Antimicrobial structure activity relationship of five anthraquinones of emodine type isolated from Vismia laurentii

Plants belonging to Vismia genus have been studied since 1979 [1] because of their biological activity due to secondary metabolites that they contain. Different parts of Vismia laurentii are used in the traditional pharmacopoeia in the treatment of different affections including microbial infections [2]. Previous chemical assessments carried out on this plant have resulted in the isolation and characterization of a great number of secondary metabolites, the most significant belonging to the xanthone, benzophenone and quinone classes. Quinones especially anthraquinones, present numerous biological activities such as antiprotozoa [1, 3], antituberculous, fungicidal [4], antioxidant [5], cytotoxic and antitumor activities [6]. Xanthones and anthraquinones are known to bind irreversibly with nucleophilic amino acids in proteins, often leading to the inactivation of proteins and loss of function [7]. The rarity of plant diseases in V. laurentii is explained by the development of a natural defense system resulting in the synthesis of a multitude of antimicrobial molecules, which enable them to fight effectively against the pathogenic microbes [8, 9].

Anthraquinones are divided into two types: alizarin and emodine [10]. The alizarin type is used as natural dye in the textile industry [11], while the emodine type was formerly used as a like laxative compound [12–14]. Many studies have reported on antimicrobial activity of anthraquinone compounds of the emodine type [2, 15, 16]. Their variety and the continuuos discovery of new emodine derivate molecules always call in question, the specific properties of the most antimicrobial effective compounds. Moreover, the number of molecules extracted from the biological and/or potentially existing systems is by far higher than the capacity of analysis of their biological properties.

Facing these limitations, a solution consists of building models which allows for correlating the activity to structure within a family of compounds, hereby increasing the effectiveness of high throughput screening [17]. On the basis of their physical and chemical properties, the antimicrobial activity of natural substances can be predicted in order to have information on biomechanism, gain time of bio-prospection of new molecules and to study their use in the sectors of the production of antiseptics, disinfectants and drugs [18, 19]. The structure activity relationship (SAR) or the quantitative structure activity relationship (QSAR) offers approaches which could be useful to predict these antimicrobial activities according to the physical and chemical properties of the molecules concerned. This approach could constitute a first stage of molecules screening and thus making it possible to reduce the number of compounds to be tested in the laboratory.

The purpose of this study was hence to establish a relationship between antimicrobial activities and physicochemical properties of some anthraquinone molecules of emodine type isolated from Vismia laurentii.

The differences observed during the growth kinetics at pH5 and 7 of the tested strains can be associated to the different nature of their cell walls. In fact, the effect of pH that is mostly described on the cell internal pH [27] is most important on bacteria than on fungi. Moreover, homeostasis regulations that sometimes involve ATP dependent processes may also include cell membrane lipid modifications to reduce fluidity [28]. Fungi resistance to pH also depends on their high cell wall thickness and composition mainly made of 80–90% glucomannoproteines, glucanes and chitins; this last compound being higher in Candida albicans with respect to other yeast species [29].

The emodine derived compounds tested in this work were highly colored and hence permitted only evaluation of the MBC and MFC. The microbiocidal concentrations observed for the different compounds can be explained by the compound interference with the cell wall, the membrane, nucleic acid and enzymes [30, 31]. The presence of an external membrane on the Gram negative bacteria can explain the difference of sensibility observed between the two groups of bacteria. Kosanić and Ranković [32] suggested that the cell wall structure and composition of bacteria and fungi could account for the different sensitivity to antimicrobial compounds. On the other hand, Gram positive bacteria and Candida albicans cells have their cell walls exposed, and compounds that can interact with these cell walls should have a long aliphatic chain to help disorder the cell wall. This is the case of compound A compared to compounds C and D. Sikkema et al. [33] observed that saturated alcanes had very low antimicrobial activity, while [34] have demonstrated that unsaturated aldehydes had more antimicrobial activity than saturated ones. This explains the difference between compounds D and B. Moreover, another possible mechanism was proposed by [35] who demonstrated that the antimicrobial activity of emodine on Helicobacter pylori was also due to the interference with saturated and unsaturated fatty acid elongation by inhibiting the β-hydroxyacyl-Acp dehydratase (HpFabZ).

Regarding the physicochemical properties and their relation with the antimicrobial activity of the compounds tested, different methods as described before were used to assess them. The molecular weight (MW) which is an indication of a compound’s steric effect is also one of the key parameters of compounds with pharmacological properties (Lipinski et al., [23]). This author also proposes the ideal molecular weight to be lower than 500 g. The numbers of electron donors (HD) and acceptors (HA) indicate the capacity of a compound to form hydrogen bonds with the cell membrane compounds. Moreover, the partition coefficient (Log K_o/w) gives information on the lipophilicity of the compound and hence its capacity to distribute itself in the membrane lipidic phase and the aqueous environmental or cytoplasmic phase. Regarding the degree of polarization, it is the tendency of a compound to negatively charge itself and hence be easily modified in the presence of positively charged compounds. The surface tension on the other hand represents the internal attractive forces of a compound that limits bond creation with other compounds. The QSAR obtained for Candida albicans indicated that HA and Polarz were the common significant variables of the equations for each pH. Both physicochemical parameters are associated to the compound capacity to bind to other molecules by forming bonds with positively charged atoms. In fact the growing importance of HA and Polarz in the equations as pH decreases (protons concentration increase) in the environment confirm this. At pH7, the concentration of protons is lower and hence the increase of the Polarz negatively affects the fungicidal property. The fact that the antifungal property of the compounds depended on the pH, and that the pH did not affect the growth of Candida albicans may suggest that the effect of pH on the compounds antimicrobial effect is associated with compound modification.

The logical structure activity relationship deduced for the bacteria strains tested indicates that the substitution in position 2 of the emodine compound is detrimental for the antibacterial activity of these compounds while the insaturation of the substitute isimportant for this activity. Moreover the increase of the aliphatic chain length of the methoxy substitute in position 3 increases the lipophilicity of the compound. The antimicrobial property which is increased by the lipophilicity of the compound is reduced as the compound molecular weight increases. In fact the lipophilicity associated to the Log (Ko/w) denotes the capacity to integrate the cell wall and membrane capacity which may be slowed by the steric effect of the compound, revealed here by the molecular weight.

In conclusion, the present work has demonstrated the antifungal and antibacterial properties of some anthraquinones of emodine type isolated from Vismia laurentii. This antimicrobial property is increased by the presence of a long aliphatic chain methoxy group substituted in position 2 of the emodine structure. The mathematical equations produced demonstrate that QSAR can contributeto understanding the diversity of compound antimicrobial activity present in plant extracts.