Elsevier

Antiviral Research

Volume 39, Issue 1, July 1998, Pages 55-61
Antiviral Research

Antiviral activity of Viracea® against acyclovir susceptible and acyclovir resistant strains of herpes simplex virus

https://doi.org/10.1016/S0166-3542(98)00027-8Get rights and content

Abstract

Viracea, a topical microbicide, is a blend of benzalkonium chloride and phytochemicals derived from Echinacea purpurea and is a proprietary formula from Destiny BioMediX Corp. Viracea was tested against 40 strains of herpes simplex virus (HSV): 15 strains (five HSV-1 and ten HSV-2) were resistant to acyclovir (ACV-R) and 25 strains (13 HSV-1 and 12 HSV-2) were susceptible to ACV (ACV-S). The median ED50 of Viracea for the five ACV-R strains of HSV-1 was a 1:100 dilution of the drug with a range of 1:50–1:400. The median ED50 of Viracea for the ten ACV-R strains of HSV-2 was 1:200 with a range of 1:50–1:3200. For the ACV-S strains of HSV-1 and HSV-2, the median ED50 of Viracea was 1:100 and 1:200, respectively. The cytotoxicity of Viracea was evaluated in a standard neutral red dye uptake assay in human foreskin fibroblasts. The cytotoxicity of Viracea approached only 50% at the highest concentration of the drug tested, a 1:2 dilution, indicating that Viracea is non-toxic in this cell cytotoxicity assay. Although the active component(s) in Viracea that has anti-HSV activity is not known, it appears that this extract has good antiviral activity against both ACV resistant and ACV susceptible strains of HSV-1 and HSV-2.

Introduction

Herpes simplex virus type 1 (HSV-1) infections are common in the US population and oropharyngeal infections are the most frequent infections caused by this virus (Corey and Spear, 1986a, Corey and Spear, 1986b). Genital herpes is an important sexually transmitted disease that is usually caused by HSV-2, but HSV-1 can also cause these infections (Johnson et al., 1989). In addition, one report suggests that HSV-2 may be a risk factor for the transmission of human immunodeficiency virus (Holmberg et al., 1988).

Recently, Fleming et al. (1997)reported that the seroprevalence of HSV-2 in persons 12 years of age or older in the USA is 21.9%, a 30% increase over the previous survey for all groups. The initial infection with HSV can be quite severe but is often mild or subclinical. Less than 10% of those who were seropositive reported a history of genital herpes (Fleming et al., 1997) and recurrences with HSV can vary from none to frequent. In immunocompromised patients and neonates, however, HSV infections can be severe and may cause systemic illnesses.

A large number of plants have been used by various societies for the treatment of diseases. Some traditional medicines consisting of these natural products have been shown by in vitro techniques to possess antiviral activity (Beuscher et al., 1994Taylor et al., 1996). Ferrea et al. (1993)found that methanolic extracts of dried Combretum micranthum leaves contained antiviral activity against HSV-1 and HSV-2. Meyer et al. (1996)described the inhibition of the cytopathic effect of HSV-1 in human lung fibroblasts using an aqueous extract of Helichrysum aureonitens (Asteraceae), a southern African medicinal plant. Of the 142 traditional medicines used in China, Indonesia, and Japan and evaluated by Kurokawa et al. (1993), 32 were found to have activity against HSV-1. The Indian medicinal plant, Pongamia pinnata, also has been shown to have in vitro activity against HSV (Elanchezhiyan et al., 1993). These materials were all prepared as extracts and the modes of action of most of these extracts are not known; however, an extract of the culture medium of mycelia from the edible mushroom Lentinus edodes appears to block the late stage of replication of HSV-1 (Sarkar et al., 1993). Yip et al. (1995)isolated the compound fluoranthene from the leaves and twigs of Elsholtzia ciliata, a Chinese medicinal plant, that has activity against Sindbis and murine cytomegalovirus.

HSV infections continue to increase, and ACV resistance has become a serious problem in certain patients; therefore, new anti-HSV drugs continue to be developed (Cassady and Whitley, 1997). Various phytochemicals have been shown to have in vitro antiviral activity and may be a good source of new antiviral agents. These agents have the potential to be modified and become more effective analogs than the original compounds. Viracea, a proprietary formula from Destiny BioMediX Corp., is a blend of benzalkonium chloride and phytochemicals derived from the aerial parts of Echinacea purpurea (L.) Moench. E. purpurea, the purple coneflower, is a native American plant and a member of the Asteraceae family. This study was designed to determine the in vitro antiviral activity of Viracea against ACV susceptible and resistant strains of HSV.

Section snippets

Antiviral compounds

A stock solution of ACV (Glaxo-Wellcome) was prepared in water at a concentration of 10 mM and frozen in aliquots at −20°C. The stock ACV was diluted in Eagle's minimal essential medium (MEM; BioWhittaker, Walkersville, MD) for use in susceptibility assays.

Echinacea purpurea powder, provided by Destiny BioMediX, Chicago, IL, was prepared from the aerial parts of mature E. purpurea plants. The plants were dried, powdered and sterilized with ethylene oxide. Viracea was made into an aqueous

The effect of temperature on the extraction of Viracea

The two control strains of HSV, HSV-1 strain F and HSV-2 strain G, were tested with the three extracts prepared at 37, 60 and 100°C. Each preparation was tested ten times. No differences were seen in the activity of Viracea prepared by extraction at either 37 or 60°C. The median ED50 for the preparations that were extracted at 37 or 60°C was the 1:200 dilution of Viracea. The median ED50 for the preparation extracted at 100°C was a 1:25 dilution. Since extracts prepared at either 37 or 60°C

Discussion

We examined the anti-HSV activity of Viracea against ACV susceptible and resistant strains of HSV-1 and HSV-2. The clinical strains were passed a limited number of times before the isolates were tested with ACV or Viracea. These isolates were characterized only as to their susceptibility to ACV and the mutations associated with ACV resistance in these clinical strains are not known.

Viracea was shown in this study to have good in vitro activity against clinical strains of HSV. No difference was

Acknowledgements

This work was supported in part by Destiny BioMediX, Chicago, IL.

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