortened by taking a loading dose over 3 days (1 tablet per day x 3 consecutive days). Traditionally it is said that mefloquine prophylaxis should be started before departure. This guideline is based on the consideration that intolerance to the drug can be monitored in this way. It is safe to begin the medication 15 days before departure so that 3 tablets are taken before leaving. In this way 75% of the side effects can be detected. At the prophylactic dosage (adults one 250 mg tablet per week) side effects occur in 2 to 3% of people, which require that the prophylaxis be discontinued. Rarely (1 in 12,000 to 15,000) preventive dosages may trigger epilepsy or psychosis may occur. Epilepsy and arrhythmias (including the use of beta-blockers, calcium antagonists and digitalis) are contra-indications for the use of this product. The product was initially not recommended in pregnant women, but in practice has proved to be safe in the second and third trimesters. The prophylactic use of mefloquine used to be limited to 3 months, but new data indicate that the product is safe if taken for longer. It is best if people who carry out critical motor operations (e.g. aeroplane pilots) do not take mefloquine. In 1994 an anecdotal case was described, of a central anticholinergic syndrome (delirium and stupor, mydriasis, hyperpyrexia) after taking mefloquine. This was very swiftly reversible after injecting 2 mg physostigmine (a cholinesterase inhibitor similar to prostigmine).

The first case of mefloquine resistance was described in Thailand in 1982. There is already mefloquine resistance on a small scale in many countries, but this can be significant locally: e.g. the cure rate in East Thailand was only 41% in 1993. P. falciparum malaria can thus sometimes occur in spite of correct prophylactic use of mefloquine. Mefloquine does not kill sporozoites (therefore P. vivax and P. ovale malaria are still possible after leaving an endemic zone and after discontinuing mefloquine).

In Southeast Asia resistance is common, but in other regions this problem is still limited. There is limited cross resistance with mefloquine. Cases of insufficient clinical response (5% of cases) are usually due to inadequate absorption. Sometimes Halfan® is used as the drug of first choice, or as a stand-by treatment for travellers, but this has been superseded by Malarone. It should be stressed that halofantrine cannot be given IV. Use in severe malaria might therefore be problematic (possibly via gastric tube). During treatment with halofantrine there is prolongation of the PR-interval and the QT time. Severe cardiac problems have been described in people who previously had a cardiac conduction disorder. Prolonged QT_c time, thiamine deficiency, conduction disturbances, ion deficiency (hypokalemia, hypomagnesaemia) and concomitant administration of quinine or mefloquine are contra-indications. If there is a falciparum malaria during mefloquine prophylaxis, Halfan® is not a good choice due to possible cross-resistance and cumulative cardiotoxicity. Other products which may also cause QT-time prolongation, such as cisapride [Prepulsid®], terfenadine [Triludan®], tricyclic antidepressants such as clomipramine [Anafranil®] and amitriptyline [Redomex®] are best avoided during Halfan® administration. An ECG is therefore advisable before administration of Halfan®. Halfan® cannot be used as prophylaxis. [A reminder: the QT-time is the time from the beginning of the Q-wave to the end of the T-wave. The correction for cardiac rate using Bazett’s formula (QT/Ö RR) gives the QTc.] If there is a prolonged QT-time (longer than 440 msec) there is an increased risk of ventricular tachycardia, more specifically "torsade de pointes". This is characterised by polymorphous QRS complexes which vary in amplitude and appear to oscillate around the isoelectric line.

Qinghaosu ("essence of qinghao") originates from a Chinese plant, Artemisia annua (sweet wormwood). This is a plant with composite flowers related to absinthe and tarragon. As the name suggests it is an annual plant, from which seeds need to be collected each year for the next cultivation. The leaves and flower heads contain the highest concentration of drug and are harvested. The variant which grows in South China and North Vietnam contains a high concentration of active constituents (up to 1.5% dry weight), unlike the variants which grow outside this zone (Europe, Virginia). From it are produced artemisinin and the derivatives artesunate (the hemisuccinate), arteether (the ethyl ether) and artemether (the methyl ether). (A reminder: an ether has the formula R-O-R’). Artelininic acid is a more water-soluble derivative (artelinate). The products are rapidly converted after ingestion to the equally active dihydro-artemisinin. Inhibitors of the cytochrome P450 (CYP 3A 4/5) such as grapefruit juice, increase the plasma levels of artemether to a significant extent (e.g. double). After oral administration there is a significant first-pass effect in the liver. The plasma half-life of artemether and its active metabolite is very short: both 1 hour.

The substance is not active upon liver stages, but upon both the immature sexual and the early-stage asexual blood forms. It does not kill mature gametocytes of P. falciparum. Since it reduces the number of gametocyte carriers, it helps to prevent malaria transmission. These drugs are used chiefly in Asia (including China, Thailand and Vietnam). They are also available at present in many African countries. One important advantage of artemisinin derivatives is the very rapid action (faster than quinine). Artemisinin and artesunate may be administered as suppositories if the patient is too ill to take oral medication (e.g. Plasmotrim Rectocaps®). The bio-availability of the drug via this route is approximately equal to that of the oral form. Artemisinin induces an enzyme which strongly promotes its own breakdown. In monotherapy relapse is frequently seen (depending on the dose and duration of therapy). Should these products be used wrongly, swift appearance of artemisinin-resistant falciparum strains can be expected. Neurotoxicity has been observed in animal trials (selective damage to certain brain stem nuclei and to the auditory nuclei in rats, dogs and Rhesus monkeys treated with IM artemether and arteether). Apparently no ototoxicity or neurotoxicity occurs in humans. The drugs also suppress reticulocytosis, but the clinical importance is still unclear. During treatment with artemether a transitory bradycardia and prolonged QT-interval was observed in approximately 1% of cases, but with no direct clinical repercussions. Artemether and dihydroartemisinin reduce the number of parasites by approximately 10,000 for each asexual cycle. After two cycles (3 days’ treatment) there is a 10⁸-fold reduction of the parasitaemia. Five days’ treatment can result in a 10¹²-fold reduction in the parasitic biomass. In functional asplenia artemether is less effective. People who are heterozygous for haemoglobin E have a swifter parasite clearance with artemisinin derivatives than those without this haemoglobin variant.

Artemisinin is poorly soluble in water and oil, but can be given as suppositories.

The derivative artemether (Paluther®, Arteminth®, Cotexcin®, Artenam®) is oil-soluble and can be used for IM administration (e.g. 600 mg/day x 5 days; recommended dose 3.2 mg/kg IV on day 1 followed by 1.6 mg/kg per day x 5 days). It should be protected from light during storage. The ampoule must be clear (there is sometimes a cloudy precipitate). There is also an oral form. Absorption improves considerably with concurrent ingestion of grapefruit juice (possible role of intestinal CYP3A4). The fixed combination of artemether with mefloquine produced good results in Southeast Asia.

Artesunate (Artenam®, Artesunate®, Arsumax®, Artemax®, Arinate®, Plasmotrim®) is the fastest-acting artemisinin derivative. It can be administered parenterally (IV, IM), rectally or orally. The dose for adults is 200 to 400 mg on day 1, 100-200 mg/day once daily on the following days to a total of 5 days. Since the molecule is not stable in water, the dry powder (60 mg) should be dissolved just before the injection with 0.6 ml sodium bicarbonate and 5.4 ml dextrose or dextrose/physiological fluid.

The combination of dihydroartemisinin and piperaquine, a chemical related to chloroquine, is known as Artekin in China. Dihydroartemisinin-piperaquine is believed to be one of the most effective drug combinations to treat malaria.

Lumefantrine (= benflumetol) was registered in China in 1987 for the treatment of P. falciparum malaria. It is an arylamine alcohol which was synthesised at the Chinese Institute of Military Medical Sciences. The half-life in the blood is approximately 4 days. The product is not active on the liver stages or gametocytes. Lumefantrine, like chloroquine, probably destroys haem polymerisation (a detoxifying pathway for the parasite). It is synergistic with artemether. The combination artemether-lumefantrine is used at a ratio of 1:6. Each tablet contains 20 mg artemether and 120 mg lumefantrine. This combination is also known as co-artemether (Riamet®, Co-Artem®). This combined product has been available since 1999 in Switzerland and in various African countries. There is as yet no paediatric syrup or injectable form. The recommended dose for an adult is 4 tablets once per day for 4 days (semi-immune person). For an adult with little or no immunity to malaria the advice is to take 4 tablets at the time of diagnosis, 4 tablets 8 hours later, 4 tablets 24 hours later and 4 tablets 48 hours later (i.e. 4 x 4 tablets).

Lumefantrine is a lipophilic molecule with very low toxicity. It is a yellow powder which is only partially soluble in water. Absorption in the intestine is highly variable from person to person and is greatly increased (up to 16-fold) by fatty food (similar to halofantrine). Since people who are ill generally do not eat much, this has important consequences. Early in the treatment very little lumefantrine is absorbed. In combinations such as co-artemether, the artemether is responsible for the initial important reduction in the number of parasites and the low residual numbers of parasites is then cleared up by lumefantrine.

In cases of P. vivax or P. ovale malaria, hypnozoites still remain in the liver after therapy with chloroquine. These may be destroyed by primaquine. Generally 15 mg base per day is used for 14 days [26 mg primaquine biphosphate = 15 mg primaquine base]. This is contra-indicated in pregnant women and where there is a significant deficiency of G6PD (glucose-6-phosphate dehydrogenase), an enzyme in the red blood cells (risk of haemolysis). In significant G6PD-deficiency no primaquine is given, or 0.75 mg/kg once per week for 8 weeks or 30 mg once per week for 15 weeks. There are P. vivax strains (e.g. P. vivax Chesson) which are less sensitive (India, Southeast Asia) and here 22.5 mg is given per day for 21 days. This therapy is certainly sensible for those who are finally returning from the tropics or from a malaria region. Sometimes what is called terminal prophylaxis is used. Primaquine can sometimes cause nausea, especially high doses taken on an empty stomach. Nausea is much less common if primaquine is taken with food. Primaquine also acts on gametocytes and in some circumstances (e.g. refugee camps) may be given to reduce transmission. Older products which are structurally related to primaquine [8-aminoquinolines], such as pamaquine (Plasmoquine, Praequine), rhodoquine (Plasmocide), quinocide, pentaquine and isopentaquine are almost never used any more.

Mild methaemoglobinemia (usually <13%) is frequent with primaquine, but as long as the concentration of methaemoglobin is less than 20%, there will be no clinical consequences. There is a spontaneous recovery after two weeks. People who have an inborn deficiency in methaemoglobin reductase are very susceptible to primaquine-induced methaemoglobinemia.

Etaquine or tafenoquine is a new 8-aminoquinoline, derived from primaquine. It has a half-life of two weeks, which is much longer than the half-life of primaquine. It may be taken orally and has low toxicity. It is active against P. falciparum and P. vivax. It is an effective schizonticide and is also active on the pre-erythrocytic stages, including the hypnozoites of P. vivax. The mechanism of action is still unclear, but it probably disturbs the action of the parasites’ mitochondria and the Golgi complex. There is also inhibition of the polymerisation of haematin to haemozoin (as with chloroquine). The product is administered as tafenoquine succinate. A dosage of 100 mg base corresponds to 125 mg salt. Ingestion with food increases the absorption by 50% and reduces the gastro-intestinal side effects. Absorption is slow, reaching a maximum plasma concentration after 12 hours. Tafenoquine is concentrated in red blood cells (3 times higher than in plasma). Tafenoquine is not eliminated via the kidneys. The optimum curative dosage has not yet been determined, but 300 mg per day x 7 days yielded a cure rate of 100% (P. vivax). A possible role in the treatment of P. falciparum is being investigated, although problems with its slow onset of action have still be to overcome. It is also effective as a causal prophylactic agent. People with G6PD-deficiency may develop severe haemolysis after ingestion. Development of methaemoglobinaemia (3-15% metHb) is common, but generally subclinical. Nevertheless account should be taken of this by mountain climbers, pilots or individuals with underlying cardiopulmonary disease. The product is not yet on the market.

These are biguanides which are converted in the body to the active product cycloguanil. The enzyme which catalyses this oxidative activation is probably mephenytoin hydroxylase. Absorption is delayed by simultaneous ingestion of magnesium trisilicate (e.g. stomach powders). The concentration of proguanil in red blood cells is 6 times higher than that in plasma. There is genetic polymorphism for the conversion of the prodrug to the active product. Persons are "extensive metabolisers" or "poor metabolisers". One way to determine this is by measuring the proguanil/cycloguanil ratio in the plasma. Poor metabolisers have a lower plasma level of the active form (= higher P/C ratio) and theoretically are more at risk that the drug will fail. Nevertheless, the importance of phenotype status is not really known. There is also a lack of clarity concerning the various metabolites of the product. Approximately 20% of the population of Southeast Asia are said to convert proguanil to cycloguanil scarcely if at all. In Kenya this is 35%. This does not, however, appear to diminish the efficacy of Malarone®. Chlorproguanil has chlorcycloguanil as its active metabolite. The combination of chlorproguanil with dapsone is also known as Lapdap®. It is used as a cheap, short-half-life antifolate. It may be combined with artesunate (combination known as "CDA"). Proguanil is excreted via the kidneys.

Cycloguanil is a powerful inhibitor of dihydrofolic acid reductase in the parasite. That is how the synthesis of nucleic acids in the parasite is disturbed. It has a slow action, and therefore cannot be used in monotherapy as a curative agent in an acute attack. There is swift development of resistance if proguanil is taken as the only prophylaxis. The therapeutic role of proguanil has changed recently, since the confirmation that atovaquone has fulfilled its initial promise (synergistic effect). As a prophylactic proguanil is given as one dose of 100-200 mg per day and chlorguanil as 20 mg per week.

Atovaquone (Wellvone®, Mepron®) is a lipophilic hydroxynaphthoquinone. A related product (lapinone) was discovered 50 years ago, but at that time was abandoned since it could only be given parenterally. A fatty meal increases the absorption of atovaquone in the intestines. In the blood the molecule is highly protein bound, but there are probably no significant interactions with other protein-bound drugs. Atovaquone is eliminated by the liver and can therefore be used in renal impairment. Malarone® cannot, however, be used as prophylaxis in renal failure because the blood levels of proguanil/cycloguanil are much higher. Simultaneous use of Malarone® and rifampicin is not recommended (blood levels 50% lower). Its safety during pregnancy is not known.

Atovaquone is a powerful schizonticide for P. falciparum and P. vivax. It inhibits the mitochondria of the parasite (inhibition of the electron transport) and the de novo pyrimidine synthesis. It has a half-life of 2-3 days in adults and 1-2 days in children. On monotherapy recrudescence occurs very quickly. To avoid this problem it is combined with proguanil (brand name of the atovaquone + proguanil combination = Malarone®). These products are synergistic. The combination of atovaquone + doxycycline is also active. Absorption of Malarone® via the intestine improves if it is taken with food. The recommended curative dose is 1000 mg atovaquone + 400 mg proguanil, once daily for 3 days (adults). In practice the curative regimen for an adult is: four tablets once daily for three days. An adjusted dosage is used for children, e.g. children from 11-20 kg: 1 tablet per day for 3 days, children 21-30 kg: 2 tablets par day for 3 days, children 31-40 kg: 3 tablets per day for 3 days. Malarone is not given to children weighting less than 10kg. The combination (250 mg atovaquone + 100 mg proguanil daily) can also be used for malaria prophylaxis. The dose is then 1 tablet per day, beginning the day before travelling and then taken daily until 7 days after return. There is also a causal prophylactic effect. There is no known cross-resistance with other anti-malaria products. The first cases of Malarone resistance were reported in 2001, i.e. very soon after introduction of the drug. The product is also being studied in toxoplasmosis, babesiosis, leishmaniasis, microsporidiosis and in Pneumocystis carinii pneumonia. In the treatment of babesiosis it proved more active in some animal studies than the combination of clindamycin/quinine.

This cellular organelle may possibly form a binding site for new drugs. Various metabolic reaction chains are present in this organelle. In 1998 it was demonstrated that in some Apicomplexa the "shikimate pathway" is present. The name "shikimate" comes from "shikimi no ki", the Japanese name for star aniseed (Illicium religiosum), from which shikimic acid was first isolated. The shikimate pathway is a biochemical reaction chain which occurs in algae, higher plants, fungi and various micro-organisms. It does not occur in mammals, however. It is important for the synthesis of aromatic amino acids such as phenylalanine, tyrosine and tryptophan, as well as for the production of ubiquinone and other substances. In the Apicomplexa it possibly supplies folic acid precursors which are needed for growth. The herbicide glyphosate ("Round-up") is a known inhibitor of an enzyme from this reaction chain and can block the growth of parasites. It also appears that the apicoplast synthesises isoprenoids in a manner which clearly differs from that of mammals. Isoprenoids are used for the production of cholesterol, steroid hormones, coenzyme Q and for enzyme prenylation. Humans synthesise these substances via what is called the "mevalonate pathway", known as a target for cholesterol-lowering substances such as HMG-CoA reductase inhibitors (statins, e.g. simvastatin = Zocor®). The apicoplast on the other hand, uses what is called the "methylerythritol or DOXP pathway" (1-deoxy-D-xylulose-5-phosphate). Fosmidomycin is a substance which actively blocks these reaction chains in P. falciparum. It is hoped that better knowledge of the biochemical details will lead to new therapeutic products for the treatment of, for example, toxoplasmosis, cryptosporidiosis and malaria.

In chloroquine sensitive P. falciparum the drug is concentrated in the parasite. There is only slow outflow (t½ = 50 minutes) of chloroquine from the sensitive parasite. In resistant parasites t½ for outflow = 1 to 2 minutes. Resistance is thus not due to inactivation, breakdown or neutralisation of chloroquine. The parasite quickly pumps the product away to the blood, so that the concentration of chloroquine within the parasite is low. At present this cannot be counteracted in humans (in vitro reversible with verapamil). The resistance potential is a function of the total biomass of the parasite. The lower the total biomass, the lower the probability of resistance. PCR technology [polymerase chain reaction] is required to differentiate a recrudescence (or relapse) in an endemic region from a re-infection with the same species. Several polymorphic loci are analysed. Every combination of alleles which is tested, is in itself rare and permits differentiation between strains.

The problem of chloroquine resistance in P. falciparum became manifest in the 1960s in South America (Colombia and Brazil) and in Southeast Asia. Gradually the resistance spread to other continents. Resistance also developed against other drugs, including Fansidar®. The situation is evolving rapidly and is getting worse. The highest resistance is found in some regions in Southeast Asia, including Cambodia, the Thailand-Cambodia border, and the Thailand-Burma border. It is only a question of time before these resistant strains spread further geographically. There are several reasons for this increasing resistance. Important factors include inadequate individual patient compliance, treatments that are are often discontinued prematurely, frequent underdosing, earlier mass-treatment campaigns reaching only part of the population and therapy being sometimes only partly administered, as well as the use of chloroquinated salt (Cambodia). Among the causes of the swift increase in geographical spread are the large-scale migrations of today, and the ability to move rapidly from place to place. Some products are eliminated slowly from the body (e.g. mefloquine t¹/₂ = 2 weeks) so that for some weeks a subtherapeutic level of the product is present in the body. When malaria parasites are exposed to such low concentrations, partially resistant strains have a selective advantage. The occurrence of subclinical cases functions as a source and reservoir for transmission of parasites with reduced sensitivity. Since the cost price of alternative drugs is generally higher than that of traditional treatments, under-dosing with new drugs will become even more important in future. New drugs are developed only slowly. Combination therapies with for example artemether-mefloquine, co-artemether, atovaquone-proguanil, comparable to the present-day quinine-doxycyclin, will become more common in future. The principle behind this approach can be compared with the present combination treatments for AIDS, tuberculosis and leukaemia.

Direct consequences of the increasing resistance of P. falciparum malaria:

• an increase in mortality and morbidity
• a delayed initial therapeutic response
• a shorter interval before recrudescence occurs.