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SARS-CoV-2 rapid spread required urgent, accurate, and prompt diagnosis to control the virus dissemination and pandemic management. Several sensors were developed using different biorecognition elements to obtain high specificity and sensitivity. However, the task to achieve these parameters in combination with fast detection, simplicity, and portability to identify the biorecognition element even in low concentration remains a challenge. Therefore, we developed an electrochemical biosensor based on polypyrrole nanotubes coupled via Ni(OH)2 ligation to an engineered antigen-binding fragment of heavy chain-only antibodies (VHH) termed Sb#15. Herein we report Sb#15-His6 expression, purification, and characterization of its interaction with the receptor-binding domain (RBD) of SARS-CoV-2 in addition to the construction and validation of a biosensor. The recombinant Sb#15 is correctly folded and interacts with the RBD with a dissociation constant (KD) of 27.1 ± 6.4 nmol/L. The biosensing platform was developed using polypyrrole nanotubes and Ni(OH)2, which can properly orientate the immobilization of Sb#15-His6 at the electrode surface through His-tag interaction for the sensitive SARS-CoV-2 antigen detection. The quantification limit was determined as 0.01 pg/mL using recombinant RBD, which was expressively lower than commercial monoclonal antibodies. In pre-characterized saliva, both Omicron and Delta SARS-CoV-2 were accurately detected only in positive samples, meeting all the requirements recommended by the World Health Organization for in vitro diagnostics. A low sample volume of saliva is needed to perform the detection, providing results within 15 min without further sample preparations. In summary, a new perspective allying recombinant VHHs with biosensor development and real sample detection was explored, addressing the need for accurate, rapid, and sensitive biosensors.

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BACKGROUND: There is great comorbidity and similarity between chronic pain and major depressive disorders. We have recently shown that 10 days of social defeat stress (SDS) induces hyperalgesia regardless depressive-like behavior in mice. Here we aimed to investigate whether social stress predisposes to chronic pain and, inversely, whether chronic pain predisposes to stress-induced depression. METHODS: Firstly, we used the 10 days SDS paradigm in mice followed by a mild protocol of repetitive inflammatory stimulus to evaluate if SDS would predispose to persistent hyperalgesia development. Secondly, we used the intense protocol of repetitive inflammatory stimulus followed by a subthreshold SDS to evaluate if persistent hyperalgesia would predispose to depressive-like behavior of social avoidance. RESULTS: Our results showed that SDS predispose to chronic pain, since stressed mice injected with PGE2 for 7 days (mild protocol), stimuli normally not sufficient to trigger chronic pain, showed persistent hyperalgesia. Also, we showed that persistent hyperalgesia induced by repetitive inflammatory stimuli predispose to long-lasting depressive-like behavior of social avoidance induced by subthreshold SDS. LIMITATIONS: We did not analyze molecular mechanism associated with chronic pain and depressive-like behavior induced by SDS. However, we hypothesized that SDS and 14 days of PGE2 would generate neuroplasticity on brain areas shared by chronic pain and depression, predisposing to pain chronification and depressive-like behavior, respectively. CONCLUSIONS: We can conclude social stress as a key and a common factor for chronic pain and depression. We can also conclude that SDS predisposes to chronic pain and, inversely, chronic pain predisposes to depressive-like behavior.

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Major depressive disorders (MDD) and chronic pain (CP) affect significant portion of the world's population and have high comorbidity rate. Social defeat stress (SDS) model was standardized in mice and can trigger depressive-like behavior and chronic pain. Based especially on clinical trials showing an effective preventive and therapeutic effect of physical exercise on CP and symptoms associated with MDD, this study aimed to investigate if the voluntary running wheel exercise can exert these effects in mice submitted to the 10-day SDS protocol, using fluoxetine as positive control. For this, we ran two set of experiments: in the first set mice started performing voluntary running wheel exercise after submitted to SDS and, in the second set, mice performed voluntary running wheel exercise before and during SDS. Mechanical and chemical hyperalgesia was analyzed through electronic von Frey and capsaicin test, respectively. Depressive-like behavior was assessed through social interaction test. Our results showed that the voluntary running wheel exercise was more effective than fluoxetine reversing the SDS-induced persistent hyperalgesia and both, fluoxetine and voluntary running wheel exercise, was effective reversing SDS-induced social avoidance. Also, voluntary running wheel exercise is an effective tool preventing both hyperalgesia and social avoidance induced by SDS. To the best of our knowledge, this was the first study using physical exercise as a therapeutic and preventive tool for chronic pain and depressive-like behavior simultaneously induced by social stress.

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The bovine appeasing substance (BAS) is expected to have calming effects in cattle experiencing stressful situations. Therefore, this study investigated the impacts of BAS administration during two of the most stressful events within beef production systems: weaning and feedlot entry. In experiment 1, 186 Bos indicus-influenced calves (73 heifers, 113 bulls) were weaned at 211 ± 1 days of age (day 0). At weaning, calves were ranked by sex and BW, and assigned to receive BAS (Nutricorp, Araras, SP, Brazil; n = 94) or water (CON; n = 92). Treatments (5 ml) were topically applied to the nuchal skin area of each animal. Calf BW was recorded and samples of blood and tail-switch hair were collected on days 0, 15 and 45. Calves that received BAS had greater (P < 0.01) BW gain from day 0 to 15 compared with CON. Overall BW gain (days 0 to 45) and BW on days 15 and 45 were also greater (P ≤ 0.03) in BAS v. CON. Plasma haptoglobin concentration was less (P < 0.01) in BAS v. CON on day 15, whereas cortisol concentrations in plasma and tail-switch hair did not differ between treatments (P ≥ 0.13). In experiment 2, 140 B. indicus-influenced bulls (∼27 months of age) from 2 different pasture-based systems (70 bulls/origin) were transported to a commercial feedlot (≤ 200-km transport; day -1). On day 0, bulls were ranked by source and BW, and assigned to receive BAS (n = 70) or CON (n = 70) and the same sampling procedures as in experiment 1. Bulls receiving BAS had greater (P = 0.04) BW gain from day 0 to 15, but less (P < 0.01) BW gain from day 15 to 45 compared to CON. No other treatment effects were detected (P > 0.14). Therefore, BAS administration to beef calves alleviated the haptoglobin response associated with weaning, and improved calf growth during the subsequent 45 days. Administration of BAS to beef bulls at feedlot entry improved BW gain during the initial 15 days, but these benefits were not sustained throughout the 45-day experiment.

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This study evaluated the effects of 6 weeks of whole-body vibration (WBV) exercise on flexibility and the rating of perceived exertion (RPE) in metabolic syndrome (MetS) individuals using 2 biomechanical conditions (fixed frequency [FF] and variable frequency [VF]). Nineteen MetS individuals were randomly allocated in FF-WBV (n = 9, 7 women and 2 men) and VF-WBV (n = 10, 8 women and 2 men) groups. Anterior trunk flexion (ATF) and RPE were determined before and after each session. The acute cumulative exposure effects were analyzed. The FF-WBV group was exposed to 5 Hz on a side alternating vibrating platform (SAVP), exposed to 10 and 50 seconds with the SAVP turned off. The VF-WBV group individuals were intermittently exposed (1 minute WBV exercise/1 minute rest) to 5 to 16 Hz, increased by 1 Hz per session and the peak-to-peak displacement (PPD) were 2.5, 5.0, and 7.5 mm. Regarding to ATF, significant improvements (P < .05) were observed in the in the acute (VF group) and cumulative intervention (FF and VF-WBV groups). The RPE significantly (P < .05) improved only in VF-WBV (cumulative intervention). In conclusion, WBV exercise improved the flexibility and decreased the RPE in MetS individuals. These findings suggest that WBV exercise can be incorporated into physical activities for MetS individuals.

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Type 1 diabetes mellitus (T1DM) is a disorder resulting from chronic autoimmune destruction of insulin-producing pancreatic ß-cells, lack of insulin production and hyperglycaemia. The aim of this study was to evaluate the hypothesis that streptozotocin-diabetic mice treated with Saccharomyces boulardii THT 500101 strain present improvement of glucose and triglycerides metabolism, reduction of liver inflammation concomitant with a beneficial impact in the gut microbiota profile. C57BL/6 male mice were randomly assigned into three groups: Control, Diabetes, Diabetes+Probiotic, and were euthanised 8 weeks after probiotic chronic administration. Mice submitted to treatment presented reduced glycemia in comparison with the diabetic group, which was correlated with an increase in C-peptide level and in hepatic glycogen content. Fat metabolism was significantly altered in streptozotocin-induced diabetic group, and S. boulardii treatment regulated it, leading to a decrease in serum triglycerides secretion, increase in hepatic triglycerides storage and modulation of inflammatory profile. The phenotypic changes seen from chronic S. boulardii treatment were found to be broadly associated with the changes in microbioma of diabetic animals, with increased proportion in Bacteroidetes, Firmicutes and Deferribacteres, and a decreased proportion of Proteobacteria and Verrucomicrobia phylum. Thus, the data presented here show up a novel potential therapeutic role of S. boulardii for the treatment and attenuation of diabetes-induced complications.

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Adolfo Lutz Institute in Sao Paolo State performs mycobacterial identification for many healthcare units, and in 2008 identified a possible outbreak involving patients submitted to bronchoscopy at the same hospital. This study aimed to analyse the clonality of isolates. Mycobacterium abscessus subsp. massiliense isolated from 28 patients, water from one bronchoscope and water from four automated endoscope reprocessing machines presented high similarity by pulsed-field gel electrophoresis. This strain was not found in the water supply, and it was hypothesized that an infected patient contaminated the bronchoscope, with further false-positive cultures from subsequent patients.

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The primary aim of this study was to evaluate penile endothelial microvascular function in patients with primary arterial hypertension and age-matched normotensive subjects using laser speckle contrast imaging (LSCI). Additionally, we analyzed the acute penile microvascular effects induced by oral phosphodiesterase type 5 inhibitor (sildenafil; SIL) administration. Endothelium-dependent microvascular reactivity was evaluated in the penises and forearms of hypertensive patients (aged 58.8±6.6 years, n=34) and age-matched healthy volunteers (n=33) at rest and 60 min following oral SIL (100 mg) administration. LSCI was coupled with cutaneous acetylcholine (ACh) iontophoresis using increasing anodal currents. Basal penile cutaneous vascular conductance (CVC) values were not significantly different between control subjects and hypertensive individuals. Penile CVC values increased significantly after SIL administration in control (P<0.0001) and hypertensive (P<0.0001) subjects. Peak CVC values were not different between the two groups during penile ACh iontophoresis before SIL administration (P=0.2052). Peak CVC values were higher in control subjects than in hypertensive subjects after SIL administration (P=0.0427). Penile endothelium-dependent microvascular function is, to some extent, preserved in patients presenting with primary arterial hypertension under effective anti-hypertensive treatment. LSCI may be a valuable non-invasive tool for the evaluation of penile vascular responses to phosphodiesterase type 5 inhibitor.

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This experiment compared insulin sensitivity, milk production, and reproductive outcomes in dairy cows consuming excessive energy during mid to late lactation and receiving in a 2 × 2 factorial design (1) concentrate based on ground corn (CRN; n = 20) or including 8% (DM basis) of Ca salts of palm oil (CSPO; n = 20), and (2) supplemented (n = 20) or not (n = 20) with 2.5 g/d of Cr-propionate. During the experiment (d 0-203), 40 multiparous, nonpregnant, lactating 3/4 Holstein × 1/4 Gir cows (initial days in milk = 81 ± 2; mean ± SE) were offered corn silage for ad libitum consumption, and individually received concentrate formulated to allow diets to provide 160% of their daily net energy for lactation requirements. From d -15 to 203, milk production was recorded daily, blood samples collected weekly, and cow body weight (BW) and body condition score (BCS) recorded on d 0 and 203. For dry matter intake evaluation, cows from both treatments were randomly divided in 5 groups of 8 cows each, and allocated to 8 individual feeding stations for 3 d. Intake was evaluated 6 times/group. Glucose tolerance tests (GTT; 0.5 g of glucose/kg of BW) were performed on d -3, 100, and 200. Follicle aspiration for in vitro embryo production was performed via transvaginal ovum pick-up on d -1, 98, and 198. Mean DMI, net energy for lactation intake, as well as BW and BCS change were similar across treatments. On average, cows gained 40 kg of BW and 0.49 BCS during the experiment. Within weekly blood samples, CRN cows had lower serum concentrations of glucose, insulin, fatty acids, and insulin-to-glucose ratio compared with CSPO cows, suggesting increased insulin sensitivity in CRN cows. During the GTT, insulin-sensitivity traits were also greater in CRN versus CSPO cows. Supplemental Cr-propionate resulted in lower serum insulin concentrations and insulin-to-glucose ratio within CRN cows only, indicating that Cr-propionate improved basal insulin sensitivity in CRN but not in CSPO cows. During the GTT, however, Cr-propionate supplementation reduced hyperinsulinemia and insulin-to-glucose ratio across CSPO and CRN cows. Milk production, as well as number of viable oocytes collected and embryos produced within each aspiration, were not affected by treatments. Hence, replacing corn by Ca salts of palm oil in the concentrate did not improve insulin sensitivity in Holstein × Gir dairy cows consuming excessive energy during mid to late lactation, whereas Cr-supplementation was effective in improving basal insulin sensitivity in cows not receiving Ca salts of palm oil.

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This study compared vaginal temperature, physiologic, and productive parameters in lactating dairy cows supplemented or not with Omnigen-AF (Phibro Animal Health, Teaneck, NJ) during the summer months in a tropical environment. Thirty-two lactating, primiparous (n = 16) and multiparous (n = 16) pregnant Holstein × Gir cows were ranked by parity, days in milk, body weight, and body condition score (BCS), and assigned to receive (SUPP; n = 16) or not (CON; n = 16) Omnigen-AF (Phibro Animal Health, Teaneck, NJ) at 56 g/cow daily (as-fed basis). During the experimental period (d -6 to 56), cows were maintained in a single drylot pen with ad libitum access to water and a total mixed ration, and milked twice daily. Cows received Omnigen-AF mixed with 200 g of corn (as-fed basis) after the daily morning milking through self-locking head gates, whereas CON cows concurrently received 56 g of kaolin mixed with 200 g of corn. For feed intake evaluation, cows from both treatments were randomly divided in 4 groups of 8 cows each, and allocated to 8 individual feeding stations for 3 d. Intake was evaluated 4 times per group from d 1 to 56. From d -6 to 0, d 15 to 28, and d 43 to 56, cow vaginal temperature was recorded hourly. Environmental temperature-humidity index (THI) was also recorded hourly from d 15 to 28 and d 43 to 56. Cows were evaluated for body weight and BCS on d -6 and 56, individual milk production was recorded daily from d -6 to 56, and milk samples were collected on d -6, 0, 7, 14, 21, 28, 35, 42, 49, and 56 for analyses of somatic cell count and milk components. Blood samples were collected on d -6, -3, 0, 9, 15, 18, 21, 24, 27, 36, 45, 48, 51, 54, and 56. Results from samples or observations collected from d -6 to 0 were included as an independent covariate in each respective analysis. Environmental THI was 74.2 ± 0.5 and cows were exposed to THI >68 for 633 h within a total of 672 h of evaluation. Cows assigned to CON had greater vaginal temperature on d 28, 43, 45, and from d 48 to 55 (by 0.38 to 0.52%), as well as greater mean somatic cell count (by 97%) and serum haptoglobin concentrations (by 89%) compared with SUPP cows. Cows assigned to SUPP had greater mean dry matter intake (by 7%), BCS on d 56 (by 11%), and mean serum insulin concentrations (by 35%) compared with CON cows. Hence, SUPP ameliorated hyperthermia, improved nutritional status, and modulated systemic and mammary gland immune parameters in lactating dairy cows exposed to heat stress conditions.

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This experiment compared insulin sensitivity parameters, milk production and reproductive outcomes in lactating dairy cows consuming excessive energy, and receiving in a 2×2 factorial arrangement design: (1) concentrate based on ground corn (CRN; n=13) or citrus pulp (PLP; n=13), and (2) supplemented (n=14) or not (n=12) with 2.5 g/day of chromium (Cr)-propionate. During the experiment (day 0 to 182), 26 multiparous, non-pregnant, lactating Gir×Holstein cows (initial days in milk=80±2) were offered corn silage for ad libitum consumption, and individually received concentrate formulated to allow diets to provide 160% of their daily requirements of net energy for lactation. Cow BW and body condition score (BCS) were recorded weekly. Milk production was recorded daily and milk samples collected weekly. Blood samples were collected weekly before the morning concentrate feeding. Glucose tolerance tests (GTT; 0.5 g of glucose/kg of BW) were performed on days -3, 60, 120 and 180. Follicle aspiration for in vitro embryo production was performed via transvaginal ovum pick-up on days -1, 82 and 162. No treatment differences were detected (P⩾0.25) for BW and BCS change during the experiment. Within weekly blood samples, concentrations of serum insulin and glucose, as well as insulin : glucose ratio were similar among treatments (P⩾0.19), whereas CRN had less (P<0.01) non-esterified fatty acid concentrations compared with PLP (0.177 v. 0.215 mmol/l; SEM=0.009). During the GTT, no treatment differences were detected (P⩾0.16) for serum glucose concentration, glucose clearance rate, glucose half-life and insulin : glucose ratio. Serum insulin concentrations were less (P=0.04) in CRN supplemented with Cr-propionate compared with non-supplemented CRN (8.2 v. 13.5 µIU/ml, respectively; SEM=1.7), whereas Cr-propionate supplementation did not impact (P=0.70) serum insulin within PLP cows. Milk production, milk fat and solid concentrations were similar (P⩾0.48) between treatments. However, CRN had greater (P<0.01) milk protein concentration compared with PLP (3.54% v. 3.14%, respectively; SEM=0.08). No treatment differences were detected (P⩾0.35) on number of viable oocytes collected and embryos produced within each aspiration. In summary, feeding a citrus pulp-based concentrate to lactating dairy cows consuming excessive energy did not improve insulin sensitivity, milk production and reproductive outcomes, whereas Cr-propionate supplementation only enhanced insulin sensitivity in cows receiving a corn-based concentrate during a GTT.

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Objective To evaluate the effect of short-term hormone replacement therapy with tibolone 2.5 mg daily on endothelial function of healthy postmenopausal women, using flow-mediated dilation (FMD) of the brachial artery. Methods We performed a randomized, double-blinded, placebo-controlled study. A total of 100 healthy postmenopausal women were randomly allocated to receive tibolone (n = 50) or placebo (n = 50) for 28 days. Measurement of the FMD of the brachial artery was performed before and after the use of tibolone and placebo. Results A total of 31 women completed the study in the tibolone group, and 32 women completed the study in the control group. The results of the FMD measurements obtained from the women in the two groups before treatment were similar (0.018 and 0.091, for tibolone and placebo, p = 0.57). The values of the FMD in women who used tibolone and placebo, before and after the treatment, were similar in both groups. The numbers of women who presented an increase in the values of the FMD in both groups were also similar. Conclusion Our results demonstrate that the administration of 2.5 mg tibolone to healthy postmenopausal women for 28 days does not promote endothelial-dependent vasodilation, measured by FMD of the brachial artery.

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STUDY DESIGN: Cross-sectional study. OBJECTIVES: The present study aims to analyze and correlate the interlimb reflexes (ILRs), through a standard methodology, in tetraplegic and healthy subjects. The study of the connectivity between the injured spinal cord and the ILR transmission empowers new rehabilitation pathways for tetraplegic patients. SETTING: University Hospital-UNICAMP, Campinas, Brazil. METHODS: A total of 15 chronic tetraplegic patients and 10 healthy subjects were analyzed with the same methodology. Two tests were performed: (i) In test 1, the stimulus was applied to the right-arm radial nerve and the electromyography (EMG) signal collected in contralateral left tibial muscle. (ii) In test 2, the stimulus was applied to the left-leg fibular nerve and EMG collected in contralateral limb biceps, exploring the opposite direction of the pathway. In both tests, the subjects were stimulated with intensities from 5 to 30 mA (5 mA step) and 40 × 500 µs current modulated pulses. Reflexes were detected from the averaging of the 40 EMG sweeps. RESULTS: Each group was analyzed with regard to the reflexes' incidence, amplitude and latency. ILRs were found with similar prominence in both groups. A correlation between the ILR amplitude and the subject injury level was verified. Significant differences were found in the correlation of ILR latency with stimulation charge between healthy and tetraplegic subjects. CONCLUSION: The ILR transmission parameters of healthy and tetraplegic subjects were studied. The results obtained strongly suggest a different ILR transmission between healthy and tetraplegic subjects, reinforcing the hypothesis of nerve regeneration after injury.

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Considering the importance of bidimensional electrophoresis and image analysis in comparative proteomics, the parameters that influence the analysis of protein expression of transgenic and non-transgenic soybean seeds were evaluated. The loaded mass of the proteins (150-500 microg), the pH separation range (3-10 or 4-7), and manual/automatic image editing were evaluated. Additionally, after optimizing the conditions, histograms and matchings were obtained in order to accurately analyze the variations (90%) in protein expression. From this, 10 proteins displayed significant differences in expression, and eight of them were characterized and identified by mass spectrometry.

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During seedling establishment, cotyledons of the rain forest tree Hymenaea courbaril mobilize storage cell wall xyloglucan to sustain growth. The polysaccharide is degraded and its products are transported to growing sink tissues. Auxin from the shoot controls the level of xyloglucan hydrolytic enzymes. It is not yet known how important the expression of these genes is for the control of storage xyloglucan degradation. In this work, partial cDNAs of the genes xyloglucan transglycosylase hydrolase (HcXTH1) and beta-galactosidase (HcBGAL1), both related to xyloglucan degradation, and two other genes related to sucrose metabolism [alkaline invertase (HcAlkIN1) and sucrose synthase (HcSUS1)], were isolated. The partial sequences were characterized by comparison with sequences available in the literature, and phylogenetic trees were assembled. Gene expression was evaluated at intervals of 6 h during 24 h in cotyledons, hypocotyl, roots, and leaves, using 45-d-old plantlets. HcXTH1 and HcBGAL1 were correlated to xyloglucan degradation and responded to auxin and light, being down-regulated when transport of auxin was prevented by N-1-naphthylphthalamic acid (NPA) and stimulated by constant light. Genes related to sucrose metabolism, HcAlkIN1 and HcSUS1, responded to inhibition of auxin transport in consonance with storage mobilization in the cotyledons. A model is proposed suggesting that auxin and light are involved in the control of the expression of genes related to storage xyloglucan mobilization in seedlings of H. courbaril. It is concluded that gene expression plays a role in the control of the intercommunication system of the source-sink relationship during seeding growth, favouring its establishment in the shaded environment of the rain forest understorey.

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Solanum species are traditionally employed as antiherpes and anticancer agents in different countries. S. paniculatum has widespread ethnomedical uses in Brazil, including the treatment of viral infections. This paper reports on the isolation of neotigogenin (1) and the new compound delta25(27)-tigogenin-3-O-beta-D-glucopyranoside (2), obtained as a mixture of R and S diastereoisomers at C22 from an ethanol extract of S. paniculatum leaves, along with the determination of their cytotoxicity against Vero cells and antiviral effect against human herpes virus type 1 (HHV-1), murine encephalomyocarditis virus (EMCv), and vaccinia virus strain Western Reserve (VACV-WR). The extract of S. paniculatum inhibited HHV-1 replication [EC50 = (298.0 +/- 11.2) microg/ml] and showed no effect on EMCv and VACV-WR. On its turn, 1 was inactive against the assayed strains but presented high cytotoxicity [CC50 = (2.03 +/- 0.03) microg/ml], whereas 2 exhibited significant antiherpes [EC50 = (170.8 +/- 1.7) microg/ml] and antivaccinia virus effects [EC50 = (177.0 +/- 3.3) microg/ml], with low cytotoxicity (CC50 > 400 microg/ml). The results corroborate Solanum paniculatum as a source of cytotoxic and antiviral compounds.

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