Phytochemicals from Beilschmiedia anacardioides and Their Biological Significance

Medicinal plants provide a vast array of raw materials for primary health care in Africa and other countries of the world. The World Health Organization (W.H.O) estimates that about 80% of Africans living in the continent have resort to traditional medical practitioners and the use of traditional medicine for the treatment of their diverse ailments. This practice has a considerable importance within the economic and cultural milieu of Africa.


Introduction
Medicinal plants provide a vast array of raw materials for primary health care in Africa and other countries of the world.The World Health Organization (W.H.O) estimates that about 80% of Africans living in the continent have resort to traditional medical practitioners and the use of traditional medicine for the treatment of their diverse ailments.This practice has a considerable importance within the economic and cultural milieu of Africa.
It is estimated that less than 10% of the world's genetic resources have been studied seriously as sources of medicines.Yet from this small fraction, humanity has reaped enormous benefits.
The search for bioactive plant natural products from higher plants is gathering momentum, as they have potential to provide new lead compounds or to be of use directly.There is an increasing sense of urgency about this search due to the destruction of natural resources.With regards to these plants, it has been estimated that 25-30 million hectares of the world's rainforests are lost each year.The crucial problem already expressed by several scientists then is how to search efficiently and rapidly for bioactive components from the vast number of unstudied plants.Part of the solution is to narrow down the search-selection.
Six approaches to the selection of plant materials for study exist: the locally random, the taxonomic, the ethnobotanic, the phytochemical, the information based, and serendipity.The ethnomedical approach appears to be the method of choice for natural product chemists working in Africa and other developing countries.In this method only plants used in traditional medicine are collected.
Very little attention has been paid to Beilshmiedia species.Previous studies concern trees and herbs of Beilshmiedia species, with the aim of cultivating herbs containing the same endiandric acid derivatives as trees.Other studies led to a patent on interesting synthesis of endiandric acid derivatives.
In our own search for prospective pharmacological products from ethnobotanic data, we have been looking at some traditional medicines whose therapeutic efficiency is scientifically established towards biomedical analyses of patients on treatment in a specialized clinic.We have selected a traditional medicine based on one plant Beilshmiedia anacardioides (Lauraceae), for its proven efficiency on genital infections and rheumatisms through clinical research.No phytochemical studies of Beilshmiedia anacardioides are however to our knowledge available in the literature.We propose that phytochemists looking for novel bioactive natural products should investigate the medicinal plants whose therapeutic efficiency has been established through clinical research on African medicine.
We have initiated a systematic phytochemical investigation of the extracts of Beilshmiedia anacardioides as well as the antibacterial activity of the eight new compounds isolated, towards five strains of microbes, namely Bacillus subtilis, Micrococcus luteus, Streptococcus faecalis, Pseudomonas palida, and Escherichia coli.
The methods used for the isolation of the compounds were mainly column chromatography and preparative TLC.The structures of all compounds were elucidated by means of modern spectroscopic techniques such as 1D-NMR ( 1 H-NMR, 13 C-NMR with DEPT experiments), and 2D-NMR ( 1 H-1 H-COSY, HMQC,HMBC,NOESY), MS , IR and X-Ray spectroscopies.
The antibacterial activities of the new compounds were examined using the dilution technique with respect to the zone of inhibition (ZI) and minimum inhibitory concentration (MIC).
We report here the results we have so far obtained and published in three renowned scientific journals (Chouna et al.,2009;2010;2011).

Study of the ethnomedical preparation
The ethnomedical preparation is a decoction.The decoction is prepared as follows: Boil about 80 g dry stem bark powder in 3 litres of water for 15 minutes.Filter when lukewarm.Drink a glass twice daily for ten days.
A treatment for fibromes could last about two to three months, depending on the patient's age.

Study setting
Cameroon is a bridge between Central Africa and West Africa, humid Africa and dry sahelian Africa, French speaking and English speaking Africa ( French and English are official languages).The country is open to the Gulf of Guinea in his south-west border.Lake Chad is at the extreme -North border.A country of 475.442 square kilometers, Cameroon is borded in the west by Nigeria, on the east by Chad and the Central African Republic, and on the south by Congo, Gabon, and Equatorial Guinea.
The Bamoun are a Bantu people living in the west Region of Cameroon.They number more than half a million.They have a rich cultural Heritage, including famous traditional Healers.Sultan Njoya wrote a book on Bamoun traditional medicine.Important Bamoun towns are Foumban, Foumbot, Koutaba, Massangam, Magba, Malantouen.Among important villages are Mahoua, Manki 1 and Manki 2, where the plant Beilshmiedia anacardioides was collected.

Generalities on Beilschmiedia anacardioides
B. anacardioides is found in Central Africa, especially in Cameroon, Tchad and Gabon.In Cameroon, this species is found in the Adamaoua and the West Region (Eyog et al., 2006;Fouilloy, 1974).It is synonymous with B. ngriki and B. Jacques-felixii and it is commonly named ntseum (in Bamoun language) in the Noun subdivision of the West region of Cameroon (Eyog et al., 2006;Fouilloy, 1974;Tchouala, 2001).

Uses of Beilschmiedia species in traditional medicine
B. anacardioides stem bark is used in the Noun sub-division of the West Region of Cameroon to treat uterine tumours, rubella, rheumatisms, bacterial and fungal infections (Tchoula, 2001).Seeds are used as spices (Eyog et al., 2006).B. lancilimba is used in the same region to cure skin bacterial infections (Tchouala, 2001).B. manii is used to treat dysentery and headache.It is also used as an appetite stimulant (Iwu, 1993).

Phytochemistry of plant constituents of Beilshmiedia species
A review of the literature revealed that no phytochemial studies have been carried out on Beilshmiedia anacardioides prior to the initiation of our study.The various phytochemical and pharmacological studies performed and reported in the literature on the beilshmiedia genus are discussed below.

Alkaloids isolated from the Beilschmiedia genus
Very few alkaloids have been isolated from the Beilschmiedia genus.Table 1.Structure of some alkaloids isolated from the Beilschmiedia genus www.intechopen.com

Pharmacological importance of alkaloids isolated from the Beilschmiedia genus
A bisbenzylisoquinoline alkaloid dehatrine ( 9) isolated from the wood of B. madang, exhibited potent inhibitory activity (IC 50 value of 0.017 µM) against the proliferation of the malaria pathogen P. falciparum (Kitagawa et al., 1993).Paulo and coworkers (1992) demonstrated the antimicrobial properties of laurelliptine (10).Leaf of B. volckii (Banfield et al., 1994) Table 2. Structure of some lignans and neolignans isolated from Beilschmiedia genus

Pharmacological importance of the endiandric acids
Very few pharmacological studies have been done in this class of metabolites.Endiandric acid H ( 41) is used for the manufacture of medication, in particular for the treatment of asthmatic disorders or concomitant inflammatory symptoms of asthma (Eder et al., 2004).

Results of our own studies
We have initiated a systematic phytochemical investigation of the extracts of Beilshmiedia anacardioides and have so far obtained the following results which have led to three publications in renowned scientific journals (Chouna et al.,2009;2010;2011).
Air-dried and ground stem bark of B. anacardioides was extracted successively at room temperature with MeOH.The methanol extract was re-extracted in turn with CH 2 Cl 2 and EtOAc.These extracts were concentrated to dryness under reduced pressure.

Biological activity and the significance of some compounds
Our preliminary antibacterial studies on the new endiandric acid derivatives have yielded chemical entities that have been shown to possess significant activities (Chouna et al., 2009) .

Antibacterial assay on some compounds isolated from B.anacardioides
Compounds 1-8 were tested in vitro for their antibacterial activity against Bacillus subtilis, Streptococcus ferus, Streptococcus minor, Micrococcus luteus, Escherichia coli, and Pseudomonas agarici, using the dilution technique.
The ZI (Table 1) and MIC (Table 2) obtained for these compounds indicated that they possessed strong to weak antibacterial activity against gram positive bacteria.
Beilshmiedic acid C (3) demonstrated the best potency against B. subtilis and M. luteus, compared to the reference drug ampicillin.The MIC values (Table 2) of Beilshmiedic acids B( 2), C (3) and G( 7), against B. subtilis and Beilshmiedic acid C (3) against M. luteus were found to be greater than that of standard drug ampicillin, indicating that this series of compounds might be possible candidates as antibacterial drugs.
None of the tested compounds was active against Gram negative P. palida and E. Coli.Therefore, they might be well tolerated as antibiotics even for long term treatments.Based on the skeletal features, it is difficult at this stage to define the contribution of the different functional groups with respect to the activity.The mechanism of action of this class of metabolites on these strains is not yet known.Further investigations will help to establish the mode of action of this particular skeleton.These interesting results highlight the potency of this rare class of metabolites that might be investigated for the search of new antibacterial drugs.

Conclusion
In our hypothesis we proposed that phytochemists looking for novel bioactive natural products should investigate the medicinal plants whose therapeutic efficiency has been established through clinical research on African medicine.
We suggested that Natural products from Beilshmiedia anacardioides may play a role in treating genital infections due to B-subtilis and M. luteus, and rheumatisms due Streptococcus ferus and S. minor.The biological activities of some of the constituents isolated in our studies, Beilshmiedic acid C presented above, more than lend support to this suggestion.
It is certain that as more and more data become available from the phytochemical and biological analysis of the constituents of therapeutic efficient medicinal plants selected after clinical research, the role of these plants in the treatment of diseases will become more defined.Thus African Traditional medicine will gain universal status.

Acknowledgments
Financial support for this survey from the Alango Foundation (Centre de Phytomédecine Africaine ) at Dschang / Cameroon is gratefully acknowledged.

4. 2
Phenolic and phenolic derived compounds 4.2.1 Lignans and neolignans Lignans and neolignans and flavonoids are the main phenolic compounds encountered in the Beilschmiedia genus.
Beilshmiedic acid C (3) was more active than Beilshmiedic acid D (4) .The enhanced activity may be due to the additional hydroxyl group at C-4 position in Beilshmiedic acid C (3) .Beilshmiedic acid B(2) which possesses one hydroxyl group more than Beilshmiedic acid A (1) and Beilshmiedic acid C (3) was less active.Beilshmiedic acid A (1) was more active than Beilshmiedic acid C (3) .They are epimers at C-4 position; the modification of the configuration at this position influences significantly the activity.