In vitro Antiviral Activity of Leaf Extracts Plantago major, Plantago lanceolata, Rubus idaeus

Abstract

Advances in organic chemistry, biochemistry, biotechnology and molecular virology have made it possible to synthesize a large number of antiviral drugs belonging to different pharmacological groups. However, one but significant disadvantage of these drugs is their significant toxicity. Therefore, along with the screening of new drugs among synthetic compounds, scientists are actively conducting research on antiviral agents of natural origin. Natural products with antiviral properties have advantages over synthetic compounds due to their low toxicity, minimal side effects, and mild action by various mechanisms. The aim of the study was to investigate the antiviral properties of aqueous-alcoholic extracts of plantain leaves (Plantago major L. and Plantago lanceolata L.), wild and garden raspberry leaves (Rubus idaeus L.) and their fermented variants on the model of human adenoviruses (HAdV3, HAdV5 and HAdV7). Methods. Determination of cytotoxicity and antiviral action of extracts was performed by standard methods using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). The titer of the virus, synthesized in the presence of drugs was determined by the end of dilution of the virus, which causes 50% development of the cytopathic effect of the virus on cells (СPE). Neoflazid was used as a reference drug. Neoflazid, developed by Ecopharm (Ukraine), it contains carboxylic acids and flavonoid glycosides isolated from wild cereals Deschampsia caespitosa L. (pike, turf) and Calamagrostis epigeios L. (dugout). All studies were performed in three replicates; the number of parallel determinations was 3–4. Calculated mean values, standard deviation, mean error. Differences in averages were considered significant at p<0.05. The research results were processed using Microsoft Office Excel 2010. Results. Low cytotoxicity of extracts of Plantago major L. and Rubus idaeus L. (wild) leaves and their fermented variants was shown, CC50 is >3 mg/mL. For extracts from the leaves of Plantago lanceolata L. and Rubus idaeus L. (garden) and their fermented variants, the toxicity was slightly higher (CC50=1.5 mg/mL). Extracts showed either a slight antiviral effect or its complete absence when used in a prophylactic regimen. We observed effective inhibition of reproduction of adenoviruses, when using extracts after adsorption of viruses. Extract of plantain leaves in concentrations of 0.06–3 mg/mL inhibited the reproduction of HAdV5 by 68–83% and inhibited the reproduction of HAdV3 and HAdV7 in concentration 3 mg/mL by 55% and 11%, respectively. Extract of Rubus idaeus L. (wild) leaves in the concentration range of 0.06–3 mg/mL inhibited the reproduction of HAdV5 by 65–89%, HAdV3 by 41–84% and HAdV7 by 22–59%. The maximum inhibition of reproduction of HAdV3 by 34% is shown for the extract from the leaves of Rubus idaeus L. (garden) at a concentration of 0.38 mg/mL, the reproduction of other viruses was suppressed by only 4–22%. It has been shown that the extracts of plantain and wild raspberry significantly affected the infectivity of viral offspring. Extract of plantain at a concentration of 3 mg/mL inhibited the reproduction of HAdV5 by 1.5 lg, fermented extract of plantain – by 1 lg. Fermented plantain extract at a concentration of 0.06 mg/mL inhibited the formation of new viral offspring, the index of reproductive inhibition (IRI) was 1.6 lg. Both fermented and unfermented Rubus idaeus L. (wild) extract had almost the same antiviral activity, IRI was 1.45 lg – 1.6 lg. Extracts of plantain and raspberry, regardless of the concentrations used, effectively inhibited the formation of infectious offspring of the HAdV3. The maximum IRI was 1.44 lg for plantain extract and 1.5 lg for fermented plantain extract. Both raspberry extracts (fermented and non-fermented) inhibited the synthesis of adenovirus serotype 3 by 1.46 – 1.54 lg. The drug Neoflazid completely inhibited the formation of infectious adenovirus at a concentration of 7.1 μg/mL. Virulicidal activity of all extracts against human adenoviruses 3, 5 and 7 serotypes was not detected. We found different antiviral activity of extracts of wild and garden raspberry leaves; we can assume that the flavonoid composition of the extracts plays an important role in their activity. Conclusions. Our new data on a wide range of anti-adenoviral activity of plantain and raspberry extracts is a prerequisite for further studies of the properties of individual components of extracts, in order to create an anti-adenoviral drug and recommendations for its pharmacological use.