Anti-herpes simplex virus activity by Spirogyra spp. extracts

ฤทธิ์การยับยั้งไวรัสก่อโรคเริมโดยสารสกัด Spirogyra spp. / Aomhatai Deethae

Name: Aomhatai Deethae

Classification :.DDC: 579.8

LCSH: Algae -- Microbiology

LCSH: Herpes simplex

LCSH: Herpesvirus diseases

LCSH: Antiviral agents

Abstract: Herpes simplex virus (HSV) type 1 (HSV-1) and type 2 (HSV-2) are large DNA viruses, causing localized skin infections and can produce severity of infection leading to death in acute encephalitis. The diseases that are occurred as cold sore or fever blister on the lip is the most manifestation of HSV infection. HSV disease is not entirely therapy and it is easy for transmission. Many effective drug such as acyclovir, valaciclovir and penciclovir have disadvantages such as side effects, expensive cost and drug resistant strains may occur after long term treatment. Therefore, the antiviral agent of natural substance from algae becomes of considerable interest to study an efficacy against HSV infection. Anti-HSV activity of green freshwater macroalga, Spirogyra spp. or Tao that was locally found in Northern of Thailand, was investigated in this research. These macroalga were extracted with water, 95% ethanol and methanol. The result showed that the highest percentage yield of this algal extract was obtained from methanolic extract (ME) of Spirogyra spp., which was 23.52%. The extracts were examined for cytotoxicity on Vero cell by MTT assay. The result revealed that the aqueous extract (AE) of Spirogyra spp. showed the lowest cytotoxicity with CD50 value of 4,363.30 µg/ml, while the ME extract of Spirogyra spp. revealed the highest cytotoxicity in Vero cells with CD50 value of 250.80 µg/ml. Then, nontoxic concentrations of extracts were used to determine against HSV-1F, five samples of ACV-resistant HSV-1 isolates and HSV-2G by plaque reduction assay. It was found that the ME and ethanolic (EE) extract of Spirogyra spp. during and after viral attachment to Vero cell showed the highest anti-HSV against both types of HSV. Thus, the study of anti-viral effect on various stages of HSV infection was performed. Direct inactivation kinetic of HSV particles revealed the high ability of EE and ME extracts on inactivation of HSV-1F, ACV-resistant HSV-1 isolates and HSV-2G particles. Interestingly, HSV-2G was the highest inactivated by ME extract to completely inhibition within 2 hours, while the HSV-1F and ACV-resistant HSV-1 isolates were inactivated to negligible amounts within 4 hours. Moreover, the EE extract showed the highest ability to inhibit both types of HSV and ACV-resistant HSV-1 isolates during viral attachment, and after viral attachment. The EE extract also exhibited the highest inhibition on replication of HSV-1F and ACV-resistant HSV-1 isolates replication at 30 hours post infection, while the highest inhibition of HSV-2G was shown after treatment by AE extract. Moreover, the EE extract of Spirogyra spp. showed the potential inhibitory effect on HSV-1F and ACV-resistant HSV-1 viral DNA and protein synthesis, while the highest inhibition of HSV-2G DNA was demonstrated after treatment by AE extract. The EE and AE extracts of Spirogyra spp. were selected to further fractionate in order to determine the phytochemical analysis. The result illustrated that the EE01 and EE02 fractions which separated from the EE extract of Spirogyra spp. composed of alkaloid, essential oil and terpenoid. The EE01 and EE02 fractions showed the highest efficacy against HSV infection. Furthermore, the results of two-dimensional electrophoresis (2-DE) and protein-protein interactions were analyzed in this research. The result of 2-DE revealed the protein spots were decreased in infected cells after treatment with Spirogyra spp. extract. The results of the HSV-1F infected cells and HSV-1F infected cells after treatment with EE extract of Spirogyra spp. revealed nineteen differentially expressed protein spots. Moreover, the comparison of the HSV-2G infected cells and HSV-2G infected cells after treatment with AE extract of Spirogyra spp., reported twenty-six differentially expressed protein spots. The identification of differentially expressed HSV-1F and HSV-2G protein spots were analyzed by Mascot program. The protein interaction between HSV-1 and host cell proteins after treatment with EE extract suggested that Nesprin-1 protein was up-regulated but the integrin beta-2 and E3 ubiquitin were down-regulated. Whereas, the functional mechanism of changed global proteins were included the regulation of cell cycle and intracellular non-membrane bounded organelle of HSV-2G infected cell after treatment with AE extract. Therefore, algal gel product containing crude ethanolic extract of Spirogyra spp. was developed and determined for anti-HSV effect and stability. It was found that, the algal gel also retained the anti-viral effect against HSV-1F, ACV-resistant HSV-1 isolates and HSV-2G after 7 months of storage. Moreover, anti-HSV activity of the algal gel were also retained after 6 cycles of heating-cooling when storage at 4ºC for 48 hours and 45ºC for 48 hours. The algal gel also did not cause any irritation on skin of volunteers. Therefore, the knowledge from this research will be used for development of effective anti-HSV agent from Spirogyra spp. macroalgae extract.

Chiang Mai University Library

Address: CHIANG MAI

Email: cmulibref@cmu.ac.th

Name: Sittiruk Roytrakul

Role: Chairman

Name: Yingmanee Tragoolpua

Role: Member

Name: Yuwadee Peerapornpisal

Role: Member

Name: Padchanee Sangthong

Role: Member

Name: Jeeraporn Pekkoh

Role: Member

Name: Itthayakorn Promputtha

Role: Member

Created: 2016

Modified: 2020-01-09

Issued: 2020-01-09

วิทยานิพนธ์/Thesis

application/pdf

CallNumber: Th 579.8 A638A

eng

DegreeName: Doctor of Philosophy

Level: Doctoral Degree

Descipline: Applied Microbiology

Grantor: Chiang Mai University

©copyrights Chiang Mai University

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