Asian Pacific Journal of Tropical Disease

7-8

Authors: Sora Yasri, Viroj Wiwanitkit

The problem of Ebola virus infection is the big global concern. Preparedness for ongoing Ebola virus infection is the topic that should be discussed. In fact, it is necessary to set up a biosecurity system to protect against the present Ebola outbreak. The medical personnel have to prepare for fighting the problem. The management of the present outbreak requires international collaboration and control of cross-border disease transmission is also the big challenge. The good case study is the Hajj scenario.

11-12

Authors: Kamon Chaiyasit, Viroj Wiwanitkit

Ebola haemorrhagic fever is a viral infection causing a major health problem worldwide. In this short article, the authors briefly review and discuss on the nutritional management (energy, protein, fat and micronutrient) in management of Ebola infection.

15-18

Authors: Beena Uppal, Prabhav Aggarwal, Naz Perween, Anuj Sud

Objective: To determine the seroprevalence of Toxoplasma infection in the HIV infected and HIV non-infected individuals in our region, including antenatal women. Methods: Five mL of blood sample was collected from a total of 1 181 individuals aged 12 years and above. These included 661 (55.9%) from HIV positive patients and 520 (44.1%) from HIV negative individuals. A total of 238 samples out of the 520 HIV negative patients were collected from the antenatal women. Demographic profile of the subjects was recorded. Immunoglobulin G ELISA was performed for all the samples, while only the samples received from antenatal women were tested by immunoglobulin M capture ELISA . Results: Seroprevalence among HIV infected and non-infected was found to be 21.3% (95% confidence interval = 18.4%-24.6%) and 14.2% (95% confidence interval = 11.5%-17.5%), respectively. The difference was statistically significant (P = 0.003). No significant gender differences were found. Seroprevalence increased from 9.1% to 30% with increasing age in the HIV infected patients. Only 2 (0.84%) samples of antenatal women were positive for immunoglobulin M capture ELSIA , while one sample was equivocally reactive. Conclusions: Seroprevalence of latent toxoplasmosis in our region is moderately high, particularly in the HIV infected patients, exposing them to the risk of reactivation. This suggests that serologic testing of all HIV infected patients is essential to initiate Toxoplasma prophylaxis. Similarly, screening for active Toxoplasma gondii infection during antenatal care and preventive education is essential to prevent and minimize congenital toxoplasmosis.

27-30

Authors: Gavendra Singh, Soam Prakash

Objective: Entomopathogenic fungi like Beauveria bassiana, Metarhizium anisopliae have been significantly pathogenic for mosquito vectors. Although few have been used for control. Moreover, the genus Verticillium encompasses a cosmopolitan group of ascomycete fungi. It is a major plant pathogen and parasitic on other fungi and insects. Methods: The culture filtrates released from the Verticillium lecanii ( MTCC 3692) were grown potato carrot broth media. These filtrates were purified with Whatman-1 filter paper and flash chromatograph respectively. Results: The results demonstrated LC 50 , LC 90 , and LC 99 values of 0.6, 4.2, and 4.86, for Culex quinquefasciatus , and 1.3, 2.32, and 3.36 (µL/cm 2 ) for Anopheles stephensi after exposure for 10 h. LT 90 values were 6.76 for Culex quinquefasciatus and Anopheles stephensi 3.54 for Anopheles stephensi. Moreover, the Aedes aegypti were completely susceptible at all selected doses. Conclusions: The fungal culture filtrates of Verticillium sp. can reduce malaria, dengue, and filarial transmission on a par with chemical insecticides providing efficient delivery system can be developed.

33-41

Authors: Vijayakumar Subramaniyan, Ramesh Venkatachalam

Objective: To analyze the E and NS proteins of four dengue virus ( DENV) serotypes which involve in the formation of dengue fever and to find out the active binding sites on the surfaces of the above proteins to access with the effective drug molecule to counter dengue. Methods: ClustalW analysis for phylogram, BLASTP analysis for E and NS proteins of DENV serotypes and Swiss-Prot for 3D structure of E and NS proteins were used. Results: E proteins were grouped into two clusters, DENV-1 , DENV-2 and DENV-3 forming one cluster and DENV - 4 alone forming a different serotype. On the surfaces of E proteins of four serotypes, five binding sites for ligands were located in different DENV which the fourth binding site was identified as the active ligand binding site in DENV-2 , DENV-3 and DENV-4 serotypes and second binding site in the DENV-1 . Among the seven NS proteins, NS-1, NS-3 and NS-5 were identified as significant molecules involved in dengue disease. Conclusions: Binding sites, essential for ligand binding are found on all the E and NS proteins which active binding sites have been identified.

45-46

Authors: Beuy Joob , Viroj Wiwanitkit

Dengue virus infection is common in tropical world and becomes the big public health threat in tropical medicine. The changing of its epidemiology is very interesting. The good transportation is an important consideration. The concern on the imported dengue virus is interesting. On the other hand, the new alternative method corresponding to the infection is interesting. Here, the authors make some discussions on imported dengue virus and use of adjunctive recombinant activated factor VIIa in dengue.

51-53

Authors: Adil Raza, Sumit Kumar Varshney, Haris Manzoor Khan, Mohammad Ashraf Malik, Abbas Ali Mehdi, Indu Shukla

Objective: To estimate superoxide dismutase ( SOD ) activity in erythrocytes infected with Plasmodium falciparum in the cases of cerebral malaria. Methods: The diagnosis of cerebral malaria was made clinically and by Giemsa stained peripheral blood smear examination, quantitative buffy coat ( QBC ) examination and rapid antigen detection test ( RDT ). Parasitemia per micro litre of blood was evaluated by counting 200 white blood corpuscles and used to calculate parasite density considering 8000 white blood corpuscles per micro litre. SOD activity was estimated by the method given by Joe M. McCord and Irwin Fridovich spectrophotometrically. Statistical analysis was performed by using SPSS software version 17. Results: The SOD activity in the cases was found to be (1.06 ± 0.50) nmol/mL and that in the controls was (3.55 ± 0.07) nmol/mL. The SOD activity in the cases was significantly decreased (P < 0.05) as compared to the controls. The Pearson’s coefficient of correlation between SOD activity and parasitemia was found to be -0.93 showing strong negative relationship. Conclusions: There is severe oxidative stress in falciparum malaria due to reactive oxygen species and supplementation of antioxidants may modify the course and outcome of the disease.

59-66

Authors: Periyasamy Senthamarai Selvan, Arulsamy Jebanesan, Govindaraj Divya and Velu Ramesh

Objective: To study the diversity and distribution of mosquitoes in Western Ghats of Coimbatore and Nilgiris District, Tamilnadu, India. Methods: Random collections were carried out during August-2013 to July-2014 in cesspits, animal footprints, rock holes, tree holes, drainages at study areas of Marudhamalai, Valparai, Mettupalayam in Coimbatore District and Dhottapeta, Coonoor, Gudalur in Nilgiris District of Tamilnadu, India by using suction tube and kerosene pump. Mosquitoes were identified by standard entomological procedures. Results: A total of 1 018 mosquitoes (larvae and pupae) were collected from all over the study areas comprising 6 genera and 23 species. They are, Culex mimulus, Culex pseudovishnui, Culex quinquefasciatus, Culex vishnui, Culex khazani, Culex uniformis, Heizmannia chandi, Heizmannia grenii, Heizmannia indica, Oclerotatus anureostriatus, Oclerotatus albotaeniatis, Oclerotatus deccanus, Oclerotatus gubernatoris, Aedes aegypti, Aedes albopictus, Aedes edwardsi, Aedes krombeini, Toxorhynchites minimus, Toxorhynchites splendens, Anopheles aitkenii, Anopheles barbirostris, Anopheles culiciformis and Anopheles maculatus. Shannon- Weaver diversity index, Margalef’s index of richness and Simpsons dominance index was also studied. From 6 sites, the highest mosquitoes were collected from Marudhamalai (309) and the least mosquitoes were collected in Mettupalayam (68). The study determined whether physicochemical characteristics differ between habitats with high and low presence of mosquito larvae. Based on Margalef’s index of richness (D mg ), the highest values were present in Mettupalayam (5.214) study area and the lowest in Marudhamalai (3.837). It can be concluded from Shanon-Weaver index of diversity that, the highest values were present in Mettupalayam (2.947) and the least value were in Gudalur (2.410) during the study period. Conclusions: In areas with reservoirs of disease, mosquito abundance information can help to identify the areas at higher risk of disease transmission. The study identifies mosquito species density and diversity of culicine and anopheline larvae. Hence, this research contribution has more significance for basic biological research and developing control strategies for vector borne diseases.

73-76

Authors: Samuel Tennyson, John Ravindran, Alex Eapen, John William

Objective: To evaluate the larvicidal activity of Ageratum houstonianum (A. houstonianum) crude leaf extracts against the immatures of vector mosquitoes. Methods: Bioassays were performed in the laboratory with hexane, ethyl acetate and methanol crude leaf extracts of A. houstonianum at concentrations of 62.5, 125, 250, 500, 1 000, 2 000, 4 000 and 8 000 mg/L against the third instar larvae of Anopheles stephensi, Aedes aegypti and Culex quinquefasciatus. Results: Poor larvicidal activity was observed. The lowest LC 50 value was noted in ethyl acetate extract against all three vector mosquito species studied and was 3 377.84, 1 952.12 and 3 558.32 mg/L respectively after 24 h. The effect of toxicity was also manifested in a shorter period when compared to the other extracts viz., hexane and methanol. In Anopheles stephensi, more than 80% mortality was however observed at higher concentrations, after 24 h exposure in all the three extracts. In Aedes aegypti and Culex quinquefasciatus, this was observed by 3 and 24 h respectively in ethyl acetate extract. Conclusions: Screening of other parts of A. houstonianum with other solvents from different places for its larvicidal activity is recommended.

80-84

Authors: Rahul C.N, Shiva Krishna K, Meera M, Sandhya Phadke, Vidya Rajesh

Objective: To characterize the sequence diversity of blood-stage Plasmodium falciparum serine repeat antigen-5 ( PfSERA5 ) which is lacking in a malaria-endemic country like India. Methods: In this study, parasitic DNA was obtained from field isolates collected from various geographic regions. Subsequently, PfSERA5 gene sequence was PCR amplified and DNA sequenced. Results: We reported the existence of unique repeat polymorphisms and novel haplotypes for both the octamer repeat (OR) and serine repeat (SR) regions of the N-terminal fragment of PfSERA5 from Indian isolates. Several isolates from India were identical to low-frequency African haplotypes. Unique finding of our study was an Indian isolate showing deletion in a perfectly conserved 14 mer sequence within octamer repeat. Indian haplotypes reported in this study were found to be distributed into the three earlier classified allelic clusters of FCR3, K1 and Honduras showcasing broad diversity as compared to worldwide haplotypes. Conclusions: This study is the first report on genetic diversity of PfSERA5 antigen from India. Further evaluation of these haplotypes by serotyping would provide useful information for investigating variant-specific immunity and aid in malaria vaccine research.

89-92

Authors: Sunil Pandey, Amrit Lama Lo, Ravi Bhakta Shrestha

Objective: To find out the prevalence of parasitic infection among school aged children and to make necessary recommendations for preventive measures. Methods: The stool samples were examined for evidence of parasitic infections by direct microscopy and confirmed by concentration methods (formal ether sedimentation technique). Results: Among 300 samples, 15 (5.00%) were found to be positive for Entamoeba histolytica 5 (1.67%) followed by parasite Giardia lamblia 4 (1.33%), by parasite Ascaris lumbricoides 3 (1.00%) by parasite Hymenolepis nana 2 (0.67%), by Cyclospora 1 (0.33%). Conclusions: Major contributors for the prevalence of parasites were found to be poor personal hygiene and the educational level of the children. This should be regarded as an issue of public health priority and demand for effective school health programs involving periodic health education and screening.

96-98

Authors: Beuy Joob, Viroj Wiwanitkit

Opisthorchiasis is a kind of trematode infection. This parasitic infestation is a chronic hepatobiliary tract infection and can cause chronic irritation that will finally lead to cholangiocarcinoma. It is highly endemic in northeastern region of Thailand and contributes to many cholangiocarcinoma cases annually. The attempt to control the disease becomes a national policy. However, the sanitation becomes a major underlying factor leading to infection and meanwhile, the poverty and low education of the local people become an important concern. In this opinion, the authors discuss the effect of local environment and culture on opisthorchiasis in northeastern Thailand. Due to the pattern change of local environment, global warming and globalization, the dynamicity can be observed.

104-108

Authors: Zahra Mohtasham-Amiri, Davoud Pourmarzi, Maryam Razi

Objective: To determine epidemiological aspects of dog bite in Guilan Province, north of Iran. Methods: An analytical cross-sectional study was conducted on 1 643 cases of dog bites who presented at rabies vaccination centers in Guilan, Iran from June 2011 to May 2012. Data including demographic characteristics of dog bite cases, characteristics of biting dog, treatment and preventive measures carried out, and dog bite incident circumstances were collected. Independent t-test, ANOVA , Fisher exact test and chi-square test were used. Results: Dog bite incidences in men and women were 179.4 and 55 in 100 000 populations, respectively. Incidences in urban and rural were 72.8, and 181.9 in 100 000 population, respectively. The highest percentage of victims (20.1%) was in 20-29 years old age group. Majority of dogs (92%) were owned. Victims in the highest percentage (26.6%) were dog owners. Most of dog bites were occurred in houses (58.5%). Entering to the dog’s guarding territory was the most common circumstances (27.6%). Injuries most commonly involved the lower extremities (51%). Rabies vaccine, rabies immunoglobulin, tetanus vaccine and tetanus immunoglobulin were administered for 100%, 23.2%, 74.8%, and 9.1%, respectively. There were significant differences between men and women in term of area and place of bite incidence and dog ownership (P < 0.05). Mean age differences among categories of dog ownership, dog bite circumstance, and bitted site of body were significant (P < 0.05). Conclusions: Designing comprehensive educational programs to reduce dog bite incidence based on gender and age of target groups can be useful.

114-115

Authors: Naima Fasih, Kauser Jabeen, Tazeen Fatima, Rashida Ahmed, Ali Bin Sarwar Zubairi, Afia Zafar

Pulmonary cryptococcosis is usually underdiagnosed entity due its highly variable clinical presentation. In tuberculosis (TB)-endemic countries like Pakistan, its diagnosis is easily overlooked as there is a high degree of overlap in clinical, laboratory and radiographic presentation with TB. For definitive diagnosis fungal culture or molecular identification is required.

121-126

Authors: Tanzia Akon, Kamal Kanta Das, Luthfun Naher Nitu, Rashed Noo

Objective: To demonstrate the antibacterial activity of cosmetic products commonly used by the community of Dhaka metropolis. Methods: A total of 10 categories of cosmetic samples (with a subtotal of 30 brands) were subjected to microbiological analysis through conventional culture and biochemical tests. Agar well diffusion method was used to determine the antibacterial trait in the tested samples which was further confirmed by the minimum inhibitory concentration method. Results: All samples were found to be populated with bacteria and fungi up to 10 5 CFU / g and 10 3 CFU /g, respectively. Growth of Staphylococcus spp., Pseudomonas spp. and Klebsiella spp. was recorded as well. Conversely, 7 out of 30 items were found to exhibit the in vitro antibacterial activity against an array of laboratory test bacterial species including Staphylococcus spp., E. coli, Bacillus spp., Pseudomonas spp., Klebsiella spp. and Listeria spp. Consequently, all the samples showed antibacterial activity below the concentration of 0.46 mg/mL as found in the minimum inhibitory concentration test. Conclusions: Overall, the presence of huge microbial population in cosmetic products is not acceptable from the point microbiological contamination level. The antibacterial trait of these items, in contrary, may draw an overall public health impact.

133-136

Authors: Ali Sharifzadeh, Hossein Momeni, Payam Ghasemi-Dehkordi, Abbas Doosti

Objective: To investigate the presence of Listeria monocytogenes in the silage samples. Methods: Silage samples obtained from 150 different farms in Shahrekord city (Iran) and after DNA extraction, all samples were analyzed by PCR technique using one pair of primers for presence of this pathogen. The amplified products were detected on 1.5% agarose gel electrophoresis. Results: Listeria monocytogenes was isolated in 4 (2%) of the 150 samples. The detection of this bacterium from silage samples in Shahrekord city indicated that these products could create a serious risk in public health of animal and human. The findings showed that in positive silage samples for Listeria monocytogenes, the pH value was about five and it was due to bacterial activity in these products. Conclusions: The quality of silage and hygiene parameters and good herd health management play an important role in the microbiological quality of herd and farm. Considering the high specificity and sensitivity of the employed PCR technique, it is recommended to be useful technique for identification of Listeria monocytogenes.

142-144

Authors: Aziza Saif Rashid Alkhawalidy, Mohammad Amzad Hossain

Objective: To use the leaves of Annona squamosa (A. squamosa) for the preparation of crude extracts and all crude extracts were used to evaluate their antioxidant activity and total phenolics content collected from Jabal Al Akhdar. Methods: The leaves powder samples of A. squamosa were used for the preparation of crude extracts with methanol by Soxhlet extractor method. Defatted methanol crude extract by water was extracted by different polarities of solvents with increasing polarities. All crude extracts were used for the determination of total phenolics content and antioxidant activity by Folin- Ciocalteu reagent and 2,2-diphenyl-1-picrylhydrazyl methods. Results: The high amount of total phenolics content found in the leaves crude extracts was in the range of 54.75 to 352.0 mg/100 g of dry powder samples. The highest antioxidant activity was found in methanol and the lowest was found in ethyl acetate crude extract. Conclusions: The crude extracts from A. squamosa leaves showed strong antioxidant activity. Therefore, it could be used as a medicine for the treatment food borne diseases.

151-154

Authors: Rafiqul Islam, Md. Shahedur Rahman, Riad Hossain, Nazmun Nahar, Belal Hossin, Abir Ahad, Shaikh Mizanur Rahman

Objective: To find out the combined antibacterial efficacy of Aegle marmelos, Aphanamixis polystachya, Cuscuta reflexa and Aesclynomene indica against bacterial pathogens. Methods: Antibacterial potency of combined plant extracts has been tested against Bacillus subtilis IFO 3026 , Sarcina lutea IFO 3232 , Xanthomonas campestris IAM 1671 , Escherichia coli IFO 3007 , Klebsiella pneumoniae ATTC 10031 , Proteus vulgaris MTCC 321 and Pseudomonas denitrificans KACC 32026 by disc diffusion assay. Commercially available standard antibiotic discs were also used to find out antibiotic resistance pattern of test organisms. Results: Among the test organisms, Escherichia coli, Proteus vulgaris, Klebsiella pneumoniae and Proteus denitrificans showed resistance against multiple commercially available antibiotics. On the other hand, these multiple drug resistant organisms showed susceptibility against combined plant extracts. Conclusions: These combined plants extracts showed synergistic antibacterial activity and could lead to new antibacterial drug designing.

159-165

Authors: Emmanuel Anyachukwu Irondi, Ganiyu Oboh, Afolabi Akintunde Akindahunsi, Aline Augusti Boligon, Margareth Linde Athayde

Objective: To quantify some major pharmacologically important flavonoids and phenolic acids in Brachystegia eurycoma seed flour (BESF) and evaluate its antidiabetic activity in type 2 diabetic rats. Method: Flavonoids and phenolic acids were quantified using a reverse-phase high pressure liquid chromatrography coupled with diode array detection. Type 2 diabetes was induced in rats using high-fat diet, low-dose streptozotocin (HFD/STZ) model, by feeding the rats with HFD for 2 weeks followed by single dose administration of STZ (40 mg/kg body weight, intraperitoneally). The diabetic rats were later fed BESF-supplemented (10% and 20%) diets, or administered with metformin (25 mg/kg b.w.) for 21 days; the control rats were fed basal diet during this period. After the dietary regimen, the rats were sacrificed, and their blood, liver and pancreas samples were collected for biochemical assays. Results: The flavonoids (catechin, rutin, quercitrin, quercetin and kaempferol) and phenolic acids (gallic acid, caffeic, chlorogenic and ellagic acid) were abundant in BESF. BESF- supplemented diets (BESF-SD) significantly (P < 0.05) decreased the levels of fasting blood glucose (FBG); ameliorated the lipid profile, and significantly (P < 0.05) increased the hepatic glycogen of the diabetic rats. BESF-SD also significantly (P < 0.05) attenuated the oxidative stress induced by STZ in the liver and pancreas of the diabetic rats. The ameliorative effects of 20% BESF-SD compared (P > 0.05) with metformin administration in some of the biomarkers. Conclusion: The flavonoids and phenolic acids in BESF may have acted synergistically to produce the observed antidiabetic effects. BESF could therefore be an effective and affordable dietary therapy for the management of T2DM; and an excellent source for drug discovery.

169-174

Authors: Reham Zakaria Hamza, Mohammad Salem Al-Harbi

Objective: To evaluate the ameliorative role of silymarin or/and Nigella sativa (N. sativa) water extract against N-acetyl-p-aminophenol ( APAP )-induced renal function deterioration in male mice at the biochemical levels. Methods: The mice were divided into seven groups (10/group). The first group was served as control. The second group was treated with dose of APAP . The third and fourth groups were treated with silymarin alone and N. sativa water extract alone, respectively. The fifth and sixth groups were treated with combination of APAP with silymarin and APAP with N. sativa water extract, respectively. The seventh group was treated with a combination of both ameliorative compounds (silymarin and N. sativa water extract) with APAP and all animals were treated for a period of 30 days. Results: Exposure to APAP at the treated dose for mice led to an alteration of kidney function parameters, increase in the level of serum urea and creatinine. Also, paracetamol administration induced oxidative stress in kidney homogenates by increasing malondialdhyde level and decreasing superoxide dismutase and catalase activities and this stress was ameliorated by administration of either silymarin or N. sativa water extract. Conclusions: Administration of silymarin or/and N. sativa water extract to APAP -treated mice alleviate the toxicity of APAP , and this appeared clearly by biochemical improvement of kidney function parameters and antioxidant parameters. But, the alleviation is more pronounced with the both antioxidants. Thus, the pronounce effect of silymarin and N. sativa water extract is most effective in reducing the toxicity induced by APAP and improving the kidney function parameters and antioxidant status of kidney of male mice.

178-180

Authors: Manish Garg, Abha Mangal, Harsh Tak, Dharmendra Pal Singh, Atul Soni

Hydatid cysts of the spleen are uncommon (2-3.5%). Primary involvement of the spleen is even rarer. We present a case of a large primary hydatid cyst of the spleen in a 55-year-old woman. Diagnosis was based on computed tomography (CT) scan along with a high degree of clinical suspicion. En-masse excision was successfully performed, and the patient was discharged with prophylactic albendazole therapy. Unless there is a high degree of clinical suspicion, this life- threatening condition may be missed; hence, the importance of this report.

184-186

Authors: Shyamapada Mandal

The current communication, based upon the previous published papers in various scientific web-based journals, states the scenario of over-the-counter sales of antibiotics, including many other factors, that enhance people practice antibiotic self-medication world-wide, which in the developed countries the situation is little different having some resolution with the antibiotic self-medication problems. This paper also states about the prudent use of antibiotics through medical supervision and prescription in order to combat the unwanted antibiotic side effects including emergence of antibiotic resistant bacteria from antibiotic misusage.

190-191

Authors: Varikasuvu Seshadri Reddy, Pullaiah Pasupuleti, Balananda Perugu

No

202-208

Authors: Sabira Sultana, Hafiz Muhammad Asif, Naveed Akhtar, Khalil Ahmad

Medicinal plants are the part and parcel of human society to combat against different diseases from the dawn of human civilization. According to World Health Organization, approximately 80% population of the developing countries are facing difficulties to afford synthetic drugs and are relying on traditional medicines mainly of plant origin in order to maintain their primary health care needs. Plants are being used in various disorders e.g., gastrointestinal disorders, genitourinary problems, hepatobilliary discomforts and psychological and respiratory problems through time immemorial and people in western countries are now reverting back towards herbal medicines because of their extensive biological and medicinal activities, higher safety and lesser costs. Many plants are being traditionally used in the treatment of fever and their antipyretic activities have been confirmed scientifically. The current review clearly demonstrates the importance of medicinal plants in the treatment of fever. Furthermore, this review can help the scientists and researchers to find some new antipyretic agents from traditional medicinal plants.

214-223

Authors: Fidèle Ntchapda, Emmanuel Talla, Paul Sakava, Franco Tanzi, Fernand-Nestor Tchuenguem Fohouo, Joseph Mbafor Tanyi, ThÉophile Dimo

Objective: To evaluate the pharmacological property of erythrodiol, a natural triterpenoid contained in propolis, as vasodilatory agent, and to determine its mechanism of action. Methods: Rats aortic rings were isolated and suspended in organ baths, and the effects of erythrodiol were studied by means of isometric tension recording experiments. Nitric oxide (NO) was detected by ozone-induced chemiluminescence. The technique used to evaluate changes in intracellular Ca 2+ concentration in intact endothelium was opened aortic ring and loaded with 16 µmol Fura-2/AM for 60 min at room temperature, washed and fixed by small pins with the luminal face up. In situ, ECs were visualized by an upright epifluorescence Axiolab microscope (Carl Zeiss, Oberkochen, Germany) equipped with a Zeiss×63 Achroplan objective (water immersion, 2.0 mm working distance, 0.9 numerical apertures). ECs were excited alternately at 340 and 380 nm, and the emitted light was detected at 510 nm. Results: In aortic rings with intact endothelium pre-contracted with norepinephrine (10 -4 mol/L), the addition of erythrodiol (10 -8 -10 -4 mol/L) induced vasorelaxation in a concentration-dependent manner; in endothelium-denuded rings, the relaxant response induced by erythrodiol was almost completely abolished suggesting that vasorelaxation was endothelium-dependent. They had almost no relaxant effect on depolarised or endothelium-denuded aortic segments. The relaxation was significantly attenuated by pre-treatment with the NO synthase inhibitor Nv- nitro-L-arginine-methylester. Erythrodiol (10 -4 mol/L) was able to significantly increase NOx levels. This effect was completely abolished after removal of the vascular endothelium. Erythrodiol (100 µmol/L) caused a slow, long-lasting increase in intracellular Ca 2+ concentration. These results further supported the hypothesis that erythrodiol can induce activation of the NO/soluble guanylate cyclase/cyclic guanosine monophosphate pathway, as suggested by functional studies. Conclusions: The present results suggest that the mechanism of relaxation seems to be mainly mediated by the endothelial production of NO. Such a vasorelaxation was an endothelium- dependent effect, via the NO/soluble guanylate cyclase/cyclic guanosine monophosphate pathway. This result also suggests that erythrodiol causes a slow influx of extracellular Ca 2+ release from the intracellular Ca 2+ stores and an inhibition of Ca 2+ extruding mechanism. It can be concluded that erythrodiol may have interesting therapeutic potential as a new vasodilator drug, for protecting the cardiovascular system.