Why are Artemisia infusions prophylactic? the herbicidal hypothesis

juin 3, 2016

Ἓν οἶδα ὅτι οὐδὲν οἶδα
Our association IFBV-BELHERB has received numerous anecdotic reports on the prophylactic effects of Artemisia plants. This effect has been documented in scientific papers. Patrick Ogwang from Uganda (Ogwang PE, et al. Trop J Pharm Res. 2012;11:445–53) showed that an infusion of Artemisia annua consumed once weekly reduced the risk of Plasmodium falciparum episodes due to a yet unidentified constituent. All this is an important lead as classical antimalarial drugs like quinine (D.Shanks, Am Soc Trop Med Hyg, 2016, May), chloroquine (T Sahu et al., Frontiers in Microbiology, 2015, 1, 283), artemisinin are not prophylactic. They only act on the erythrocytic stage but have no impact on the liver stage invasion. They are just designed to kill the parasites in the erythrocytes, but they leave a bloody battlefield and a depressed immune system. Some even enhance the gametocytogenesis.
Many medicinal plants used against malaria in endemic areas are aimed to treat the acute symptoms of the disease such as fevers and their action is limited to these symptoms. In some endemic areas of the Brazilian Amazon region one medicinal plant seems to be an exception: Ampelozyziphus amazonicus, localla named “Indian beer” used to prevent the disease when taken daily as a cold suspension of powdered dried roots (VF Andrade-Neto et al., Int J Parasitology, 2008, 38, 1505-11). In infections induced by sporozoites, chickens treated with extracts of this plant were partially protected against Plasmodium gallinaceum. Some animals did not become infected, whereas others had a delayed prepatent period, lower parasitemia and a reduction in mortality. A delay in the time until parasites establish blood stage infection is not a minor effect. It reduces the risk of severe and cerebral malaria.
In a previous document published on http://www.malariaworld.org “Pentacyclic triterpenes in antimalarial plants, a new paradigm” we alerted to the important role these acids play in malaria control. And indeed Ampelozyziphus amazonicus is very rich in pentacyclic triterpenes, mainly betulinic acid (D do Carmo et al., Pharmacognosy Magazine, 2015, 11, 244-250). Betulinic acid, maslinic acid, oleanolic acid, ursolic acid and others seem thus to be responsible for the prophylactic activity of these plants. But that does not explain why.
Another good example for the antimalarial and prophylactic effect is Phyllantus amarus. This plant is well known for these properties in Ghana (R Appiah-Opong et al., Ghana Medical Journal, 2011, 45, 143-146), in Burkina Faso (M Traore et al., Phytother Res. 2008. 22, 550-1, in Nigeria, RDCongo (L Tona aet al., J Ethnopharmacol. 2004, 93, 27-32), but also in China, India, Brazil. The aqueous extract shows suppressive and curative properties similar to standard antimalarials like chloroquine, or artesunate, but it also shows prophylactic properties by delaying the onset of infection (T Ajala et al., Asian Pacific J Trop Med 2011). The plant is very rich in pentacyclic triterpenes.
The apicoplast is a plastid organelle, homologous to chloroplasts of plants or algae, that is found in apicomplexan parasites like Plasmodium or Toxoplasma. In hindsight it seems incredible that such an organelle with a nonmammalian metabolism could so long have concealed its identity in parasites that have received as much scientific attention as Plasmodium (RF Waller et al., Curr Issues Mol Biol 7, 2005, 57-80). It is now recognized that this large group of parasites had a photosynthetic ancestry and were converted into parasitism early in the evalution of animals. Apicoplast function is necessary for both intraerythrocytic and intrahepatic development. Recently it was found that the apicoplast is also present in the gametocytogenesis, in the sexual stage of Plasmodium falciparum. But only the female macrogametes have an apicoplast, the male microgametes don’t (N Okamoto et al., Eukaryot Cell 8(1) 2009, 128-132).
Attempts have been made to find antimalarials which selectively attack the apicoplast. Several antibiotics are known to interfer with this organelle. They do not kill parasites in the first generation, but the progeny of drug-treated parasites suffer a delayed death, probably because they lose their apicoplast and the second generation of merozoites is unable to penetrate erythrocytes. Antibiotic treatment specifically inhibits the biogenesis and inheritance of the apicoplast in Plasmodium falciparum liverstage, resulting in continuous liver stage maturation but subsequent failure to establish blood-stage infection. This process of maturation of numerous merozoites in the liver induces potent immune protection for subsequent infections. This prophylactic protection obtained by several antibiotics was demonstrated to be exceptionally robust (J Friesen et al., http://www.ScienceTranslationalMedicine.org, 2010, 2).
Many herbicides also have an activity against the malaria protozoan (S O Duke, Weed Science, 58, 2010, 334-339). Herbicides interfer with plant cells by interrupting mitosis and the formation of multinucleated cells. A good example is the action of herbicides from the nitroanilin family (trifluralin, oryzalin). They have a strong action on Plasmodium falciparum. The herbicide amiprophosmethyl also has antimalarial activity and a significant action on schizogony, i.e. the formation of multinucleated parasites. A more surprising example is the effect of the well known glyphosate on several apicomplexa like Plasmodium falciparum, Toxoplasma gondii (F Roberts et al., Nature 393, 801-805, 1998).
Several herbicides including those of the cyclohexanedione family act by perturbing the apicoplast fatty acid biosynthesis (C Goodman et al., Int J Parasitol. 2014, 44, 285-289). This synthesis is dispensable in blood stages of human and rodent malaria, but vital for for the liver stage.
Artemisia annua has strong allelopathic properties as was documented by Mediplant in Switzerland. In other words the plant becomes invasive and inhibits the growth of other plants or cash crop on fields where Artemisia has been planted for the extraction of artemisinin. A recent paper from Iran (MH Bijeh Keshhavarzi et al, J Biol & Envir Sci. Nov 2014) describes the allelopathic effects of Artemisia annua on lettuce Lactuca sativa. The aqueous extract on an outside plot significantly reduced germination percentage and rate, fresh and dry weight. Another paper (Seyed Mohsen et al., Annals of Biological Research, 2011, 2-6, 687-69) describes the allelopathic effect of Artemisia annua aqueous extracts on vegetables and plants like Portulaca olearcea (pursley), Chenopodium album (goose-foot), Avena ludoviciana (oat), Plantago ovata (plantain). For the latter the effects are noticeable on germination percentage, germination rate, plumule length, radicle length, wet weight, dry weight. A Chinese paper (Shen He et al., Ying Yong Sheng Tai Xue Bao. 2005 Apr;16(4):740-3) had previously studied the allelopathy of different plants. Artemisia annua affected the seedling height and fresh weight of radish, cucumber, wheat and maize around 50%.
But the allelopathy of Artemisia remains a controversial issue. Although in vitro trials on seed germination of various plants show an impact of artemisinin and flavonoids, this remains far from field effects noticed and may only offer a partial explanation, Artemisinin and flavonoids are hardly soluble in water and rapidly degraded in the soil.
A recent paper offers an explanation which is very attractive proposing the role of pentacyclic triterpenoids in the allelopathic effects of Alstonia scholaris. (Wang CM1, J Chem Ecol. 2014 Jan;40(1):90-8). Alstonia scholaris is a tropical evergreen tree native to South and Southeast Asia. Alstonia forests frequently lack understory species. However, potential mechanisms, particularly the allelochemicals involved remain unclear. They identified allelochemicals of A. scholaris, and clarified the role of allelopathic substances from A. scholaris in interactions with neighboring plants and showed that the allelochemicals from leaves, litter, and soil from A. scholaris were identified as pentacyclic triterpenoids, including betulinic acid, oleanolic acid, and ursolic acid. In the field, ursolic acid accumulated abundantly in the soil in A. scholaris forests, and suppressed weed growth during summer and winter. A. scholaris pentacyclic triterpenoids influence the growth of neighboring weeds by inhibiting seed germination, radicle growth, and functioning of the photosystem.
These molecules are stable in powder form and in water for months (P Puttarak et al., Natural Product Sciences, 2016, 22, 1-20)
If the pentacyclic triterpenes have an allelopathic, herbicidal effect, this offers a completely new hypothesis and opportunity for destroying the malaria parasite in all its forms, from hepatocytes to gametocytes.
As the primary interface between the body and the outside environment, the skin protects the host against invading organisms.
It has always been assumed that sporozoites rapidly exit the injection site and enter the blood circulation. But it was demonstrated by PCR that the majority of the infective sporozoites remain in the skin for hours (L M Yamauchi et al., Cellular Microbiology, 2007, 9, 1215-22). The same authors had shown in preliminary studies no decrease in the sporozoite loads in the skin up to 3h. These findings imply that there is ample time for host and parasite to interact at the inoculation site and that tailored treatments of the skin might inhibit the survival of the sporozoites before they enter the bloodstream. At 37°C the time required for sporozoite penetration in hepatoma cells is 3 hours (VF Andrade-Neto op.cit.). Malaria-specific CD8⁺ T cells are primed in the skin draining lymphs (S Chakravarty et al., Nat Med 2007, 13, 1035-1041). A significant reduction in anti-sporozoite CD8⁺T cell response was observed in animals that had their draining lymph nodes removed prior to sporozoite infection. Pentacyclic triterpenes could play a role. Topical application of ursolic and maslinic acid for example significantly reduces epidermal inflammation, NF-κB, Cox-2 and skin tumor proliferation (Jiyoon Cho et al., Oncotarget, 6-36, 39292-305).
The high concentration of pentacyclic triterpenes in the skin of fruits or barks of trees evidently has the function to repel parasites and molds and to prevent their entry into the fruit or plant. Betulinic acid acts as an antifeedant (SG Jagadesh et al., J Agric Food Chem. 1998, 46, 2297-99). It affects the growth of larvae and pupae (V Lingampally et al., Asian J Plant Sc and Res. 2012, 2, 198-206).
Unlike many other microbial organisms that utilize the phagocytic properties of their host for invasion, sporozoites actively invade hepatocytes. They even pass through several hepatocytes prior to the final hepatocyte in which they develop. The reason for this process is not well understood; it is likely that the sporozoites choose the best environment for their differentiation into merozoites. Sporozoites have specialized secreting organelles in the apical region which play a central role in host cell invasion. The migration through several hepatocytes increases sporozoite competency for the formation of these apical organelles.
To survive and develop in the parasitophorous vacuole inside the hepatocyte the parasite has developed several strategies including depletion of CD-8 lymphocytes and suppression of NF-κB to prevent cell death. The circumsporozoite protein (CSP) plays a key role in this inhibition of NF-κB.
Memory CD8⁺T cell populations residing in the liver provide the first line of defence against naïve infections, but more even in subsequent infections and are the key players of the recall response. Any immune strengthening approach, including vaccines, requires the generation of a robust, stable memory population. In particular, CD4⁺cell help was shown to be required for CD8⁺memory cell responses against malaria (She-Wak Tse et al., Mem Inst Oswaldo Cruz 2011, 106, 172-178). CD4⁺helper T cells are critical orchestrators of immune responses. CD4⁺T cells decrease the threshold required for protective immunity and this immunity may last for months (NW Schmidt et al., PNAS, 2008, 105, 14017-22).
It was shown that oleanolic acid upregulates the CD4⁺and CD8⁺populations (J Wang et al., International Immunopharmacology 2012, 14-4). It was also found that betulinic acid increased the total number of thimocytes, splenocytes, lymphocytes. It is a potential biological response modifier and may strengthen the immune response of its host (Y Jine Polish J Veterinary Sc. 2012, 15, 305-13). This is in line with our findings in Katanga where we confirmed that administration of capsules containing Artemisia leaf powder raised the CD4⁺ (Constant Kansongo Tchandema, personal communication). This strengthening of the immune system by Artemisia plants may also be related to the pentacyclic triterpenes they contain.
Delayed apoptosis of infected hepacytes is another strategy of defence of the sporozoites. Blocking apoptosis allows the parasite to complete its full cycle and the merozoite burden is strongly enhanced. It fully exploits host cell resources and ultimately produces tens of thousands of merozoites which are released from the hepatocyte. Sensitizing the infected hepatocyte to apoptosis may substantially reduce parasite burden. Recent evidence suggests that Plasmodium infected hepatocytes are similar to cancer cells. The authors demonstrate that the anticancer drug obatoclax reduced the number of infected hepatocytes by > 70%, but had limited effect on uninfected cells (A Kaushansky et al., Cell Death and Disease, 4. E762). Many pentacyclic triterpenes, like betulinic acid, are used in cancer treatment. It seems worthwile to study their effect in malaria infection.
Several antibiotics also have an effect during the hepatocytic stage. Others, including tetracyclines and clindamycin used for the treatment of malaria, have little action on the pre-erythrocytic stages.
An extensive study has been made on the antibiotic thiostrepton and the effects of this drug on life-cycle stages of the malaria parasite in vivo. Preincubation of mature infective sporozoites with thiostrepton has no observable effect on their infectivity. Sporozoite infection both by mosquito bite and sporozoite injection was prevented by pretreatment of mice with thiostrepton. Thiostrepton eliminates infection with erythrocytic forms of Plasmodium berghei in mice. (M Sullivan et al., Mol Biochem Parasit 2000 109, 17-23).
After its release from the hepatocyte the merozoites penetrate the erythrocytes very rapidly. Parasite entry into erythrocytes is a complex, dynamic process. The invading merozoite orients its apical end toward the junction of invasion. Invagination of the erythrocyte bilayer then results in engulfment of the parasite. Merozoites without their apicoplast are unable to penetrate red blood cells.
A study of the Walter Reed Army Institute already in 1992 found that the herbicide Trifluralin showed strong anti-malarial effects not only on Plasmodium falciparum in cultures, but also transmission blocking by inhibiting gametocyte maturation and viability (J Nath et al., 19th Army Science Conference, 1994).
Thiostrepton (op.cit) treatment of infected mice reduces transmission of parasites by more than ten-fold, indicating that the plastid has a role in sexual development of the parasite. These results also indicate that the plastid function is accessible to drug action in vivo and important to the development of both sexual and asexual forms of the parasite.
Supply of the isoprenoid building blocks isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) is the essential metabolic function of the apicoplast for isoprenoid biosynthesis, particularly during gametocytogenesis. When IPP supplementation was removed early in gametocytogenesis, developmental defects were observed, supporting the essential role of isoprenoids for normal gametocytogenesis. Furthermore, mosquitoes infected with gametocytes lacking the apicoplast developed fewer and smaller oocysts that failed to produce sporozoites. This finding further supports the essential role of the apicoplast in establishing a successful infection in the mosquito vector (J Wiley et al., Eukaryotic Cell 2015, 14, 128-133)


Was the Nobel price for artemisinin a fatal error

avril 14, 2016

Was the Nobel prize for artemisinin a fatal error?

April 2, 2016 – 06:26 — Irene Teis
In 2015 a Nobel Price was attributed to Youyou Tu, almost 50 years after a report describing artemisinin’s structure, pharmacology, and efficacy had been published in 1979 by the “Qinghaosu Anti-Malarial Coordinating Research Group,” where she was a member of. Mr Huang Shuze, Deputy Minister of Health, stated in his 1981 summary report “Project 523 mobilized multiple departments ; thirty scientific research units and medical schools in 1975”.

WHO for decades hesitated in considering this traditional medicine approach. Only at the end of the nineties, when chloroquine’s resistance became overwhelming did first clinical trials take place. But artemisinin was not water soluble, hardly bioavailable, metabolized very rapidly and gave premature signs of resistance. WHO then prescribed in 1998 extremely high doses up to 1 200 mg of artemisinine for a person of 60 kg on the first day of treatment (WHO/MAL/98.1086) , at the verge of severe neurotoxicty and hepatoxicity.

A RECENT PAPER RINGS AN ALARM BELL. Plasmodium chabaudi malaria parasites through a step-wise increase in artesunate dose evolve extremely rapidly slow clearance rates. These slower clearance rates provide fitness advantages to the parasite through increased overall density, recrudescence after treatment and increased transmission potential. Removal of only the susceptible parasites by artesunate treatment led to substantial increases in the densities of resistant parasites (LC Pollit et al., PloS Pathogens 2014, 10,4, e1004019). The traditional view has been that aggressive chemotherapy , involving high doses applied for sufficiently long time to eliminate parasites, best minimizes the evolution of resistance. For this reason WHO in several statements has condemned the use of Artemisia annua herbal treatment because of low artemisinin concentrations in the infusions. But numerous clinical trials, small and large, demonstrated that Artemisia annua and Artemisia afra infusion or powdered leaves reduce parasitaemia much more efficiently than ACTs and eliminate all gametocytes (see Breaking News for clinical trials with Artemisia plants, on http://www.malariaworld.org).

Previous work using the anti-malarial pyrimethamine has shown that removing susceptible competitors through drug treatment can lead to dramatic increases in the density of resistant parasites.

Resistance can also be affected by the dormancy effect (A. Codd et al., Malaria Journal, 10:56, 2011). One of the side effects of the higher doses of artemisinin is this effect induced in plasmodium. The parasite encapsulates itself against the aggressive peroxide artesunate and reawakens at the end of the treatment. The same effect is called quiescence by a French research team (B. Witkowski et al., Antimicrob Agents Chemother. Doi:10.1128/AAC.01636-09). Under a very high dose of artesunate, a Plasmodium falciparum ring-stage sub-population persists in culture and continues its cycle of development normally after drug removal. This may be one of the causes of resistance.

During the Ebola crisis in West Africa. Medecins sans Frontières administered 1.5 million treatments of artesunate-amodiaquine in Sierra Leone in a mass drug administration campaign. This is the largest-ever mass distribution of antimalarials. It is impossible to find any results on PubMed? Was it a failure with dramatic consequences, deathtoll or resurgence?

Irene Teis

MSF-Coarsucam clinical trial with horrendous death toll

Submitted by Marc Vanacker (not verified) on April 4, 2016 – 17:46

There is indeed a peer reviewed paper (E Guignoux et al., NEJM, 2016 ;374 :23-32) which refers to the MDA clinical trials made by Médecins sans Frontières with artesunate-amodiaquine during the 2014 Ebola crisis. Not in Sierra Leone but in Liberia. Not with 1 500 000 patients but with 382 (three hundred eighty two).
As per Table 2 there were three branches in the trial : 194 patients for Coartem, 71 for Coarsucam and 63 with no antimalarial drug prescription.
In the Coartem group 125 (64.4%) died, in the Coarsucam group 36 (50.7%) and in the no drug group 41 (65.1%).
It is surprising that the total number of patients quoted in the abstract : 382, does not match the total number of patients in table 1 : 278, neither in table 2: 328, nor table 3: 282 nor in table 4: 295. This needs to be clarified as the statistics become dubious
Whatever, the authors make the dazzling claim that the 71 patients who were prescribed artesunate-amodiaquine had a lower risk of death than did patients who were prescribed artemether-lumefantrine.
A pyrrhic victory for Médecins sans Frontières on a bloody battlefield. Sanofi-Aventis will evidently continue to subsidize them. Amodiaquine is banned in France and artesunate-amodiaquine will now probably become available in French pharmacies

Artemisia afra will save Africa

avril 14, 2016

March 23, 2016 – 16:23 — Pierre Lutgen lutgenp@gms.lu
A very recent paper of a South African research team shows that among 8 medicinal plants Artemisia afra has the lowest IC50 for impairing the development of late stage gametocytes (P Moyo et al., J of Ethnopharmacology, acceopted 15 March). A very important finding as not many plants have such a significant gametocytocidal effect.

It confirms the in vivo results obtained end of 2015 in a large scale, double blind randomized clinical trials in Maniema, RDCongo (see Breaking news from clinical trials with Artemisia plants) where Artemisia afra was one of the branches of the test. Artemisia herbal tea completely eliminated gametocytes but they were still present on day 28 in 10% of those treated with Coartem In 2013 already Dr Constant Kansongo in Katanga had found in a trial with 44 Plasmodium falciparum infected patients that after 7 days of treatment with 20 gr of capsules containing A afra powder the gametocytes had completely disappeared, except for one patient. Artemisia afra does not contain any artemisinin. The best explanation available is the high arginine content of Artemisia plants (see « Arginine, a deadly weapon against gametocytes » on malariaworld.org). Frank van der Kooy at the University of Leiden found that Artemisia afra has anti-HIV properties stronger than Artemisia annua.

The situation is completely different for artemisinin derivatives and ACTs, it is even alarming. A paper from Mali published in February clearly shows it (AA Djimbe et al., Parasite, 2016. 23, 3). Artesunate does not clear mature gametocytes during oral artesunate treatment and does not prevent the appearance of new gametocytes. The same recrudescence with oral artemisinin monotherapy had already been observed in Vietnam in 2001 (PT Giao et al., Am J Trop Med Hyg, 2001 65 690-695). The conclusion of the authors was that artemisinin monotherapy may offer rapid recovery and fast parasite clearance, but recrudescence is frequent. For up to 20 percent of the cases on day 28, although gametocytes had completely disappeared on day 7. Extending the duration of the monotherapy from 5 to 7 days did not reduce recrudescence. A study from Kenya had also found that gametocyte carriage was much lower on day 14 than on day 28 and 42 for artemether lumefantrine, but not for dihydroartemisinin-piperaquine (P Sawa et al., J Infect Dis, 2013, 207, 1637-45). It is well known that artemisinin drugs are gametocytocidal for immature, but not mature gametocytes (GO Ghotosho et al., Mem Inst Oswaldo Cruz 2011, 106 no5). A paper of the Swiss Tropical and Public Health Institute (BJ Huho et al., Malaria Journal, 2012 11:118) comes to the conclusion that in high perennial transmission settings case management with ACT may have little impact on overall infectiousness of the human population. They even found in their study, that the most direct indicator of human-to-mosquito transmission, namely oocyst prevalence was substantially higher after ACT introduction. A study from Burkina Faso found in a recheck 12 months after a clinical trial with ACTs that the number of symptomatic malaria episodes was even slightly higher in the ACT arm than in the control arm and that after several treatments the prevalence of gametocyte carriers was the same in both arms (AB Tiono et al.,Malaria Journal 2013, 12:79). Another study found that ACT did not significantly reduce the proportion of infectious children. Submicroscopic gametocytaemia is common after treatment and contributes considerably to mosquito infection. (JT Bousema J Infect Dis., 2006, 193, 1151-59). Because of the short half-life of artemisinin and because high doses induce dormancy in the asexual parasite, asexual forms, mostly rings, remaining after completion of ACT may develop into mature gametocytes 7-15 days later. Some patients have the first appearance of gametocytemia 4-8/day after completion of a 3 day-ACT. (Wilairatana P, et al.,Southeast Asian J Trop Med Public Health. 2010 Nov;41(6):1306-11). What worries the authors of the study from Mali is not only that similar results had been found in a study in 2002-2004, but the fact that baseline gametocyte carriage was significantly higher 6 years after deployment of ACTs in this setting. If artemisinin derivatives really enhance recrudescence and gametocyte carriage, this is indeed alarming. It would mean that ACTs will not eradicate malaria but enhance it in the long run.

When IFBV-BELHERB had raised this concern with WHO Geneva and ITG Antwerp the blunt answer received from one of the experts was: “Your arguments do not make any sense from a public health point of view ».

Artemisia afra is growing wild from The Cape to Addis Abeba.

No further need to import Nobel prize validated pharmaceutical drugs from China or Europe

Malaria, folates and PABA

avril 14, 2016

Malaria, folates and PABA

April 13, 2016 – 17:38 — Pierre Lutgen

Folates combine three molecules : pretidine & para-aminobenzoic acid (PABA) & glutamate. They were discovered around 1940 and first isolated from spinach leaves. The term folate is derived from the latin word folium.
The malaria parasite has a unique feature of being able to salvage exogenous folate derivatives and/or synthesize them de novo. Due to its high rate of replication, the parasite has a high demand for folates. Folate metabolism is the target of several antimalarials.
Food fortified with folic acid has been available for consumption in North America for over two decades. African countries are now embracing this concept; however, because folate promotes malaria parasite division, as it does in cancer cells, there is a possibility of malaria exacerbation if folate intake is increased. (Nzila A1Food Nutr Bull. 2016 Mar 4. pii: 0379572116634511).
The detrimental role of PABA (para-amino benzoic acid) on malaria has already been described 60 years ago (F Hawking, British Medical Journal, 1954, Feb, 425-429). Rats fed on a milk diet were insusceptible to infection with Plasmodium berghei. Milk does not contain PABA or only traces. This insusceptibility was reversed by the addition of PABA or folic acid. The same experiences were repeated on monkeys and gave the same results. It is likely that the relative immunity to malaria shown by infants in many parts of the tropics may be due to a deficiency of PABA in their mother’s milk.
In 1991 it was found that feeding wistar albino rats on low protein and low energy diet caused suppression of P berghei parasitaemia. When PABA was added to the diet parasitaemia re-elevated (A Bhatia et al Indian J Malariol 1991, 28 237-42). The same effect had already been inadvertely noticed in in vitro trials (CF Gilks et al., Parasitology, 1989 89 175-177).
Another research team showed that dietary folate deficiency protects primates against malaria (KC Das et al., Blood, 1992, 80-281). Blood infected with Plasmodium cynomolgi was injected into-folate deficient animals and folate-replete control animal. All control animals developed malaria and several died.
A more recent study extensively studies the effect of dietary PABA on murine Plasmodium yoelii infection (GA Kiczska et al., JID, 2003, 188, 1776-81). Plasmodium species, unlike humans, can utilize PABA for de novo generation of folate. The authors show that, despite the presence of biosynthetioc machinery to synthesize PABA, Plasmodium yoelii, a rodent malaria species, requires exogenous dietary PABA for survival. Mice fed low-PABA-diets do not die from lethal doses of P.yoelii. The initiation of a PABA-deficient diet after P.yoelii infection is established, leads to the clearance of parasites and subsequent resistance to infection by P. yoelii. An intact immune system is not necessary for protection given that mice with severe combined immunodeficiency were also protected by PABA-deficient diet.
In a trial made in The Gambia involving 600 children with uncomplicated falciparum malaria, among children who received the antifolate sulfadoxine-pyrimethamine, the treatment failure rate was significantly higher in those given folic acid than those given placebo (MB van Hensbroek et al., Trans R Soc Prop Med Hyg 1995,89, 672-6). And the authors suggest that the WHO recommendation of universal folic acid supplementation should exclude children in areas of high prevalence.
In a randomized, double blind prophylactic trial in Zanzibar the authors had to conclude that the routine supplementation with iron and folic acid in preschool children in a population with high rates of malaria can result in an increased risk of severe illness and death. (Sazawal S et al., Lancet. 2006 28;367(9507):302.
Another more recent study confirmed that high dietary folate in mice alters immune response and reduces survival after malarial infection (DN. Meadows, et al., PLOS One 2015 Nov 24. doi: 10.1371/journal.pone.0143738)
In case of malaria infection diet should be low in folates or PABA. Swamping Africa with multiple micronutrient powders (MNPs)from Switzerland, , nutraceuticals from the US, “compléments alimentaires” from France, all containing folates, is questionable. The folic acid fortified milk market is booming. Business on the verge of crime.
A plant which could be detrimental during malaria infection is Moringa oleifera. The average folate in vegetables is 40 microg/100g but in Moringa oleifera it goes up to 540 microg/100g DW (K Witt Echo Research Note No 1, 2012). Moringa is rich in glutamic acid – 5 times more than Artemisia- and para-aminobenzoic acid (PABA), two of the building blocks of folate (G Magnani et al., Biochem J, 2013455, 149-155). PABA is a major constituent in Moringa oleifera and soya (L Mbanga et al., Adv Biochem & Biotechnol., 2015, 1, 1-13). It was never detected in Artemisia annua. A recent paper studied the relative bioavailability of folate from the traditional food plant Moringa oleifera L. as evaluated in a rat model. The bioavailability of folate from dried leaves was 81.9%, which is much higher than the values of 50% known for other plants (Saini RK et al., J Food Sci Technol. 2016 ;53:511-20).
Several recent large scale trials in RDCongo (see “Breaking news from clinical trials with Artemisia plants, malariaworld.org) have shown that Artemisia annua and Artemisia afra completely eliminate gametocytes from malaria infected patients.
This is very encouraging for those who really want to eradicate malaria.
But another research team found that PABA administered to gametocyte-carrying mice increased the number of oocysts in mosquitoes fed on them (W Peters Ann Trop Med Parasitol 1980, 74, 275-82). High PABA content in the diet leads to the selection of drug resistant parasites in mice. A higher yield of resistance was related to the higher parasitaemia generated by PABA (B Merkli et al., Exp Parasit 1983 55, 372-6).
That is worrying. We are supposed to know. But the malaria experts from WHO and Tropical Medicine Institutes close their eyes on it.
Pierre Lutgen

Combiner les antirétroviraux avec les antipaludiques peut être fatal

février 14, 2016

Lors de la conférence du 19 mai 2014 à l’Université du Nebraska qui portait sur le SIDA, F.A. Fehintola montrait que la nevapirine prescrite simultanément avec l’artemether-lumefantrine (Coartem) réduisait de 70% la concentration du principe actif lumefantrine dans le sang infecté.

Ceci ne fait que confirmer des résultats obtenus en Afrique du Sud (T Kredo , Antimicrob Agents Chemother. 2011 Dec; 55(12):5616-23). L’artemether et la nevapirine sont métabolisés par le cytochrome P450 3A4 induit par la nevapirine.
Pauline Byakik à la Makerere University a elle aussi constaté que l’administration simultanée du Coartem avec efavirens diminuait les effets de l’artemether et de la lumefantrine. (J Antimicrob Chemother 2012, 67, 2213-2221). Le même effet a été décrit par T Kakuda et al., (HIV Therapy Workshop, Barcelona, 16-18 Mar 2012).
L’antagonisme a été observé non seulement avec les dérivés de l’artemisinine mais également avec les dérivés de la quinine chez la femme enceinte. (K Kayentao, Am J Trop Med Hyg. 2014 Mar; 90(3):530-4). L’amodiaquine est 40% moins concentrée dans le sang des patients traités par neviparine (K Scarsi et al., J Antimicrob Chemother 2014, 69, 1370-76).
Le traitement intermittent à la méfloquine se traduit par une transmission accrue du VIH de la mère à l’enfant. (R Gonzalez et al., PLOS Medecine 2014, 11-9).
Beaucoup de touristes européens utilisent la malarone comme prophylactique. La molécule active de ce remède, l’atovaquone-proguanil se retrouve à de plus faibles concentrations chez les patients prenant en même temps de l’efavirens, du lopinavir, du ritonavir ou du atazanavir (van Luin M1, AIDS. 2010 May 15;24(8):1223-6).
Ces observations ont été résumées par Van Geertruyden JP. (Clin Microbiol Infect. 2014 Apr;20(4):278-85). Les interactions pathophysiologiques et epidemiologiques entre VIH. et pathogènes tropicaux, surtout ceux du paludisme, commencent à créer de fortes appréhensions dans les pays du Sud. Deux décennies de recherches confirment que la liaison entre l’immunosuppression du VIH est corrélée avec l’incidence du paludisme, avec les échecs dans son traitement et les cas de malaria sévère. Le risque d’un développement de la résistance aux traitements prescrits par l’OMS se trouve donc multiplié. La recommandation de l’OMS de juin 2013 de traiter les mères qui viennent d’accoucher et les nouveau-nés pendant la période d’allaitement avec des ARV chaque jour pendant six semaines n’a pas trouvé de confirmation scientifique de ses bienfaits lors d’essais cliniques. Le prétendu effet contre le paludisme et sa transmission n’a pas pu être constaté (P Natureebaet al., JID 2014 :210). The Lancet a publié une étude qui montre plutôt un effet contraire : le ritonavir et le saquinavir réduisent de plus de 50% le nombre d’erythrocytes phagocytés par les macrophages (S Nathoo et al., The Lancet, 362, 2003. 1039-1041).

Le plus grave c’est que le traitement antiretroviral augmente la fréquence des porteurs asymptomatiques de Plasmodium. De 15.4 à 44.4 selon une étude du Niger. Et ces porteurs asymptomatiques ont une densité de parasites beaucoup plus élevée. (Adetifa et al., Niger Postgrad Med J, 2008 15. 141-145).
Une bombe à retardement pour l’Afrique.
Déjà en 2006 F Nosten et U d’Alessandro avaient insisté pour qu’on fasse des études approfondies sur les interactions entre antirétrovirax et antipaludiques, surtout chez les femmes enceintes (Curr Drug Saf 2006, Jan 1-15).
Ceci n’a pas empêché une inondation du marché africain avec ces deux remèdes. Un business qui se chiffre dans les milliards de dollars.
Pierre Lutgen

Breaking news from clinical trials with Artemisia plants

février 9, 2016

A team of medical doctors in RDCongo, Jerome Munyangi and Michel Idumbo, have run randomized clinical trials on a large scale in the Maniema province with the participation of some 1000 malaria infected patients. The trials were run in conformity with the WHO procedures and compared Artemisia annua and Artemisia afra with ACTs (Coartem and ASAQ). For all the parameters tested herbal treatment was significantly better than ACTs: faster clearance for fever and parasitemia, absence of parasites on day 28 for 99.5% of the Artemisia treatments and 79.5% only for the ACT treatments. A total absence of side effects was evident for the treatments with the plants, but for the 498 patients treated with ACTs, 210 suffered from diarrhea, and/or nausea, pruritus, hypoglycemia etc. The efficiency was equivalent for Artemisia annua and Artemisia afra. More important even is the observation for the total absence of gametocytes after 7 days treatment with the herb. A tremendous hope for malaria eradication. The results have been communicated to the local health authorities, and to the Ministries of Health and Research in the RDCongo who were supportive of these trials. The draft of a paper is almost ready and will be submitted to a peer reviewed scientific journal.
The financial support for the trials comes from the association MoreforLess in Paris.
This is not the only clinical trial run in 2015 with Artemisia annua aqueous infusions. In Benin the University of Abomey, together with the Universities of Louvain and of Liege, run a large scale trial with Artemisia annua grown in Benin. The trial involved 130 malaria infected patients. Fever clearance was evident after 48 hours and parasitemia decreased by 70% already on the first day, and remained 100 % absent on days 14 and 28. No side effects were noticed. The research team from Benin strongly recommends for African countries to replace the expensive and now often ineffective ACTs by Artemisia annua tea (H Zime-Diawara et al., Int J Biol Chem Sci 2015, 9-2, 692-702).
These results confirm results obtained by the association IFBV-Belherb and her partners in many small scale trials in several African countries over the last 6 years. Therapeutic efficiency always was > 95% and prophylaxy was noticed and documented. The abstracts or peer reviewed papers of all these trials are available on request.

Artemisia defeats schistosomiasis

février 9, 2016

In parallel with the clinical trials run by a team of medical doctors in the province of Maniema on the efficiency of Artemisia annua and Artemisia afra against malaria, (see Breaking News Jan.5 on http://www.malariaworld.org) they have completed another large scale randomized, double blind trial against schistosomiasis, Artemisia vs Praziquantel.
The results confirm previous anecdotic results from several countries in Africa. Both arms in this trial had 400 infected patients. The treatment efficiency was 97 % in the Artemisia arm and 71% in the Praziquantel arm. No side effects were noticed in the Artemisia treatment. Praziquantel caused vomiting in 26.5% of the patients, abdominal pain in 18.5%, cephalalgy in 15.5%.
Very impressive is the fact that the Artemisia treatment led to an unexpected almost complete absence of eggs in feces after 2 months.
Schistosomiasis kills 150 000 Africans per year and more than 70 000 000 are infected. A neglected disease in neglected, poor populations where the only existing drug, Praziquantel, loses efficiency year after year.
These young African doctors have run these trials despite the oposition and intimidations of BigpharmaWHO. They will win the battle against neglected tropical diseases where Western chemical monotherapies have failed

Jerome Munyangi, Michel Idumbo, Lucile Cornet-Vernet, Pierre Lutgen

More on diabetes and malaria from Palestine

décembre 18, 2015

n diabetes and malaria from PalestineAs stated in previous blogs diabetes is on the rise everywhere. It is a debilitating and often fatal disease per se but it also increases the incidence of malaria because higher glucose contents in the blood are a fertile terrain for Plasmodium falciparum.
It is the general belief that malaria patients are poor in antioxidant defenses and that supplementation with antioxidants (vitamins, polyphenols) will alleviate the severity of malaria infections. It is thus surprising that the University of Al Quds finds that the total antioxidant status in type 2 diabetic patients in Palestine in significantly higher compared to control subjects. The study involved 212 diabetic subjects and 208 normal subjects (AT Kharroubi et al., J Diabetes Research, 2015 ID461271).
A similar finding had already been made in Romania on a reduced number of 15 patients: the total antioxidant capacity of plasma increased in type 2 diabetes (O Saviu et al, J Internat Med Res. 2012, 40, 709-716).
Already in 1999 it had been found that in diabetic condition antioxidant enzyme levels are elevated (M Ramanathan et al., Indian J Exp Biol 1999, 37, 182-3). It is generally accepted that reactive oxygen species and oxidative stress impair beta-cell function in the pancreas and reduce insulin secretion. Pancreatic islets are known for their extremely low antioxidative defense status and their unusual susceptibility to ROS (E Gurgul et al., Diabetes 2004, 53, 2271-78). And any increase in ROS can markedly impair insulin secretion.
Confronted with low levels of insulin, which has anti-inflammatory properties (Aljada et al., Metab Clin Exp 55, 2006, 1177-85), diabetic beta-cells try self-protection against oxidative stress through an adaptive up-regulation of their antioxidant defenses (Gregory Lacraz et al, PLoS ONE 2009, 4-8, e6500). But often this protection is not sufficient and it is often proposed to supplement antioxidants in the treatment of diabetes. A review published in 2011 reviews all human clinical trials where antioxidants were studied as an adjuvant to standard diabetes treatment. The authors came to the surprising conclusion that there is not any established benefit for antioxidants use in the management of diabetic complications. They interfer with the physiologic redox balance. Therefore, routine vitamin of mineral supplementation should not be generally recommended (S Golbidi et al, Curr Diabetes Rev 20117, 106-25).

One of the neglected side effects of diabetes is the generation of large quantities of hydrogen peroxide.
Healthy pancreatic beta-cells exhibit a dramatic response to nutrients and glucose through hypersecretion of insulin in order to maintain energy homeostasis. Many studies have suggested that chronic exposure of pancreatic beta-cells to high levels of glucose may contribute to impaired beta-cell function, leading to the production of ROS. Intriguingly, compared to other tissues, beta-cells have a lower abundance of antioxidant defense such as superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPx). The ROS produced include superoxide and hydroxyl radicals. These may subsequently be converted to hydrogen peroxide (Y Ihara et al., Diabetes. 1999 , 48 927-32). A study from Tunesia shows that the hydrogen peroxide concentration in plasma is increased fourfold in type 2 diabetes compared to controls (A Msolly et al., J Cardiovascul Disease, 2013, 1, 48-51). It was based on 200 confirmed type 2 diabetes patients and 200 controls recruited in the regional blood transfusion center of Sousse. There was a strong positive linear correlation between glycated hemoglobin and hydrogen peroxide, between free fatty acids and hydrogen peroxide concentration and a negative correlation between quantitative insulin index and hydrogen peroxide. Another paper documents that insulin treatment reduces the hydrogen peroxide concentration in blood (A Bravard et al., Am J Physiol Endocrinol Metab, 2011, 300, E581-E591).
At sites of inflammation hydrogen peroxide appears to modulate the inflammatory process and inactivates NFκB. Due to its permeability in many tissues, it operates as an intracellular and intercellular messenger, but at high concentrations it becomes toxic, especially in insulin producing cells known for their extremely low antioxidant equipment against hydrogen peroxide (B Halliwell et al., FEBS Letters 2000, 486, 10-13). Hydrogen peroxide could also plays a role in the regulation of renal function. Renal medullary hydrogen peroxide production is increased in diabetics (D Patinha et al., Life Sci, 2014, 108, 71-9).
Hydrogen peroxide has another side effect. It decreases endothelial nitric oxide synthase NOS promoter activity (Kumar S et al., DNA Cell Biol. 2009, 2:119-29). It is well documented that NOS and NO generated by this enzyme have important endothelial functions and its inhibition may explain most of the cardiovascular problems generated by diabetes.

Several biomarkers of metabolic acidosis, including lower plasma bicarbonate have been associated with insulin in cross-sectional studies. A team from Canada conducted a study called the “Nurses Health Study”. Plasma bicarbonate was measured in 630 women who did not have type 2 diabetes mellitus at the time of the blood draw in 1989 but developed the disease during 10 years of follow-up. The outcome was that higher plasma bicarbonate levels were associated with lower odds of incident type 2 diabetes mellitus (EI Mandel et al., CMAJ, 2012, 184, E179)
The pancreas generates a lot of sodium bicarbonate. The rate of decomposition of hydrogen peroxide has been measured in aqueous sodium carbonate solutions (0.1-1.0 M). It is decomposed in a few minutes and 10 times faster than in a sodium hydroxide solution (HH Lee et al., 2000, Tappi Journal)(U von Gunten et al, Ozone Science and Engineering Journal, 2000, 22, 305-328).
The role of bicarbonate and hydrogen peroxide in diabetes deserve much more research. Because it has an impact on malaria too. As the University of Quds has shown bicarbonate strongly contributes to the inhibition of beta-hematin crystallization (Suhair Jaber et al., J Pharmacy Pharmacol 2015, 3, 63-72).

It is astonishing and frustrating that the impact of diabetes on Plasmodium proliferation is known since more than 100 years. In 1912 Bass and Johnson reported that the addition of glucose was necessary for successful in vitro cultivation of the human malaria parasites Plasmodium falciparum and vivax. However, blood from a diabetic person was successfully used in the culture medium without addition of glucose. They also reported that the amount of quinine sufficient to control malaria infections was ineffective in this case. As a result of these observations they came to the conclusion that the elevated blood sugar from a diabetic patient provided a medium which could better support the growth and reproduction of parasites than could the blood of a non-diabetic person.

Pierre Lutgen
Mutaz Akkawi
17 december 2015.

Does artesunate promote malaria transmission?

décembre 12, 2015

The amino acid arginine is the only molecule in our food known to generate nitric oxide NO via NOS enzymes. It plays a key role in malaria therapy and cerebral malaria as described in previous blogs on http://www.malariaworld.org. NO derived from arginine is not only lethal for merozoites but also for gametocytes. NO is efficient against other diseases like leishmaniasis or filariasis (R O’Connor et al., Infection and Immunity, 2000, 68, 6101-6107).

The inducible nitric oxide synthase (iNOS) is NF-kB-regulated. Many botanical medicinal herbs and drugs derived from these herbs have been shown to have effects on the NO signaling pathway.

Polysaccharides may enhance this production, through a potent macrophage/monocyte activation. This has been demonstrated for acidic polysaccharides from Artemisia tripartita (Gang Xie et al., Photochemistry, 2008, 69, 1359-71), from Tanacetum vulgare (Gang Xie et al., Int Immunopharmacol 2007, 7, 1639-50), from mushrooms (JJ Volman et al., Mol Nutr Food Res, 2010, 54, 268-76). Also for Ginseng saponins and polysaccharides through the activation of NF-kB in macrophages (JY Kim et al., Biosc Biotechnol Biochem. 2005, 69, 891-895). But the polysaccharide with the strongest stimulation of NO synthesis is inulin (HN Koo et al, J Nutr Biochem, 2003, 14, 598-605).) The activation of NF-kB by inulin plays a key role (F Bahmani et al., J Am Coll Nutr, 2015 2, 1-8) and may explain the antimicrobial and tumoricidal activity. This explains why current research is directed to develop inulin as an adjuvant for influenza vaccines, hepatitis B vaccines, malaria vaccines and anti-tumour drugs (D Silva et al., Immunol Cell Biology 2004, 82, 611-16).

On the opposite, vitamins, particularly vitamin E and C inhibit iNOS and are detrimental for this reason during a malaria infection (see « Vitamin C and Malaria : beware ! » on http://www.malariaworld.org Mar 15 2015).

Coumarins, particularly scopoletin, strongly inhibit iNOS (Tien-Ning Chang et al., Evidence Based Complement Alternat Med, 2012, 595603). Its action to this effect is as strong as that of vitamin C (Xiujuan Yao et al., Int Immunopharmac. 2012, 14, 434-462). Quercetin is a NO inhibitor too, but scopoletin is ten times stronger (Aneta Janecki, Thesis, 2012, Universität Berlin). It needs to be mentionned at this stage that all genotypes of Artemisia annua are poor in quercetin (Laboratory Celabor, personal communication). Scopoletin is well present in most species of the Artemisia family. The NO inhibitory effect by scopoletin was demonstrated in Artemisia feddei (TH Kang et al., Planta Med 1999, 65, 400-3). It was even patented for Artemisia annua (US 6337095, 2002). The authors find higher concentrations of scopoletin in the stems than in the leaves (0.3 vs 0.2%). Very important in this context appears the fact that Artemisia annua from Luxembourg genotype only contains 0.02 % of scopoletin versus 0.2 % for Artemisia annua of the high artemisinin hybrid (Rosine Chougouo, Thesis, Université des Montagnes, Cameroon, 2011) and (Laboratory Celabor, personal communication). This to some extent may explain its excellent performance in clinical trials.

But sesquiterpene lactones even appear to be stronger NO inhibitors. This was first evidenced in 1997 for Artemisia ludoviciana in Mexico (P Bork et al., FEBS letters, 1997, 402, 85-90). The sesquiterpene lactones parthenolide and isohelenin prevented NF-kB activation completely as low as 5 microM. A similar effect was confirmed for the sesquiterpene lactone helenalin (G Lyss et al., J Biol Chem. 1998, 273, 33508-16). Ergolide, a sesquiterpene lactone from Inula britannica inhibits iNOS in macrophages (Jeung Whan Han et al., Brit J Pharmacol 2001. 133, 503-12). A similar effect for artemisinin was found in Italy (E Aldieri et al., FEBS letters, 2003, 552, 141-144). A more extensive study on 5 artemisin derivatives at the University of Heidelberg showed that among the five in a mouse macrophage model artesunate revealed the highest ability to inhibit generation of NO (J.Badireenath et al., Nitric Oxide, 2008, 19, 184-191).

In our own research we had found that in vitro pure artemisinin pretreatment strongly inhibited NF-kB activation (Dr Mario Dicato, personal communication 2008). We have performed extensive chemical analysis of the Artemisia annua produced at Luxembourg in comparison with the Mediplant hybrid variety. In all laboratories (Laboratoire National de la Santé in Luxembourg, Celabor in Belgium) we confirmed that Artemisia annua from Luxembourg is poor in artemisinin (0.1%) and scopoletin (0,02%) compared with 1.2% artemisinin and 0.2 % scopoletin in the hybrid. In the froth test the Artemisia from Luxembourg is also much richer in saponin than the Artemisia hybrid from Cameroon. We have confirmed that the Luxembourg genotype has better anti-inflammatory properties i.e.lower IL-6 and IL-8 activation PM de Magalhaes et al., Food Chemistry, 2012, 134, 864-71). In all clinical trials we have run in several African countries with infusion or capsules with Artemisia annua from Luxembourg the cure rate was >95%. In the literature we find two clinical trials with the Anamed high artemisin herb. In both cases the cure rate is much lower, around 75% and the recrudescence is high (CH Blanke, G Naisabhaet al., Tropical Doctor, 208, 38 . 113.6)(MS Müller et al., Trans R Soc Trop Med Hyg 2004, 98, 318-21)

We are currently running clinical trials to study the effect of different Artemisia species on gametocytogenesis.
Besides the inhibition of NO which leeds to the proliferation of gametocytes, artemisinin derivatives are strong immunosuppressors (AF Tawfik et al., Int J Immunopharmacol. 1990, 12 385-389) opening a wide angle door for recrudescence and reinfection.

The result of greed and stubborness : a Pyrrhic victory for Bigpharma and WHO, but a genocide looming for Africa.
Pierre Lutgen
3 december 2015

Herbes médicinales, nitrates et arginine

novembre 28, 2015

Herbes médicinales, nitrates et arginine.


On nous a fait beaucoup peur au cours des dernières décennies avec les eaux potables contenant des nitrates. Serait-ce un mythe, comme d’autres avec lesquels on nous fait peur en cette fin de siècle? Le dossier mérite d’être réouvert. Surtout que les nitrates sont souvent utilisés comme médicaments (comme vasodilateurs, contre les saignements de l’oesophage, lors des accouchements difficiles) ou comme ajouts au dentifrice pour fortifier les dents. Au siècle passé on recommandait des prises de 8 g comme médicament. Le corps humain génère lui-même des nitrates. Le lait maternel est très riche en nitrates (N Hord et al., Breastfeeding Medicine, 6-6, 2011, 393-399


La salive transforme les nitrates en nitrites. Il y a à cela des raisons biologiques : les nitrites forment par acidification des oxydes d’azote qui tuent les germes pathogènes dans la bouche, dans l’oesophage et dans l’estomac. Et certains chercheurs vont jusqu’à recommander une alimentation riche en nitrates. On a en effet découvert aux Etats-Unis qu’une alimentation riche en nitrates réduisait les risques du cancer du larynx et de l’oesophage. Le médecin d’Alfred Nobel administrait déjà de la nitroglycérine à son illustre patient pour soigner son Angina pectoris sans bien savoir pourquoi. L’élucidation du mécanisme vient de valoir le prix Nobel à deux chercheurs américains. Certains sportifs prétendent même que les nitrates augmentent leurs performances (K Landsley et al., Medicine and Science in Sports and Exercice, 2011, 1125-1131).


On vient également de se rendre compte que le monoxyde d’azote (NO) qu’on avait déclaré ennemi numéro un il y quelque temps, exerce un effet bénéfique sur un grand nombre de fonctions corporelles. Un rapport de la Royal Société montre qu’il joue un rôle primordial dans la circulation sanguine et la régulation des activités du cerveau, de l’estomac, des poumons, du foie et autres. Le système immunitaire s’en sert pour combattre les infections virales, bactériennes, ainsi que les tumeurs.


La hantise de la methémoglobine et du cancer


Mais comment en est-on arrivé à proscrire les nitrates?

Les normes très sévères se basent sans doute sur des cas de methémoglobinémie ou d’intoxication constatés chez des nourrissons en Allemagne après la guerre. Il s’agissait dans la plupart des cas connus d’eaux de puits situés près de fosses à purin, riches en composés azotés, mais surtout en contamination bactérienne. Cette hypothèse a été récemment confirmée par des recherches faites aux Etats-Unis Environmental Science and Technology. 367A, September 1999.


Ce sont ces cas de methémoglobinémie chez les nourrissons et des indications que les nitrosamines pouvaient avoir une incidence sur les cancers qui ont motivé en 1961 la FAO (Food and Agriculture Organization) et l’OMS (Organisation Mondiale de la Santé) à établir une valeur pour une dose journalière admissible (DJA) dans l’alimentation. Un coefficient de sécurité fixé arbitrairement a été appliqué pour obtenir une très forte marge de sûreté. En 1980 la directive européenne sur les eaux potables a édicté la règle suivante : l’eau est potable si la concentration en NO3 est inférieure à 50 mg/l. Les normes de la plupart des pays européens ont par la suite adopté la même valeur dogmatique. Aucune de ces normes ne fait référence à des données scientifiques. Il est en effet très difficile, voire impossible, de trouver dans la littérature scientifique des données épidémiologiques sur lesquelles se baserait cette norme ou de trouver des études ayant déterminé des seuils de toxicité pour les humains.

La dose létale est de 1000 mg/kg de poids vif chez les bovins (soit 1 000 000 mg ou un kg  de nitrate pour une grosse vache).


La dose moyenne absorbée par un adulte par jour est de 100 mg par jour. Les légumes y contribuent pour 86% d’après une étude finnoise, et l’eau seulement pour 3%. D’après le magazine Test-Achats (Belgique) n° 376-1995 beaucoup de salades pommées dépassent les valeurs de 3000 mg/kg de nitrates autorisés en été et de 4000 mg/kg autorisés en hiver. Si on a peur des nitrates il faudrait certainement s’abstenir des bettes, des betteraves rouges et des radis qui peuvent contenir jusqu’à 5000 mg/kg.  La laitue dite biologique ne contient guère moins de nitrates que la laitue non-biologique. En mangeant notre assiette de salade ‘biologique’ nous ingérons autant de nitrates qu’en buvant 10 litres d’eau prétendue polluée.


Ils y a des gens qui aiment le jambon et la saucisse. Mais on perturbe leur plaisir avec la phobie des nitrates et nitrites. Ces salaisons en contiennent autour de 100 mg/kg, soit vingt fois moins que la salade biologique.


D’après un rapport publié par le Ministère Fédéral de la Santé de la RFA, aucun cas de methémoglobinémie n’a été constaté au cours des vingt dernières années en Allemagne de l’Ouest, là où l’eau potable est bactériologiquement saine et cela même lorsque la concentration en nitrates dépasse les valeurs admises. Les quatre cas de méthémoglobinémie connus en France entre 1989 et 1992 sont dus à des biberons ou des soupes de carottes contaminés par des bactéries et laissés à  la température ambiante, ce qui favorise la transformation des nitrates en nitrite.


De nombreuses eaux de puits en Californie contiennent plus 2000 mg/l et pourtant aucun cas de méthémoglobinémie n’y a été détecté (M.Apfelbaum, Risques et peurs alimentaires, Editions Odile Jacob, nov 1998).


Concernant les cancers, les expériences animales, même faites à des doses massives n’ont jamais montré d’augmentation de fréquence. Les variations dans le taux de cancer gastrique entre différentes régions d’Italie n’ont pas de corrélation avec une alimentation plus riche en nitrates. La même conclusion a été tirée dans une étude statistique faite dans l’Etat de Wisconsin. Une étude similaire a Taiwan a montré que les cancers du colon sont en relation avec la dureté de l’eau et nullement avec les nitrates(C, Yang et al., Arch. Environm. Contamination and Toxicology, 35, 148, 1998). L’influence des nitrates de l’eau potable sur les tumeurs du cerveau a été étudiée à l’université de Heidelberg. Aucune corrélation n’a pu être trouvée. La plus récente de ces études a porté sur 70 000 personnes à Bocholt BG Volkmer et al., BJU Int 95, 972, 2005.en Allemagne. La majorité avaient consommé pendant 28 années une eau à 60 mg/l de nitrates, les autres à 10 mg/l. 527 cas de cancers rénaux ou urologiques ont été détectés dans cette population, mais on ne peut pas mettre en évidence un effet néfaste significatif pour les nitrates. Pour les nitrosamines tant décriées aucun lien avec le cancer n’a pu être mis en évidence à ce jour (Deutsche Forschungsgemeinschaft, April 15th 2014).




Le secret des herbes médicinales : nitrates et arginine


Les plantes médicinales contiennent en général plus de nitrates que les légumes ou les fruits. On parle de 1 200 mg/kg de matière fraîche contre 400 pour les fruits et légumes.

Les plantes de la famille Artemisia peuvent être considérées comme accumulatrices : riches en potassium gallium, selenium, nitrates (A Traore, M Diallo…P Lutgen. Afric J Biotechnol 12-26, 4179-86, 2013). Elles poussent très bien sur les sols riches en nitrates. On a mesuré des valeurs de 3% de nitrates dans des armoises séchées.

Le rôle important du NO a été reconnu au cours des dernières décennies. Parmi les acides aminés trouvés dans la nature seule l’arginine produit le NO à l’aide des enzymes NOS. On trouve l’arginine surtout dans les poissons, la viande, les noix, l’ail, les avocats. Or il se fait également que les plantes de la famille des Artemisia sont très riches en arginine. Une étude récente d’Ukraine (O Ochkur et al., Pharm Innovat J.  2013, 2, 64-67) a analysé 8 différentes sous-espèces de cette famille et trouvé une teneur de 8 à 10 fois supérieure à celle d’autres herbes ou légumes. Artemisia annua est celle qui en contient le plus. Ceci confirme en fait les données de EA Brisibe and J Ferreira  (Food Chemistry, 2009, 115, 1240-1246).

En résumé, on peut dire qu’il y a deux sources exogènes d’oxydes d’azote: les nitrates et l’arginine provenant de l’alimentation (l’inhalation de NO provenant du trafic routier est négligeable, voire ridicule)

Mais il y a pour les composés nitrés une forte homéostasie. Le NO est très rapidement métabolisé en nitrate et nitrite. La réaction inverse est possible également. Le nitrite est un réservoir pour le NO. Cet équilibre peut être perturbé par la maladie. La concentration est même un signal pathologique. Le stress oxydatif, particulièrement dans le cas du paludisme, inhibe les enzymes NOS et la production de NO à partir d’arginine et les nitrates et nitrites servent de roue de secours dans ce cas. L’existence en parallèle de ces métabolismes est une preuve de l’importance des composés nitrés pour la santé.

Les nitrites comme remède contre le paludisme

Un article scientifique publié il y a 20 ans aurait plus dû attirer l’attention du monde médical dans la lutte contre le paludisme qui tue encore un million de personnes par année, surtout suite au paludisme sévère ou cérébral.  (NM Anstey et al., J Exp Med 1996, 184, 557-567). Dans ce dernier cas il fût démontré qu’en cas de paludisme cérébral la production de NO est inhibée, mais que l’issue fatale peut éventuellement être évitée par une stimulation de la production de NO. L’étude se basait sur des résultats in vivo chez des enfants tanzaniens. Déjà quelques années plus tôt il avait été montré in vitro que le nitrate et le nitrite étaient toxiques pour le Plasmodium à des concentrations millimolaires (KA Rockett et al., Infection and Immunity, 1991, 59, 3280-3283).

Il est fort possible qu’à l’époque ces résultats passaient aux oubliettes parce que la phobie contre les nitrates et les nitrites était des plus virulentes.


Un travail plus récent a montré que le paludisme cérébral chez les rats pouvait être réduit de 79% à 29% par l’administration de NO  et le taux de survie augmenté dans les mêmes proportions. Le traitement conduisait aussi à une meilleure irrigation sanguine du cerveau, une diminution de l’inflammation vasculaire et moins et des risques d’hémorrhagie (P Cabrales et al., J Infect Diseases, 2011,  203, 1454-63). Par son action le NO réduit l’adhésion endothéliale des parasites qui bloquent les vaisseaux, cause principale du coma.

A la Duke University (Zhang et al., poster) on a pu montrer chez des enfants souffrant de paludisme sévère une corrélation inverse entre la sévérité de la maladie et la production de NO à partir d’arginine. Le paludisme sévère se caractérise par des dysfonctions endothéliales. Des essais in vivo en Indonésie ont montré que l’arginine intraveneuse augmente le taux de NO dans l’air expiré et augmente le tonus des artères (TW Yeo et al., JEM, 2007, 11, 2693-2704).

Ce n’est donc pas surprenant que des travaux récents essaient de démontrer que la disponibilité de NO peut être efficace contre le paludisme sévère (P Sobolewski et al., Trends Parasitol, 2005, 21, 414-22). D’autres travaux ont montré que les concentrations en nitrates et nitrites étaient plus faibles en cas de paludisme cérébral. L’hypoarginemia est souvent la cause d’une faible disponibilité de composés nitrés (I Gramaglia et al., Nature Medicine, 2006, 12, 1417-22). On a mesuré la concentration d’arginine dans des échantillons de plasma cryopréservés d’enfants tanzaniens et trouvé des valeurs basses chez les patients souffrant de paludisme cérébral, intermédiaire en cas de paludisme moins sévère et des valeurs normales, donc plus élevées chez les sujets sains (Lopansri et al, The Lancet, 2003 361, 676-78).

Le NO intervient dans la destruction des parasites asexués à l’aide des globules blancs. Très encourageante est également la découverte par les mêmes auteurs que le NO inhibe les gamétocytes (T Naotunne et al., Immunology, 1993, 78, 555-562). Plus surprenante encore est la découverte très récente d’une équipe chinoise (Li Zheng et al., Parasites & Vectors, 2015, 8 :326) montrant que des suppléments d’arginine non seulement réduisent la parasitémie mais réduisent également le nombre de zygotes et d’oocystes chez les Anophèles femelles ayant ingéré du sang humain chargé d’arginine. Ceci confirme d’autres résultats chinois. Le NO inhibe la transformation des gamétocytes de Plasmodium yoelii en gamètes. (Ya-Ming Cao et al., Parasitol Internat. 1998, 47, 157-166).  C’est en fait l’exflagellation des gamétocytes qui est inhibée (YJ Liu et al., Zhongguo Ji She, 2007, 25, 206-8).


Dans la lutte contre le paludisme les sociétés pharmaceutiques se sont surtout attelées à l’élimination des parasites de la phase asexuée (mérozoïtes, trophozoïtes, schizontes) et des symptômes qu’ils causent (fièvre, nausée, inflammation, coma…). La lutte contre les gamétocytes de la phase sexuée a été négligée. On a cependant pu observer que la primaquine avait un effet gamétocytocide occasionnel mais ce médicament peut être fatal pour des gens souffrant de la déficience G6DP. La sulfadoxine-pyriméthamine et la docycline augmentent même la population des gamétocytes La chloroquine et l’artemisinine n’ont pas d’effet sur les gamétocytes murs. En fait ces remèdes n’ont aucune chance d’affecter les gamétocytes, parce qu’ils sont administrés durant 3 jours seulement et les gamétocytes se développent à partir du 10éme jour d’une infection.


L’arginine joue un rôle crucial dans l’activation des macrophages et influence favorablement d’autres mécanismes de défense du système immunitaire tels que les cellules B et T. Elle augmente les CD4. Chez les sujets infectés par la leishmaniase le parasite désactive les macrophages. On a pu démontrer in vivo le rôle essentiel joué par l’arginine dans la destruction des parasites de cette maladie. (N Wanasen, Immunol Res 2008, 41, 15-25). Le NO détruit également les trophozoïtes de Giardia lamblia. Le NO peut détruire les mycobactéries de la tuberculose. (Th Schön, Thesis, Linköping University, 2002, No 749).  En cas de schistosomiase (bilharziose) on a observé de basses teneurs en arginine dans le serum (A Senft, Comparat Biochem and Physiol, 1967, 21, 299-306).


Les activités prophylactiques de la tisane Artemisia annua contre le paludisme ont été clairement démontrées dans des essais in vivo en Ouganda (PE Ogwang et al., Trop J Pharmaceut Res « 012, 13, 445-453).








Le nitrite qui était déclaré ennemi public commence à être innocenté, ou mieux, il retrouve son rôle essentiel pour la santé. Il y a même des auteurs qui disent que beaucoup de maladies des dernières décennies sont dues à ces restrictions légales absurdes (NS Bryan., et al Free Radical Biology & Medicine 2006, 41, 691-701).


Il y a eu dissociation entre la réalité telle que décrite par la science et les fantasmes collectifs créateurs d’une autre réalité, sociale puis politique. Pourquoi nous a-t-on fait peur ?


Il est évident que plus de recherche est nécessaire pour mieux comprendre le mécanisme d’action thérapeutique et prophylactique des produits nitrés. Et il est urgent de procéder à des essais cliniques avec Artemisia annua. Malheureusement ces essais sont formellement interdits par l’OMS.


Pierre Lutgen

Lucile Cornet-Vernet