Antimicrobial and immunomodulatory properties of extracts of Asparagus setaceous Kunth and Caesalpinia volkensii Harm

Abstract

Infections due to antibiotic resistant microorganisms have become widespread in recent years, and resistance rates among key pathogens have continued to grow at an alarming rate worldwide and the search for novel antimicrobial agents to combat such pathogens have become crucial. It is now an accepted that majority of the antibiotics used in the treatment of these infectious diseases have failed and the ever-increasing resistance to current anti-infective drugs has become a major concern to the medical community. The problem of microbial resistance is still growing and the continued use of antimicrobial drugs in the future remains uncertain. Therefore, actions must be taken to reduce this problem, for example, to control the use of antibiotic, and to continue to develop new novel drugs, either synthetic or natural. The ultimate goal is to offer appropriate and efficient antimicrobial drugs to the patient. For a long period, plants have been a valuable source of natural product for maintaining human health and an impressive number of modern drugs have been isolated from them, many based on their use in traditional medicine. Currently, it is estimated that over 50 percent of all modern clinical drugs are of natural products origin. This study investigated the mode of action of two Kenyan plants namely Asparagus setaceous Kunth and Caesalpinia volkensii Harm. Asparagus setaceous belong to the family Liliaceae while Caesalpinia volkensii belongs to Caesalpinacea. Many medicinal uses of the various parts of plants from these two families have been reported in traditional folklore medicines. These have reportedly been used in the treatment of inflammatory diseases, bronchitis, pneumonia, syphilis and other venereal diseases, malaria and antihelminthic. However, their therapeutic mechanisms remain largely unclear. It is not clear how extracts of these two plants exert their beneficial and therapeutic effects. It is believed that their effects could be direct on the pathogens thus killing them, or to alter and enhance the functioning and activities of immune cells. There was need to investigate and clearly elucidate the mechanisms of actions of these two plants. The aerial part and leaf of A. setaceous and leaf, stem and root of C. volkensii were extracted using polar and non polar solvents. The dichloromethane, hexane, methanol, ethanol and aqueous extracts of A. setaceous root and aerial part and C .volkensii leaf, stem and root were evaluated for their antibacterial and antifungal properties against gram negative (Escherichia coli, Pseudomonas aeruginosa and Streptococcus faecalis) and gram positive (Bacillus subtilis, Staphylococcus aureus) and the fungus Candida albicans. Antibacterial and antifungal activity was tested using the test disc diffusion method. Minimum inhibitory concentration (MIC) of the extracts was also determined. Extracts found to be active were also tested on their ability to affect the growth kinetics of selected pathogens.

An assessment to evaluate the ability of the crude extract to affect functioning and activities of immune cells was also carried out. Extracts were administered orally and then the crude extracts were evaluated on their effect on total and differential white albino rats. The crude extracts were also evaluated on their ability to activate and enhance the function of neutrophils and macrophages. This was done through the neutrophil adherence, Candida and sheep red blood cell phagocytosis and nitroblue tetrazolium reduction tests (NBT). They were further assessed on their capacity to stimulate cellular mediated immunity through delayed type hypersensitivity (DTH) test. Dose dependent relationship tests were also carried out in order to establish the most active dosage. Data obtained from this study was analyzed using ANOVA and student t-test with the level of significance fixed at p<0.05. The organic extracts of A. setaceous and C. volkensii were demonstrated to possess activity against some of the bacteria and fungus. The aqueous extracts were largely inactive. The extracts showed minimum inhibitory concentration (MIC) activity against some of the bacteria and the fungus with MIC ranging from 6.25mg-50mg/kg. The extracts were also able to affect the growth kinetics of the selected pathogens when compared to pathogens growing in presence of media alone. In immunomodulatory tests, the methanol, ethanol and aqueous extracts were found to be active. They were able to significantly increase total white blood cells count as well as neutrophil numbers in albino rats (p<0.05). They were found to significantly increase WBCs counts in rats 9 days after cyclophosphamide treatment (p=0.000). Rats pretreated with the extracts were found to be protected from the myelosuppressive effect of cyclophosphamide. The extracts significantly enhanced ercent neutrophil adhesion, candida phagocytosis and increased nitroblue tetrazolium reduction (p= 0.000) Sheep red blood phagocytosis by macrophage as determined spectrophotometrically was found to be enhanced. The methanol, ethanol and aqueous extracts were also found to significantly (p=0.000) enhance DTH response in rats to variable degree. The activities of the extracts were also found to be dose dependent with the dose of 500mg/kg and 1000mg/kg being the most active. In most cases, the two higher doses used had similar and comparable activity. The extracts were also demonstrated to increase the percent survival rates of rats exposed to S. aureus. Phytochemical screening revealed the presence of alkaloids, flavonoids, steroids, sterols, saponins and tannins. This study goes along way in confirming and scientifically verifying and validating claims by traditional medical practitioners that these two plants have therapeutic values in managing infectious diseases. The study indicates that the main mechanism of action of the extracts of these plants is immunomodulation although further research is required to make definite conclusion. This will accelerate the integration of these medicinal plants into our health care system and of their use in the management of infectious diseases. Again the importance of demonstrating presence of immunostimulants from these plants portends a great conquest for medicine especially in this era of HIV/AIDS.