In Vitro And In Vivo Studies Of Cassia Surattensis Flower Against Aspergillus Niger
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Date
2012-01
Authors
Vello, Sumathy
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Abstract
Infectious disease is one of the commonest health problems in developing
countries with the rise of antibiotic resistance. Invasive aspergillosis is causing high
mortality and morbidity rate among immunosuppressed patients. Therefore, there is
an urgent need for novel antifungal therapy to control the fatality rate in the
population. Opting on medicinal plants has become the current trend amongst
scientific investigators as plants are rich with biological activities. Cassia surattensis
flower was studied to identify this organ as a potential antifungal agent. Screening of
selected pathogenic microorganisms which were inclusive of Gram positive bacteria,
Bacillus subtilis, Bacillus thuringiensis, Micrococcus sp and Staphylococcus aureus,
Gram negative bacteria, Escherichia coli, Proteus mirabilis and Salmonella sp and
fungi Candida albicans, Aspergillus niger and Rhizopus sp against the flower
extract revealed this flower to possess antimicrobial properties. The zone of
inhibition for microorganisms from disc diffusion assay ranged from 13 ± 0.04 mm
to 18 ± 0.00 mm. Microorganisms that were weak against the flower extract were
further evaluated with Minimum Inhibitory Concentration (MIC) assay to determine
the flower extract activity against the microorganisms in a dose dependent manner.
MIC values for both Gram positive and Gram negative bacteria ranged from 3.125
mg/mL to 25.00 mg/mL. For fungi, the MIC value ranged from 3.125 mg/mL to
6.25 mg/mL. The presence of antifungal agent in the flower extract caused damage
to the hyphae and conidiophores of A. niger. This was observed via ultrastructural examinations on A. niger which demonstrated folded, collapsed, squashed and
broken hyphae. Shrunken conidiospores were the major structural alteration detected
on the conidia. Qualitative phytochemical screening of the methanolic flower extract
found that carbohydrates, tannins, saponins, steroids, phenols and flavonoids were
present as active compounds. Toxicity study via brine shrimp assay was performed
to ensure the safety of the flower extract. Results turned out to be negative for
toxicity with LC50 3.32 mg/mL for flower extract and LC50 0.27 mg/mL for
potassium dichromate used as positive control. In vivo oral acute toxicity study was
performed to support the nontoxic data. Mice administered with single dose of 5000
mg/mL of the flower extract did not show any architectural alterations on the lung,
heart, liver, spleen and kidney sections in the histopathology examinations and thus
strongly suggested the flower extract as toxic free. Antifungal activity of C.
surattensis flower against A. niger was further studied with systemic aspergillosis
model. Mice succumbed with conidia was studied via organ fungal burden
determination, galactomannan levels and histopathology analysis. Colony counts in
the organs and Galactomannan Index (GMI) increased steadily in the negative
control as the study prolonged. Mice treated with flower extract showed a reduction
pattern in fungal burden and GMI after Day 7 onwards although the reduction size
was smaller compared to the positive control mice which received Amphotericin B.
This was supported with histological analysis whereby sections from the flower
extract treated mice featured recovery of the injured tissue for all the examined
organs compared to severe damage observed in the negative control by end of the
study. By Day 28 conidia infection reduced in liver and was cleared from lung and
kidney in positive control. As a conclusion, C. surattensis flower could be a promising candidate in the pharmaceutical industry for new antifungal drug
discovery using plant as the main ingredient.
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Keywords
Antifungal agents