Evaluation of Antiviral, Antimicrobial, Antiifungal and Anthelmintic activity of Pyrus Communis leaves extract
pyrus communis:
Taxonomy
Kingdom : Plantae
Division : Magnoliophyta
Class : Magnoliopside
Order: Rosales
Family: Rosaceae
Subfamily: Amygdaloideae
Tribe : Maleae
Sub-tribe: Malinae
Genus : Pyrus L.
Species : Communis Linn.
pyrus communis L. (pear) is an important medicinal plant that has been commonly used since prehistoric times. The pear is
known as ‘Amritphale’ due to its immense effect in health care system of human. Different compounds have been detected
in various parts of Pyrus communis L. like arbutin, kaempferol, quercetin, fredielin, isoquercitrin, sterols, ursolic acid, astragalin,
sorbitol, phloridzin and tannins that are responsible for various activities of pear such as anti-oxidant, anti-microbial, anti-
cancer, cholesterol-lowering, skin-whitening, anti-diabetic, wound healing effect, action on urinary system, immune booster
and weight loss effect.
Pyrus communis(pear) has many nutritional benefits as well as many medicinal uses. Pear is a sweet fruit that contains a
lot of calcium, vitamins and minerals. The pear comes from the tree Pyrus communis, also called
the common pear tree. The tree is 10 to 20 meters (33 to 66 feet) high. They live an average of
65 years. Kingdom of pyrus communis is planate and belongs to family rosaeace. Pear tree can
easily grow in heavy soil. Pear (Pyrus communis) is one of the most important fruit trees on the earth which origin is from Europe and North Africa (Chessell, 2011).
Pears contain special phytonutrients including anti-inflammatory flavonoids, anticancer
polyphenols and anti-ageing flavonoids. pear is also effective in diseases like a kidney stone,
decreasing cholesterol level and even diabetes (Abotaleb et al., 2019). Aerial and flowering parts of Pyrus communis are a
source of folk medicine as they show anti-inflammatory
effects (Zahid K et al., 2019; Velmurugan C and
Bhargava A, 2014). Pears are stimulate bowel peristaltic
movement and digestion, affect blood pressure, low in
calories, and show antipyretic, diuretic, and antitussive
activities (Chizzali C et al., 2016; Konarska A, 2013;
Reiland H and Slavin J, 2015). Its bark and leaves can be
used in healing of wound. Among the Arabs, buds, bark,
and leaves of the tree are domestic remedies for their
astringent effect (Velmurugan C and Bhargav A, 2013;
Nadkarni KM and Nadkarni AK, 2005). The pear flowers
are used in traditional medicine as components of
spasmolytic and analgesic drugs (Sharma K et al., 2015
Each
part of the pear tree has its importance due to its multiple role such as many medicinal as well
as nutritional values. The bark of the pear help reduces pain produced by blows, twists. Pear leaves also reduce the inflammations of the prostate (Fleming, 2013).
Flavonoid glycosides have been detected like
kaempferol 3-O--D (6”-O--L-rhamnopyranosyl)-
glucopyranoside, quercetin 3-O--D glucopyranoside and
quercetin 3-O--D-(6”-O-- L-rhamnopyranosyl)-
glucopyranoside. Triterpenes and sterols (-amyrin and
-sitosterol). Coumarins and phenolics are present in the
flowers of Pyrus communis L. Chlorogenic acid is also
identified in the flowers of Pyrus communis L. In the
stem bark of Pyrus communis L., triterpenoids are
identified (Kaur R and Arya V, 2012). Quercetin
considered as a major flavonoid in Pyrus communis juice,
having antithrombotic, antitumor, anti-inflammatory,
antioxidant, and anti-apoptotic effects. Quercetin was
reported to be effective in decreasing ischemia related
brain swelling and brain injury and also neuroprotective
in a zebrafish model (Arzoo and Parle M, 2017; Koppula
S et al., 2012).
Taxonomy
Kingdom : Plantae
Division : Magnoliophyta
Class : Magnoliopside
Order: Rosales
Family: Rosaceae
Subfamily: Amygdaloideae
Tribe : Maleae
Sub-tribe: Malinae
Genus : Pyrus L.
Species : Communis Linn.
Antiviral Test
The EV71 BrCr laboratory-adapted strain was used at a low multiplicity of infection (MOI) in a standardized antiviral assay as described earlier (Martínez-Gualda et al., 2017). Briefly, a serial dilution of the extract(s) was prepared in assay medium that was added to an empty microtiter 96-well plate, after which the virus inoculum was added, followed by a suspension of freshly harvested rhabdosarcoma (RD) cells (2 × 104 cells/well). While the cells were settling to the bottom, the virus was allowed to infect them in the presence of extract. The assay plates were incubated at 37°C, 5% CO2 with virus inoculum and compounds until full virus-induced cell death was observed in the untreated, infected controls (3–4 days post-infection). Subsequently, the antiviral effect was quantified using a colorimetric readout with 3- (4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium/phenazine methosulfate (MTS/PMS method) and the concentration of compound at which 50% inhibition of virus-induced cell death would be observed (EC50) was calculated from the antiviral dose-response curves. A similar assay setup was used to determine the adverse effect of an extract or test compound on uninfected, treated cells for calculation of the CC50 (concentration of sample that reduces overall cell health by 50% as determined by the MTS/PMS method). The final, maximal DMSO concentration that was reached in the assay wells with the highest sample input (1%) was well-tolerated by the cells. Rupintrivir was used as a positive control (Martínez-Gualda et al., 2017). In addition, the selectivity index (SI) was calculated as the ratio of the CC50 for cell growth to the EC50.The Selectivity Surface-SS, was determined from plots (CPE and CC curve) and the Therapeutic Index-TI was calculated as (SS*10logSI) (Supplementary Material).
Anthelmintic Assay
The anthelmintic assay was carried out in a 96-well microplate (flat-bottom, TPP Techno Plastic Products AG, Switzerland). Synchronized C. elegans (15 μL, ~45 L4 larvae) were added to each well of a 96-well microplate containing 184 μL of E. coli OP50 culture (OD = 0.5 at 620 nm). Subsequently, 1 μL of plant extract was added to each well; DMSO (1 μL) was used in a separate well as a solvent control. After mixing, the 96-well microplate was placed into a WMicroTracker (Phylumtech, Argentina) apparatus and incubated for 16 h at 20°C. The movement of worms in each well was measured every 30 min and recorded by the WMicroTracker. The percentage of the average movement over 16 h of test samples with extract compared to the DMSO control, was used to estimate the relative anthelmintic activity. Levamisole (final concentration 50 μM) was systematically used as a positive control.
Antifungal Test
Antifungal activity was tested against C. albicans in a similar way as for bacteria. Instead of MH broth, YPD broth was used with a smaller quantity of plant extract (to keep the final DMSO concentration below 2%). For antifungal activity, each well of a microdilution plate was inoculated with 196 μL of the diluted yeast suspension and 4 μL of the test solution was added. Control wells were prepared with 196 μL YPD broth and 4 μL extract to correct for any absorption due to extract components, or 4 μL of DMSO or MilliQ (solvent control), or the antifungal miconazole (200 μg/mL) in DMSO (positive control).
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