Antiviral activity of the marine alga Symphyocladia latiuscula against herpes simplex virus (HSV-1) in vitro and its therapeutic efficacy against HSV-1 infection in mice

The antiviral activities of extracts from 5 species of marine algae collected at Haeundae (Pusan, Korea), were examined using plaque reduction assays. Although the activity of a methanol (MeOH) extract of Sargassum ringoldianum (Sargassaceae) was the most potent against several types of viruses, it was also cytotoxic. A MeOH extract of Symphyocladia latiuscula (Rhodomelaceae) and its fractions exhibited antiviral activities against acyclovir (ACV) and phosphonoacetic acid (PAA)-resistant (AP(r)) herpes simplex type 1 (HSV-1), thymidine kinase (TK(-)) deficient HSV-1 and wild type HSV-1 in vitro without cytotoxicity. The major component, 2,3,6-tribromo-4,5-dihydroxybenzyl methyl ether (TDB) of a CH(2)Cl(2)-soluble fraction was active against wild type HSV-1, as well as AP(r) HSV-1 and TK(-) HSV-1 (IC(50) values of 5.48, 4.81 and 23.3 microg/ml, respectively). The therapeutic effectiveness of the MeOH extract and TDB from S. latiuscula was further examined in BALB/c mice that were cutaneously infected with HSV-1 strain 7401H. Three daily oral administrations of the MeOH extract and TDB significantly delayed the appearance of score 2 skin lesions (local vesicles) and limited the development of further score 6 (mild zosteriform) lesions in infected mice without toxicity compared with controls. In addition, TDB suppressed virus yields in the brain and skin. Therefore TDB should be a promising anti HSV agent.     

Antiviral and antiplasmodial spirodihydrobenzofuran terpenes from the fungus Stachybotrys nephrospora

Two known spirodihydrobenzofuran terpenes (1 and 2) were isolated from a mycelium extract of the fungus Stachybotrys nephrospora BCC 3900. Compound 1 (Mer-NF5003F or stachybotrydial) exhibited potent antiviral activity (the IC50 value of 4.32 microg/mL) comparable to the standard drug, acyclovir, while compound 2 was inactive against the HSV-1 virus. Both 1 and 2 possessed antiplasmodial activity (IC50 values of 0.85 and 0.15 microg/mL for 1 and 2, respectively), and were not toxic towards the Vero cell line. A regiospecific conversion of the dialdehyde 1 to the lactone 2 proceeded simply under acidic conditions.     

The synergistic effects of betulin with acyclovir against herpes simplex viruses

Betulin, a pentacyclic triterpenoid, was isolated from the bark of Betula papyrifera. The antiviral efficacies of betulin on herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) were evaluated using viral plaque reduction assays on Vero cells. The results indicate that betulin is active against both HSV-1 and HSV-2 infections with the 50% effective concentrations (EC(50)) of 0.40 and 4.15 microg/ml, respectively. The cytotoxicity of betulin was examined on Vero cells using a neutral red uptake assay. The 50% cytotoxic concentration (CC(50)) of betulin was 73.1 microg/ml. A synergistic antiviral effect between betulin and acyclovir (ACV) was determined by drug combination studies. Strong and moderate synergistic antiviral effects were observed for betulin and ACV against HSV-1 when the concentrations of ACV and betulin were higher than 0.068 and 0.4 microg/ml, respectively. At the concentrations lower than these, additive effect was found. Synergistic antiviral effects were also found against HSV-2 at higher concentrations than for HSV-1, i.e. 0.45 microg/ml of ACV combined with 8.4 microg/ml of betulin.     

Xanthine oxidase inhibitory activity of Vietnamese medicinal plants

Among 288 extracts, prepared from 96 medicinal plants used in Vietnamese traditional medicine to treat gout and related symptoms, 188 demonstrated xanthine oxidase (XO) inhibitory activity at 100 microg/ml, with 46 having greater than 50% inhibition. At 50 microg/ml, 168 of the extracts were active, with 21 possessing more than 50% inhibition. At 25 microg/ml, 146 extracts exhibited inhibitory activity, with 8 showing over 50% inhibition, while 126 extracts presented activity at 10 microg/ml, with 2 having greater than 50% inhibition. The MeOH extracts of Artemisia vulgaris, Caesalpinia sappan (collected at the Seven-Mountain area), Blumea balsamifera (collected in Lam Dong province), Chrysanthemum sinense and MeOH-H(2)O extract of Tetracera scandens (Khanh Hoa province) exhibited strong XO inhibitory activity with IC(50) values less than 20 microg/ml. The most active extract was the MeOH extract of the flower of C. sinense with an IC(50) value of 5.1 microg/ml. Activity-guided fractionation of the MeOH extract led to the isolation of caffeic acid (1), luteolin (2), eriodictyol (3), and 1,5-di-O-caffeoylquinic acid (4). All these compounds showed significant XO inhibitory activity in a concentration-dependent manner, and the activity of 2 was more potent (IC(50) 1.3 microM) than the clinically used drug, allopurinol (IC(50) 2.5 microM).     

Inactivation of HSV-1 and HSV-2 and prevention of cell-to-cell virus spread by Santolina insularis essential oil

The essential oil obtained in toto from Santolina insularis was investigated for its antiviral activity on herpes simplex type 1 (HSV-1) and type 2 (HSV-2) in vitro. The IC(50) values, determined by plaque reduction assays, were 0.88 and 0.7 microg/ml for HSV-1 and HSV-2, respectively, while the CC(50) determined by the MTT test on Vero cells was 112 microg/ml, indicating a CC(50)/IC(50) ratio of 127 for HSV-1 and 160 for HSV-2. Results obtained by plaque reduction assays also indicated that the antiviral activity of S. insularis was principally due to direct virucidal effects. Antiviral activity against HSV-1 and HSV-2 was not observed in a post-attachment assay, and attachment assays indicated that virus adsorption was not inhibited. Up to 80% inhibition of HSV-1 was achieved at the concentration of 40 microg/ml by yield reduction assay. Furthermore, reduction of plaque formation assays also showed that S. insularis essential oil inhibits cell-to-cell transmission of both HSV-1 and HSV-2.     

Anti-herpesvirus activity of carbocyclic oxetanocin G in vitro

A series of new compounds, carbocyclic oxetanocins, have been synthesized and their anti-herpesvirus activity determined. Carbocyclic oxetanocin G (OXT-G) was most active against herpes simplex virus (HSV) and human cytomegalovirus (HCMV) among carbocyclic oxetanocins tested; the median effective concentrations (EC50) for HSV-1, -2, and HCMV were 0.23, 0.04 and 0.40 micrograms/ml, respectively. The EC50 value of carbocyclic OXT-G against HSV-2 was significantly lower than those of acyclovir, ganciclovir (DHPG) and OXT-G, while the value for HCMV was comparable to those of DHPG and OXT-G. Carbocyclic OXT-G showed much higher activity against TK+ HSV-2 than against a TK- mutant, suggesting that this compound is a good substrate for HSV-2-induced TK. The antiviral activity of the compound was only partially reversed even by the addition of 100-fold excess deoxyguanosine. The results suggest that the mode of action of carbocyclic OXT-G is different from that of OXT-G.     

Antiplasmodial activity of the alkaloids of Peschiera fuchsiaefolia

The tertiary and quaternary alkaloids isolated from the stem bark, root bark and seeds of Peschiera fuchsiaefolia are reported. The tertiary alkaloid crude extract from the stem bark was tested in vitro against Plasmodium falciparum on the basis of the antimalarial use of the plant. It showed good activity against both the D6 strain (IC50 = 495 ng/ml) and chloroquine-resistant W2 strain (IC50 = 817 ng/ml) and voacamine was the most active of the tested alkaloids (IC50 = 238 ng/ml for D6 and 290 ng/ml for W2). The tertiary alkaloid crude extract from the root bark of the same plant is more active than voacamine (IC50 = 179 ng/ml for D6 and 282 ng/ml for W2 strain), and is particularly rich in dimeric alkaloids (0.22% of the vegetable material).     

Antiviral activity of Plantago major extracts and related compounds in vitro

Plantago major L., a popular traditional Chinese medicine, has long been used for treating various diseases varying from cold to viral hepatitis. The aim of present study was to examine the antiviral activity of aqueous extract and pure compounds of P. major. Studies were conducted on a series of viruses, namely herpesviruses (HSV-1, HSV-2) and adenoviruses (ADV-3, ADV-8, ADV-11). The antiviral activity of EC50 was defined as the concentration achieved 50% cyto-protection against virus infection and the selectivity index (SI) was determined by the ratio of CC50 (concentration of 50% cellular cytotoxicity) to EC50. Results showed that aqueous extract of P. major possessed only a slight anti-herpes virus activity. In contrast, certain pure compounds belonging to the five different classes of chemicals found in extracts of this plant exhibited potent antiviral activity. Among them, caffeic acid exhibited the strongest activity against HSV-1 (EC50=15.3 microg/ml, SI=671), HSV-2 (EC50=87.3 microg/ml, SI=118) and ADV-3 (EC50=14.2 microg/ml, SI=727), whereas chlorogenic acid possessed the strongest anti-ADV-11 (EC50=13.3 microg/ml, SI=301) activity. The present study concludes that pure compounds of P. major, which possess antiviral activities are mainly derived from the phenolic compounds, especially caffeic acid. Its mode of action against HSV-2 and ADV-3 was found to be at multiplication stages (postinfection of HSV-1: 0-12 h; ADV-3: 0-2 h), and with SI values greater than 400, suggesting the potential use of this compound for treatment of the infection by these two viruses.     

Antiviral activity of hop constituents against a series of DNA and RNA viruses

We investigated whether crude hop extracts and purified hop components representing every major chemical class of hop compound have antiviral activity. These hop constituents were tested for antiviral activity against bovine viral diarrhea virus (BVDV) as a surrogate model of hepatitis C virus (HCV), human immunodeficiency virus (HIV), influenza A virus (FLU-A), influenza B virus (FLU-B), rhinovirus (Rhino), respiratory syncytial virus (RSV), yellow fever virus (YFV), cytomegalovirus (CMV), hepatitis B virus (HBV), and herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2). The extracts all failed to prevent the replication of HIV, FLU-A, FLU-B, RSV and YFV. A xanthohumol-enriched hop extract displayed a weak to moderate antiviral activity against BVDV (therapeutic index (TI)=6.0), HSV-2 (TI=>5.3), Rhino (TI=4.0) and HSV-1 (TI=>1.9) with IC(50) values in the low microg/ml range. Pure iso-alpha-acids demonstrated low to moderate antiviral activity against both BVDV (TI=9.1) and CMV (TI=4.2) with IC(50) values in the low microg/ml range. No antiviral activity was detected using beta-acids or a hop oil extract. Ultra-pure preparations (>99% pure) were used to show that xanthohumol accounted for the antiviral activity observed in the xanthohumol-enriched hop extract against BVDV, HSV-1 and HSV-2. Xanthohumol was found to be a more potent antiviral agent against these viruses than the isomer iso-xanthohumol. With Rhino, the opposite trend was observed with iso-xanthohumol showing superior antiviral activity to that observed with xanthohumol. Xanthohumol also showed antiviral activity against CMV, suggesting that it might have a generalized anti-herpesvirus antiviral activity. Again, superior antiviral activity was observed with the xanthohumol isomer against CMV. In summary, iso-alpha-acids and xanthohumol were shown to have a low-to-moderate antiviral activity against several viruses. These hop constituents might serve as interesting lead compounds from which more active anti-HCV, anti-Rhino and anti-herpesvirus antiviral agents could be synthesized.     

Inhibition of herpes simplex virus types 1 and 2 replication in vitro by mercurithio analogs of deoxyuridine.

The in vitro antiviral activity of several 5-mercurithio analogs of 2'-deoxyuridine (dUrd) on the replication of herpes simplex virus types 1 (HSV-1) and 2 (HSV-2) were examined. Of those compounds tested, the thioglycerol analog of 5-mercuri-2'-deoxyuridine (HgdUrd) was most effective in inhibiting the replication of HSV-1 in KB cells with a 50% inhibitory dose (ID50) of 0.001 micrograms/ml while the glutathione analog of HgdUrd was the most effective in inhibiting the replication of HSV-2 with a ID50 of 0.075 micrograms/ml. Conversely in HeLa TK- cells, the mercaptoguanosine analog of HgdUrd was the most effective compound in inhibiting virus replication with ID50S of 0.098 and 0.001 micrograms/ml for HSV-1 and HSV-2 respectively. These results suggest that these mercurithio analogs of dUrd are as effective as acyclovir in preventing the replication of these herpesviruses.     

Aldose reductase inhibitors from the fruiting bodies of Ganoderma applanatum

The isolation and characterization of rat lens aldose reductase (RLAR) inhibitors from the fruiting bodies of Ganoderma applanatum were conducted. Among the extracts and fractions from G. applanatum tested, the MeOH extract and EtOAc fraction were found to exhibit potent RLAR inhibition in vitro, their IC50 being 1.7 and 0.8 microg/ml, respectively. From the active EtOAc fraction, seven compounds with diverse structural moieties were isolated and identified as D-mannitol (1), 2-methoxyfatty acids (2), cerebrosides (3), daucosterol (4), 2,5-dihydroxyacetophenone (5), 2,5-dihydroxybenzoic acid (6), and protocatechualdehyde (7). Among them, protocatechualdehyde (7) was found to be the most potent RLAR inhibitor (IC50=0.7 microg/ml), and may be useful for the prevention and/or treatment of diabetic complications.     

Jacaranone: A cytotoxic constituent from<Emphasis Type="Italic">senecio ambiguus</Emphasis> subsp.<Emphasis Type="Italic">ambiguus</Emphasis> (Biv.) DC. Against renal adenocarcinoma achn and prostate carcinoma LNCaP cells

Senecio ambiguus subsp. ambiguus (Biv.) DC. extracts were able to inhibit the in vitro proliferation of renal cell adenocarcinoma ACHN and hormone dependent prostate carcinoma LNCaP. The potential cytotoxic property of the plant was revealed by the methanolic extract action against LNCaP (IC50 of 5.51 microg/mL) and ACHN (IC50 of 38.95 microg/mL). The most potent cytotoxic activity (IC50 of 5.34 microg/mL against the prostate carcinoma cell line) was exerted by the dichloromethane extract. Through bioassay-guided fractionation of the dichloromethane extract jacaranone was isolated as the major active constituent. This quinoid showed a very strong activity against ACHN and LNcaP with IC50 of 4.32 and 7.39 microg/mL, respectively. Its structure was established by GC/MS and NMR analysis. The n-hexane extract showed an interesting inhibition on the proliferation of tumor cell lines an IC50 value of 5.23 microg/mL against LNCaP. Three compounds identified in the n-hexane extract such as nerolidol, a-humulene and g-tocopherol were found to be active aginst LNCAP with IC50 values ranged from 11.24 to 15.56 microg/mL.     

N1-substituted thymine derivatives as mitochondrial thymidine kinase (TK-2) inhibitors

Novel N1-substituted thymine derivatives related to 1-[(Z)-4-(triphenylmethoxy)-2-butenyl]thymine have been synthesized and evaluated against thymidine kinase-2 (TK-2) and related nucleoside kinases [i.e., Drosophila melanogaster deoxynucleoside kinase (Dm-dNK) and herpes simplex virus type 1 thymidine kinase (HSV-1 TK)]. The thymine base has been tethered to a distal triphenylmethoxy moiety through a polymethylene chain (n = 3-8) or through a (2-ethoxy)ethyl spacer. Moreover, substitutions at position 4 of one of the phenyl rings of the triphenylmethoxy moiety have been performed. Compounds with a hexamethylene spacer (18, 26b, 31) displayed the highest inhibitory values against TK-2 (IC50 = 0.3-0.5 microM). Compound 26b competitively inhibited TK-2 with respect to thymidine and uncompetitively with respect to ATP. A rationale for the biological data was provided by docking some representative inhibitors into a homology-based model of human TK-2. Moreover, two of the most potent TK-2 inhibitors (18 and 26b) that also inhibit HSV-1 TK were able to reverse the cytostatic activity of 1-(beta-D-arabinofuranosyl)thymine (Ara-T) and ganciclovir in HSV-1 TK-expressing OST-TK-/HSV-1 TK+ cell cultures.     

Antiplasmodial activity of Uvaria klaineana

Crude extracts of Uvaria klaineana Engler and Diels (Annonaceae) stems showed in vitro activity against chloroquine-resistant K1 strain of Plasmodium falciparum. The most active extract was the basic dichloromethane extract containing crude alkaloids (IC50 = 3.55 microg/mL). The bioassay-guided fractionation of this extract led to the isolation of the major alkaloid crotsparine (1) which showed an antiplasmodial activity against the chloroquine-sensitive Thai strain of P. falciparum and the chloroquine-resistant K1 and FcB1 strains of P. falciparum. Two minor alkaloids were also identified as crotonosine (2) and zenkerine (3). Their structures were elucidated using 2D-NMR techniques.     

Screening of some Yemeni medicinal plants for inhibitory activity against peptidases

Extracts of different polarities (dichloromethane, methanol, and aqueous extracts) from 5 Yemeni medicinal plants (Aspilia helianthoides, leaves; Ceropegia rupicola, whole plant; Kniphofia sumarae, whole plant; Pavetta longiflora, leaves; and Plectranthus cf barbatus, leaves) were screened for their inhibitory effects against angiotensin converting enzyme (ACE), neutral endopeptidase (NEP), and aminopeptidase N (APN) activities. Four extracts (methanol extracts of Ceropegia rupicola, Kniphofia sumarae, and Plectranthus cf barbatus, and the aqueous extract of Pavetta longiflora) were found able to inhibit the enzymatic activity of NEP. Significant reduction in the activity of NEP (p < 0.01) was observed at a concentration of 50 microg/ml, and above of all tested extracts. The most active extract was the methanolic extract of Ceropegia rupicola with IC50 of 111 microg/ml. Only the methanolic extract of Aspilia helianthoides was found to exhibit inhibitory effect against the ACE activity with IC50 = 133 microg/ml. None of the tested plant extracts was found active against the aminopeptidase N activity.     

Penciclovir is a potent inhibitor of feline herpesvirus-1 with susceptibility determined at the level of virus-encoded thymidine kinase

Feline herpesvirus-1 (FHV-1) is the causative agent of a severe ocular disease in cats for which a safe potent antiviral chemotherapeutic agent is highly demanded. The sensitivity of FHV-1 to inhibition by three anti-herpetic nucleoside analogues [acyclovir (ACV), penciclovir (PCV) and cidofovir (CDV)] was tested by means of yield reduction assay. ACV showed very poor ability to inhibit FHV-1 replication. At low multiplicity of infection (MOI), both PCV and CDV were nearly equally effective with IC50 values ranging between 6 and 8 microg/ml. However, when the MOI was raised to 3PFU/cell, the activity of CDV was markedly reduced (IC50 25 microg/ml), while that of PCV remained relatively low (IC50 10 microg/ml). Although FHV-1 is normally insensitive to ACV, it exhibited >1000-fold increase in sensitivity when the thymidine kinase (TK) encoded by herpes simplex virus-1 (HSV-1) was supplied in trans. Furthermore, three PCV-resistant FHV-1 variants selected in vitro were shown to carry mutations in the TK gene. Taken together, these data provided direct evidence that PCV is a potent selective inhibitor of FHV-1 and that the virus-encoded TK is an important determinant of the virus susceptibility to nucleoside analogues.     

Synthesis, antiprotozoal and cytotoxic activities of new N-(3,4-dimethyl-5-isoxazolyl)-1,2-naphthoquinone-4-amino derivatives

Three derivatives of N-(3,4-dimethyl-5-isoxazolyl)-1,2-naphthoquinone-4-amino (1), a compound which exhibits significant activity against Trypanosoma cruzi and Plasmodium falciparum but with cytotoxicity toward murine L-6 cells, were synthesized with the aim of ameliorating its cytotoxicity. The in vitro antiprotozoal and cytotoxic activities of the synthesized compounds were evaluated against T. cruzi, Trypanosoma brucei rhodesiense, P. falciparum and murine L-6 cells. The hydroxymethyl (2) and the oxime (3) derivatives were active against T. cruzi, with IC50 values in a range comparable to those of 1 (IC50: 0.65 microg/ml) and benznidazole (IC50: 0.56 microg/ml) while the carboxymethyloxime (4) was inactive. Compounds 2 and 3 were cytotoxic toward L-6 cells, with IC50 values identical to that of 1 (IC50: 0.50 microg/ml). The results did not support the suggestion that 2 and 3 may be used as prodrugs of 1.     

Efficacy of orally administered Lobelia chinensis extracts on herpes simplex virus type 1 infection in BALB/c mice

By a plaque reduction assay, methanolic extracts from Lobelia chinensis (LC) significantly blocked herpes simplex virus type 1 (HSV-1) replication in HeLa cells without apparent cytotoxicity. The 50% inhibitory concentration (IC(50)) of LC on HSV-1 replication is 139.2 microg/ml. To elucidate LC anti-HSV-1 activity in vivo, BALB/c mice were injected subcutaneously with HSV-1 (2.5 x 10(6)PFU/50 microl), treated orally thrice a day with acyclovir (60 mg/kg/dose) or LC (20 and 50 mg/kg/dose) for 7 days, and inspected daily for signs of disease. Data from the scoring system indicated that animals infected with HSV-1, developed progressive zoster lesions starting 2 days postinfection (p.i.) and appeared the most serious syndromes at 4-5 days p.i. In marked contrast to the results with control mice, treatment with acyclovir or 50 mg/kg/dose LC resulted in a sustained protective effect. The HSV-1 titers and DNA levels in ground skin samples were significantly reduced by LC. No toxic effect of LC on liver and kidney functions was apparent. These results indicated that LC was a potent inhibitor of the in vitro and in vivo replication of HSV-1.     

Hepatoprotective constituents of the seeds of Combretum quadrangulare

Hepatoprotective effect of MeOH, MeOH-H2O (1:1) and H2O extracts of Combretum quadrangulare seeds were examined on D-galactosamine (D-GalN)/tumor necrosis factor-alpha(TNF-alpha)-induced cell death in primary cultured mouse hepatocytes. The MeOH extract showed the strongest inhibitory effect on D-GalN/TNF-alpha-induced cell death (IC50, 56.4 microg/ml). Moreover, the MeOH extract also significantly lowered the serum glutamic pyruvic transaminase (sGPT) level on D-GalN/lipopolysaccharide (LPS)-induced liver injury in mice. Bioguided separation of the MeOH extract led to the isolation of 38 compounds of various classes including triterpene glucosides, lignans and catechin derivatives. Among the isolated triterpene glucosides, lupane-type (1-3; IC50, 63.1, 59.8 and 76.2 microM, respectively) and ursane-type (11, mixture of 12 and 14; IC50, 30.2 and 34.6 microM, respectively) compounds exhibited strong hepatoprotective activity. 1-O-Galloyl-6-O-(4-hydroxy-3,5-dimethoxy)benzoyl-beta-D-glucose (26; IC50, 7.2 microM), methyl gallate (28; IC50, 19.9 microM), and (-)-epicatechin (31; IC50, 71.2 microM) also had a potent hepatoprotective effect on D-GalN/TNF-alpha-induced cell death in primary cultured mouse hepatocytes.