Original article
Synthesis and biological evaluation of novel ocotillol-type triterpenoid derivatives as antibacterial agents

https://doi.org/10.1016/j.ejmech.2013.07.041Get rights and content

Highlights

  • A series of ocotillol-type derivatives derived from natural PPD were obtained.

  • Compounds 3, 5, 16 and 24 showed potent activities against Gram-(+) bacteria.

  • Compounds 3 and 16 highly sensitized Gram-(+) bacteria to antibiotics KAN and CHL.

  • Compounds 3, 5 and 16 exhibited no cytotoxicity at their MICs.

Abstract

A novel class of ocotillol-type triterpenoid derivatives have been synthesized and evaluated for their in vitro antibacterial activity against several representative pathogenic bacterial strains. Compounds 20(S)-protopanaxadiol (PPD), 3, 5, 16 and 24 exhibited potent antibacterial activity against Gram-positive bacteria. Compounds 3 and 5 also displayed promising antibacterial activity against a community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA; strain USA300). Furthermore, compounds PPD, 3 and 16 combined with two commercially available antibiotics kanamycin and chloramphenicol showed strong synergistic inhibitory effects at their sub-MIC concentrations against S. aureus USA300 and Bacillus subtilis 168. Additionally, cytotoxic activity assay showed that the compounds tested did not affect cell viability of the human epithelial kidney (HEK-293) and human cervical (HeLa) cells at their MICs.

Graphical abstract

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Compounds 3 and 16 combined with kanamycin and chloramphenicol showed strong synergistic inhibitory effects at their sub-MIC concentrations against Staphylococcus aureus USA300 and Bacillus subtilis 168.

Introduction

Since the discovery of Penicillin, the development of antibiotics has been carried out for over 80 years. In addition to the β-lactam family antibiotics, that have many structural variants, other antibiotics or synthetic antimicrobial agents have also been developed, including the macrolides, tetracyclines, quinolones and glycopeptides which have different mechanisms of antibacterial action [1]. However, due to horizontal gene transfer and unlinked point mutations, more and more bacteria with resistance to antibiotics were isolated, with methicillin-resistant Staphylococcus aureus (MRSA) infections representing a particularly serious health problem worldwide [2]. More recently, infections due to community-associated (CA) MRSA, which is responsible for diseases such as necrotizing pneumonia, severe sepsis and necrotizing fasciitis, have risen rapidly, sparking fears of an epidemic [3]. In recent decades, vancomycin-resistant S. aureus (VRSA) strains were also identified [4]. Consequently, novel antimicrobial agents with distinct structures are urgently needed to help battle emerging antibiotic-resistant bacterial infections.

Natural products have been the single most productive source of leads for the development of drugs particularly as anti-cancer agents and anti-infectives [5]. The triterpenoids, including sterols, steroids, and saponins, are a large and structurally diverse group of natural products with well-characterized biological activities [6]. Many of them such as ursolic acid (UA), oleanolic acid (OA), betulinic acid (BA) (Fig. 1) show antimicrobial activities [7]. Previous reports indicated that the weak antimicrobial activity of betulinic acid as compared to its analogs ursolic acid and oleanolic acid is probably due to lack of the E ring that is present oleanene [8]. However, surprisingly, 23-hydroxybetulinic acid (HBA) (Fig. 1) demonstrates moderate antimicrobial activity against Gram-positive bacteria Bacillus subtilis (closely related to Bacillus anthracis) strain 168 and S. aureus strain RN4220 in our preliminary testing (Table 1), which suggests the E ring of oleanene/oleanane in triterpenoid natural products may not be the unique critical factor for antimicrobial activity.

Although the antibacterial activity of some natural products may be less than that of certain marketed drugs, their lower cytotoxicity compared to the traditional antibiotics was preferred. In addition, natural products usually have an element of structural complexity which is required for the inhibition of many antibacterial protein targets [9]. Ocotillol (Fig. 1), a triterpene isolated from Fouquieria splendens Engelm, is characterized by bearing a five-membered epoxy ring at C-20 [10]. Pseudo-ginsenoside-F11 (Fig. 1) which is an ocotillol-type ginsenoside found in Panax quinquefolium (American ginseng) [11] has been proven to possess a protective effect on myocardial ischemia injury [12]. Recent research showed plant derived triterpenoid saponins commonly involve in plant defense system by perturbating membranes of bacteria or virus [13], as well as increasing membrane permeability of mammalian cells [14]. Thus, triterpenoid derivatives are potential to be developed as antimicrobial agents by affecting bacterial cell viability through membrane destruction. So far few studies have been devoted to the antimicrobial effect of ocotillol-type triterpenoid derivatives; furthermore, compared to oleanane, tetracyclic triterpenoid compounds are more potent in drug discovery for their low cytotoxicity [15] and diverse functional groups as well as their prospects for modification. Therefore, the present study described the synthesis of the tetracyclic triterpenoid derivatives from 20(S)-protopanaxadiol (PPD) and evaluated for their in vitro antibacterial, synergistic antibacterial activities and cytotoxicity [16].

Section snippets

Synthetic chemistry

We chose PPD, one of the main ginsengenin components of Panax ginseng, as our starting material (Scheme 1), as it can also be isolated from other natural sources [17]. As previously described [16], the 3, 12-diol of PPD was protected using acetic anhydride in the presence of DMAP; epoxidation of the resulting acetate 1 followed by nucleophilic addition in situ furnished a mixture of tetrahydrofuran 2. After base treatment, two stereoisomeric triols 3 and 4 were obtained with desired yields.

Results and discussion

We have designed and synthesized a series of ocotillol-type triterpenoid derivatives based on PPD. The minimum inhibitory concentration (MIC) of the synthesized compounds was determined against several bacterial strains using a standard LB medium dilution technique. Initial MIC screening results are presented in Table 1. The data demonstrated that PPD and the ocotillol-type triterpenoid derivatives 3, 5, 6, 16 and 24 inhibited the growth of Gram-positive bacteria in vitro with MIC values about

Conclusions

In this paper we have designed and synthesized a series of ocotillol-type triterpenoid derivatives based on PPD, in which compounds 3, 5, 16 and 24 showed good antibacterial activity particularly against Gram-positive bacteria, and compounds 14 and 17 exhibited notably activity against Gram-negative bacteria. Additional testing against CA-MRSA strain USA300 demonstrated compounds 3 and 5 also possess potent antibacterial activity. The subsequent synergistic antibacterial assay revealed that the

Chemistry

Most chemicals and solvents were analytical grade and, when necessary, were purified and dried with standard methods. Melting points were taken on an XT-4 micro melting point apparatus and uncorrected. 1H NMR spectra were recorded with a Bruker AV-300 or ACF 500 spectrometer in the indicated solvents (TMS as internal standard): the values of the chemical shifts are expressed in δ values (ppm) and the coupling constants (J) in Hz. High-resolution mass spectra were recorded using an Agilent QTOF

Acknowledgments

The authors are grateful to National Natural Science Foundation of China (No. 81001358), Promotive research fund for Excellent Young and Middle-aged Scientists of Shandong Province (No. BS2010YY073), Shandong Provincial Natural Science Foundation, China (No. ZR2012HM036), Open Project Program of State Key Laboratory of Natural Medicines, China Pharmaceutical University (No. 201201), Taishan Scholar Project to Fenghua Fu and Australian NHMRC grant APP1008014 for financial support.

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    These authors contributed equally to this work.

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