Zn promoted GaZrOx Ternary Solid Solution Oxide Combined with SAPO-34 Effectively Converts CO2 to Light Olefins with Low CO Selectivity.

We proposed a new strategy for CO2 hydrogenation to prepare light olefins by introducing Zn into GaZrOx to construct ZnGaZrOx ternary oxides, which was combined with SAPO-34 to prepare a high-performance ZnGaZrOx/SAPO-34 tandem catalyst for CO2 hydrogenation to light olefins. By optimizing the Zn doping content, the ratio and mode of the two-phase composite, and the process conditions, the 3.5 %ZnGaZrOx/SAPO-34 tandem catalyst showed excellent catalytic performance and good high-temperature inhibition of the reverse water-gas shift (RWGS) reaction. The catalyst achieved 26.6 % CO2 conversion, 82.1 % C2 =-C4 = selectivity and 11.8 % light olefins yield. The ZnGaZrOx formed by introducing an appropriate amount of Zn into GaZrOx significantly enhanced the spillover H2 effect and also induced the generation of abundant oxygen vacancies to effectively promote the activation of CO2. Importantly, the RWGS reaction was also significantly suppressed at high temperatures, with the CO selectivity being only 46.1 % at 390 °C.

Prevalence and Rapid Screen Method of Diagnostic Criteria for Psychosomatic Research Syndromes in Human Papillomavirus-Infected Patients.

INTRODUCTION: The early and rapid identification of psychosomatic symptoms is crucial to prevent harmful outcomes in patients with human papillomavirus (HPV) infection in busy comprehensive clinics. This study aimed to explore the prevalence and rapid screening method of the Diagnostic Criteria for Psychosomatic Research-revised (DCPR) syndromes in patients with HPV infection. METHODS: A total of 504 participants underwent a clinical assessment that included DCPR, Diagnostic and Statistical Manual of Mental Disorders, fifth edition (DSM-5), the Social Support Rating Scale (SSRS), the Simplified Coping Style Questionnaire (SCSQ), fear of disease, sociodemographic and clinical characteristics. The prevalence of DCPR syndromes and DSM-5 diagnoses were compared between the HPV-positive and negative patients using χ2 tests. We explored the rapid screen indicator through multiple logistic regression analyses of the participants' psychosocial factors, sociodemographic and clinical characteristics. RESULTS: The incidence of DCPR syndromes in HPV-positive patients (56.6%) was significantly greater than that in HPV-negative patients (17.3%) and DSM-5 diagnoses (8.5%) in the HPV-positive group. Health anxiety, irritable mood, type A behavior, and demoralization were the most common psychosomatic syndromes in HPV-positive patients. As the degree of fear increased from 0 to 5 to 10, the risk of DCPR increased from 1.27 (95% CI: 0.21-7.63) to 3.24 (score range: 1-5, 95% CI: 1.01-10.39) to 9.91 (score range: 6-10, 95% CI: 3.21-30.62) in the HPV-positive group. CONCLUSION: The degree of fear, as an independent risk factor, could be used to quickly screen outpatients with a high risk of DCPR syndrome among women with HPV infection.

Clinical Interviewing: An Essential but Neglected Method of Medicine.

Clinical interviewing is the basic method to understand how a person feels and what are the presenting complaints, obtain medical history, evaluate personal attitudes and behavior related to health and disease, give the patient information about diagnosis, prognosis, and treatment, and establish a bond between patient and physician that is crucial for shared decision making and self-management. However, the value of this basic skill is threatened by time pressures and emphasis on technology. Current health care trends privilege expensive tests and procedures and tag the time devoted to interaction with the patient as lacking cost-effectiveness. Instead, the time spent to inquire about problems and life setting may actually help to avoid further testing, procedures, and referrals. Moreover, the dialogue between patient and physician is an essential instrument to increase patient's motivation to engage in healthy behavior. The aim of this paper was to provide an overview of clinical interviewing and its optimal use in relation to style, flow and hypothesis testing, clinical domains, modifications according to settings and goals, and teaching. This review points to the primacy of interviewing in the clinical process. The quality of interviewing determines the quality of data that are collected and, eventually, of assessment and treatment. Thus, interviewing deserves more attention in educational training and more space in clinical encounters than it is currently receiving.

Protective Effect of Que Zui Tea on d-Galactose-Induced Oxidative Stress Damage in Mice via Regulating SIRT1/Nrf2 Signaling Pathway.

Que Zui tea (QT) is an important herbal tea in the diet of the 'Yi' people, an ethnic group in China, and it has shown significant antioxidant, anti-inflammatory, and hepatoprotective effects in vitro. This study aims to explore the protective effects of the aqueous-ethanol extract (QE) taken from QT against ᴅ-galactose (ᴅ-gal)-induced oxidative stress damage in mice and its potential mechanisms. QE was identified as UHPLC-HRMS/MS for its chemical composition and possible bioactive substances. Thus, QE is rich in phenolic and flavonoid compounds. Twelve compounds were identified, the main components of which were chlorogenic acid, quinic acid, and 6'-O-caffeoylarbutin. Histopathological and biochemical analysis revealed that QE significantly alleviated brain, liver, and kidney damage in ᴅ-gal-treated mice. Moreover, QE remarkably attenuated oxidative stress by activating the Nrf2/HO-1 pathway to increase the expression of antioxidant indexes, including GSH, GSH-Px, CAT, SOD, and T-AOC. In addition, QE administration could inhibit the IL-1ß and IL-6 levels, which suppress the inflammatory response. QE could noticeably alleviate apoptosis by inhibiting the expressions of Caspase-3 and Bax proteins in the brains, livers, and kidneys of mice. The anti-apoptosis mechanism may be related to the upregulation of the SIRT1 protein and the downregulation of the p53 protein induced by QE in the brain, liver, and kidney tissues of mice. Molecular docking analysis demonstrated that the main components of QE, 6'-O-caffeoylarbutin, chlorogenic acid, quinic acid, and robustaside A, had good binding ability with Nrf2 and SIRT1 proteins. The present study indicated that QE could alleviate ᴅ-gal-induced brain, liver and kidney damage in mice by inhibiting the oxidative stress and cell apoptosis; additionally, the potential mechanism may be associated with the SIRT1/Nrf2 signaling pathway.

Quality and Flavor Difference in Dry-Cured Meat Treated with Low-Sodium Salts: An Emphasis on Magnesium.

The present study aimed to develop low-sodium curing agents for dry-cured meat products. Four low-sodium formulations (SPMA, SPM, SP, and SM) were used for dry-curing meat. The physicochemical properties and flavor of the dry-cured meat were investigated. The presence of Mg2+ ions hindered the penetration of Na+ into the meat. The weight loss, moisture content, and pH of all low-sodium salt groups were lower than those of S. Mg2+ addition increased the water activity (Aw) of SPMA, SPM, and SM. Dry-curing meat with low-sodium salts promoted the production of volatile flavor compounds, with Mg2+ playing a more prominent role. Furthermore, low-sodium salts also promoted protein degradation and increased the content of free amino acids in dry-cured meat, especially in SM. Principal component analysis (PCA) showed that the low-sodium salts containing Mg2+ were conducive to improving the quality of dry-cured meat products. Therefore, low-sodium salts enriched with Mg2+ become a desirable low-sodium curing agent for achieving salt reduction in dry-cured meat products.

Chemical Constituents, Hypolipidemic, and Hypoglycemic Activities of Edgeworthia gardneri Flowers.

The flowers of Edgeworthia gardneri are used as herbal tea and medicine to treat various metabolic diseases including hyperglycemia, hypertension, and hyperlipidemia. This paper investigate the chemical constituents and biological activities of ethanolic extract and its different fractions from E. gardneri flowers. Firstly, the E. gardneri flowers was extracted by ethanol-aqueous solution to obtain crude extract (CE), which was subsequently fractionated by different polar organic solution to yield precipitated crystal (PC), dichloromethane (DCF), ethyl acetate (EAF), n-butanol (n-BuF), and residue water (RWF) fractions. UHPLC-ESI-HRMS/MS analysis resulted in the identification of 25 compounds, and the main compounds were flavonoids and coumarins. The precipitated crystal fraction showed the highest phenolic and flavonoid contents with 344.4 ± 3.38 mg GAE/g extract and 305.86 ± 0.87 mg RE/g extract. The EAF had the strongest antioxidant capacity and inhibitory effect on α-glucosidase and pancreatic lipase with IC50 values of 126.459 ± 7.82 and 23.16 ± 0.79 µg/mL. Besides, both PC and EAF significantly regulated the glucose and lipid metabolism disorders by increasing glucose consumption and reducing TG levels in HepG2 cells. Molecular docking results suggested that kaempferol-3-O-glucoside and tiliroside had good binding ability with enzymes, indicating that they may be potential α-glucosidase and pancreatic lipase inhibitors. Therefore, the E. gardneri flowers could be served as a bioactive agent for the regulation of metabolic disorders.

Phytochemicals from Anneslea fragrans Wall. and Their Hepatoprotective and Anti-Inflammatory Activities.

Anneslea fragrans Wall., popularly known as "Pangpo tea", is an edible, medicinal, and ornamental plant of the Family Theaceae. The leaves of A. fragrans were historically applied for the treatment of liver and intestinal inflammatory diseases in China. This study aimed to explore the hepatoprotective agents from A. fragrans leaves through hepatoprotective and anti-inflammatory assessment. The phytochemical investigation of the leaves of A. fragrans resulted in the isolation and identification of a total of 18 chemical compounds, including triterpenoids, aliphatic alcohol, dihydrochalcones, chalcones, flavanols, phenolic glycoside, and lignans. Compounds 1-2, 4-6, 11-12, and 16-18 were identified from A. fragrans for the first time. Compounds 7 and 14 could significantly alleviate hepatocellular damage by decreasing the contents of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) and inhibit the hepatocellular apoptosis in the HepG2 cells induced by N-acetyl-p-aminophenol (APAP). In addition, compounds 7 and 14 inhibited reactive oxygen species (ROS) and malondialdehyde (MDA) contents and increased the catalase (CAT) superoxide dismutase (SOD), and glutathione (GSH) levels for suppressing APAP-induced oxidative stress. Additionally, compounds 7, 13, and 14 also had significant anti-inflammatory effects by inhibiting interleukin-6 (IL-6), interleukin-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α) productions on LPS-induced RAW246.7 cells.

Influence of Ultra-High-Pressure Pretreatment Method on Chemical Constituents, Antioxidant and Cytoprotective Activities of Free, Esterified, and Bound Phenolics from Anneslea Fragrans Wall. Leaves.

Anneslea fragrans Wall., an edible and medicinal plant, is traditionally used to treat liver and gastrointestinal diseases. This paper aimed to investigate the influence of ultra-high pressure (UHP) pretreatment on the phenolics profiling, antioxidant, and cytoprotective activities of free (FP), esterified (EP), and bound (BP) phenolics from A. fragrans leaves. A total of 32 compounds were characterized and quantified. The davidigenin (44.46 and 113.37 mg/g extract) was the highest in A. fragrans leaves. The vitexin (9), afzelin (10), coreopsin (15), and davidigenin (28) were analyzed with MS2 fragment pathways. Results showed that UHP treated A. fragrans leaves had higher total phenolic (TPC) and total flavonoid (TFC) contents of FP, EP, and BP fractions than those in the raw leaves. Moreover, UHP pretreated A. fragrans leaves had higher scavenging activities on DPPH+• and ABTS+•, and inhibitory effects on the intracellular ROS generation in H2O2-induced HepG2 cells. UFP showed the highest inhibition of ROS production among the samples. Therefore, UHP pretreatment method might be used as an effective strategy for elevating the availabilities of A. fragrans leaves to develop functional foods.

Anti-leukemic effect and molecular mechanism of 11-methoxytabersonine from Melodinus cochinchinensis via network pharmacology, ROS-mediated mitochondrial dysfunction and PI3K/Akt signaling pathway.

Melodinus cochinchinensis (Lour.) Merr. is a Yunnan endemic folk medicine. Our previous study showed that 11-methoxytabersonine (11-MT) isolated from M. cochinchinensis has strong cytotoxicity on human T-ALL cells, but its molecular mechanism has not been studied. In current study, the cytotoxicity and possible mechanism of 11-MT on T-cell acute lymphoblastic leukemia was explored using network pharmacology and molecular biology techniques. 11-MT significantly inhibited the cell proliferations on different four human T-ALL cells (MOLT-4, Jurkat, CCRF-CEM, and CEM/C1 cells). 11-MT triggered ROS accumulation, calcium concentration and cell apoptosis, and decreased the mitochondrial membrane potential (MMP) in human T-ALL cells, especially MOLT-4 cells. Western blot analysis showed that it can induce MOLT-4 cell apoptosis by up-regulating PI3K/Akt signaling pathway. Therefore, 11-MT induces human T-ALL cells apoptosis via up-regulation of ROS-mediated mitochondrial dysfunction and down-regulation of PI3K/Akt/mTOR signaling pathway.

Elsholtzia bodinieri Vaniot Ameliorated Acute Lung Injury by NQO1, BCL2 and PTGS2 In Silico and In Vitro Analyses.

Acute lung injury (ALI) is a clinical respiratory disease caused by various factors, which lacks effective pharmacotherapy to reduce the mortality rate. Elsholtzia bodinieri Vaniot is an annual herbaceous plant used as a traditional herbal tea and folk medicine. Here we used bioinformatic databases and software to explore and analyze the potential key genes in ALI regulated by E. bodinieri Vaniot, including B cell leukemia/lymphoma 2 (Bcl2), prostaglandin-endoperoxide synthase 2 (Ptgs2) and NAD(P)H dehydrogenase, quinone 1 (Nqo1). In an inflammatory cells model, we verified bioinformatics results, and further mechanistic analysis showed that methanol extract of E. bodinieri Vaniot (EBE) could alleviate oxidative stress by upregulating the expression of NQO1, suppress pyroptosis by upregulating the expression of BCL2, and attenuate inflammation by downregulating the expression of PTGS2. In sum, our results demonstrated that EBE treatment could alleviate oxidative stress, suppress pyroptosis and attenuate inflammation by regulating NQO1, BCL2 and PTGS2 in a cells model, and E. bodinieri Vaniot might be a promising source for functional food or as a therapeutic agent.

Phenolic Constituents, Antioxidant and Cytoprotective Activities, Enzyme Inhibition Abilities of Five Fractions from Vaccinium dunalianum Wight.

The bud of Vaccinium dunalianum Wight has been traditionally consumed as health herbal tea by "Yi" people in Yunnan Province, China, which was locally named "Que Zui tea". This paper studied the chemical constituents of five fractions from Vaccinium dunalianum, and their enzyme inhibitory effects of α-glucosidase and pancreatic lipase, antioxidant activity, and cytoprotective effects on H2O2-induced oxidative damage in HepG2 cells. The methanol extract of V. dunalianum was successively partitioned with petroleum ether (PF), chloroform (CF), ethyl acetate (EF), n-butanol (BF), and aqueous (WF) to obtain five fractions. The chemical profiling of the five fractions was analyzed by ultra-high-performance liquid chromatography coupled with a tandem mass spectrometry (UHPLC-MS/MS), and 18 compounds were tentatively identified. Compared to PF, CF, BF and WF, the EF revealed the highest total phenols (TPC) and total flavonoids (TFC), and displayed the strongest enzyme inhibition ability (α-glucosidase and pancreatic lipase) and antioxidant capacity (DPPH, ABTS and FRAP). Furthermore, these five fractions, especially EF, could effectively inhibit reactive oxygen species (ROS) production and cell apoptosis on H2O2-induced oxidative damage protection in HepG2 cells. This inhibitory effect might be caused by the up-regulation of intracellular antioxidant enzyme activity (CAT, SOD, and GSH). The flavonoids and phenolic acids of V. dunalianum might be the bioactive substances responsible for enzyme inhibitory, antioxidant, and cytoprotective activities.

Synthetic Aperture Radar Image Despeckling Based on Multi-Weighted Sparse Coding.

Synthetic aperture radar (SAR) images are inherently degraded by speckle noise caused by coherent imaging, which may affect the performance of the subsequent image analysis task. To resolve this problem, this article proposes an integrated SAR image despeckling model based on dictionary learning and multi-weighted sparse coding. First, the dictionary is trained by groups composed of similar image patches, which have the same structural features. An effective orthogonal dictionary with high sparse representation ability is realized by introducing a properly tight frame. Furthermore, the data-fidelity term and regularization terms are constrained by weighting factors. The weighted sparse representation model not only fully utilizes the interblock relevance but also reflects the importance of various structural groups in despeckling processing. The proposed model is implemented with fast and effective solving steps that simultaneously perform orthogonal dictionary learning, weight parameter updating, sparse coding, and image reconstruction. The solving steps are designed using the alternative minimization method. Finally, the speckles are further suppressed by iterative regularization methods. In a comparison study with existing methods, our method demonstrated state-of-the-art performance in suppressing speckle noise and protecting the image texture details.

Clinimetric Criteria for Patient-Reported Outcome Measures.

Patient-reported outcome measures (PROMs) are self-rated scales and indices developed to improve the detection of the patients' subjective experience. Given that a considerable number of PROMs are available, it is important to evaluate their validity and usefulness in a specific research or clinical setting. Published guidelines, based on psychometric criteria, do not fit in with the complexity of clinical challenges, because of their quest for homogeneity of components and inadequate attention to sensitivity. Psychometric theory has stifled the field and led to the routine use of scales widely accepted yet with a history of poor performance. Clinimetrics, the science of clinical measurements, may provide a more suitable conceptual and methodological framework. The aims of this paper are to outline the major limitations of the psychometric model and to provide criteria for clinimetric patient-reported outcome measures (CLIPROMs). The characteristics related to reliability, sensitivity, validity, and clinical utility of instruments are critically reviewed, with particular reference to the differences between clinimetric and psychometric approaches. Of note is the fact that PROMs, rating scales, and indices developed according to psychometric criteria may display relevant clinimetric properties. The present paper underpins the importance of the clini-metric methodology in choosing the appropriate PROMs. CLIPROM criteria may also guide the development of new indices and the validation of existing PROMs to be employed in clinical settings.

4,21-Secovincanol, a Novel Immunosuppressive Monoterpenoid Indole Alkaloid from Kopsia Hainanensis.

4,21-Secovincanol (1), a novel C-21/N-4 cleavage monoterpenoid indole alkaloid, along with four analogs (2-5), were obtained from the aerial parts of Kopsia hainanensis. Structurally, compound 1 might be a derivative of epivincanol (2) via C-21/N-4 cleavage. Their structures were confirmed by means of comprehensive spectroscopic data analysis and comparison with the reported data. All isolates significantly inhibited Con A-stimulated mice splenocytes proliferation at 10-40 µM in a dose-dependent manner in vitro. Especially, compound 3 exhibited potent activities comparable to positive control (Dexamethasone, DXM).

The Bioavailability, Extraction, Biosynthesis and Distribution of Natural Dihydrochalcone: Phloridzin.

Phloridzin is an important phytochemical which was first isolated from the bark of apple trees. It is a member of the dihydrochalcones and mainly distributed in the plants of the Malus genus, therefore, the extraction method of phloridzin was similar to those of other phenolic substances. High-speed countercurrent chromatography (HSCCC), resin adsorption technology and preparative high-performance liquid chromatography (HPLC) were used to separate and purify phloridzin. Many studies showed that phloridzin had multiple pharmacological effects, such as antidiabetic, anti-inflammatory, antihyperglycaemic, anticancer and antibacterial activities. Besides, the physiological activities of phloridzin are cardioprotective, neuroprotective, hepatoprotective, immunomodulatory, antiobesity, antioxidant and so on. The present review summarizes the biosynthesis, distribution, extraction and bioavailability of the natural compound phloridzin and discusses its applications in food and medicine.

Activity Guided Isolation of Phenolic Compositions from Anneslea fragrans Wall. and Their Cytoprotective Effect against Hydrogen Peroxide Induced Oxidative Stress in HepG2 Cells.

Anneslea fragrans Wall., commonly known as "Pangpo Tea", is traditionally used as a folk medicine and healthy tea for the treatment of liver and intestine diseases. The aim of this study was to purify the antioxidative and cytoprotective polyphenols from A. fragrans leaves. After fractionation with polar and nonpolar organic solvents, the fractions of aqueous ethanol extract were evaluated for their total phenolic (TPC) and flavonoid contents (TFC) and antioxidant activities (DPPH, ABTS, and FRAP assays). The n-butanol fraction (BF) showed the highest TPC and TFC with the strongest antioxidant activity. The bio-guided chromatography of BF led to the purification of six flavonoids (1-6) and one benzoquinolethanoid (7). The structures of these compounds were determined by NMR and MS techniques. Compound 6 had the strongest antioxidant capacity, which was followed by 5 and 2. The protective effect of the isolated compounds on hydrogen peroxide (H2O2)-induced oxidative stress in HepG2 cells revealed that the compounds 5 and 6 exhibited better protective effects by inhibiting ROS productions, having no significant difference with vitamin C (p > 0.05), whereas 6 showed the best anti-apoptosis activity. The results suggest that A. fragrans could serve as a valuable antioxidant phytochemical source for developing functional food and health nutraceutical products.

Bioactivity-Guided Isolation of Phytochemicals from Vaccinium dunalianum Wight and Their Antioxidant and Enzyme Inhibitory Activities.

Vaccinium dunalianum Wight, usually processed as a traditional folk tea beverage, is widely distributed in the southwest of China. The present study aimed to investigate the antioxidant, α-glucosidase and pancreatic lipase inhibitory activities of V.dunalianum extract and isolate the bioactive components. In this study, the crude extract (CE) from the buds of V. dunalianum was prepared by the ultrasound-assisted extraction method in 70% methanol and then purified with macroporous resin D101 to obtain the purified extract (PM). Five fractions (Fr. A-E) were further obtained by MPLC column (RP-C18). Bioactivity assays revealed that Fr. B with 40% methanol and Fr. D with 80% methanol had better antioxidant with 0.48 ± 0.03 and 0.62 ± 0.01 nM Trolox equivalent (TE)/mg extract for DPPH, 0.87 ± 0.02 and 1.58 ± 0.02 nM TE/mg extract for FRAP, 14.42 ± 0.41 and 19.25 ± 0.23 nM TE/mg extract for ABTS, and enzyme inhibitory effects with IC50 values of 95.21 ± 2.21 and 74.55 ± 3.85 for α-glucosidase, and 142.53 ± 11.45 and 128.76 ± 13.85 µg/mL for pancreatic lipase. Multivariate analysis indicated that the TPC and TFC were positively related to the antioxidant activities. Further phytochemical purification led to the isolation of ten compounds (1-10). 6-O-Caffeoylarbutin (7) showed significant inhibitory effects on α-glucosidase and pancreatic lipase enzymes with values of 38.38 ± 1.84 and 97.56 ± 7.53 µg/mL, and had the highest antioxidant capacity compared to the other compounds.

Optimization of isoxazoline amide benzoxaboroles for identification of a development candidate as an oral long acting animal ectoparasiticide.

Novel isoxazoline amide benzoxaboroles were designed and synthesized to optimize the ectoparasiticide activity of this chemistry series against ticks and fleas. The study identified an orally bioavailable molecule, (S)-N-((1-hydroxy-3,3-dimethyl-1,3-dihydrobenzo[c][1,2]oxaborol-6-yl)methyl)-2-methyl-4-(5-(3,4,5-trichlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)benzamide (23), with a favorable pharmacodynamics profile in dogs (Cmax=7.42ng/mL; Tmax=26.0h; terminal half-life t1/2=127h). Compound 23, a development candidate, demonstrated 100% therapeutic effectiveness within 24h of treatment, with residual efficacy of 97% against American dog ticks (Dermacentor variabilis) on day 30 and 98% against cat fleas (Ctenocephalides felis) on day 32 after a single oral dose at 25mg/kg in dogs.

Discovery of an orally bioavailable isoxazoline benzoxaborole (AN8030) as a long acting animal ectoparasiticide.

A novel series of isoxazoline benzoxaborole small molecules was designed and synthesized for a structure-activity relationship (SAR) investigation to assess the ectoparasiticide activity against ticks and fleas. The study identified an orally bioavailable molecule, (S)-3,3-dimethyl-5-(5-(3,4,5-trichlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)benzo[c][1,2]oxaborol-1(3H)-ol (38, AN8030), which was long lasting in dogs (t1/2=22 days). Compound 38 demonstrated 97.6% therapeutic effectiveness within 24 h of treatment, with residual efficacy of 95.3% against American dog ticks (Dermacentor variabilis) on day 30% and 100% against cat fleas (Ctenocephalides felis) on day 32 after a single oral dose at 50 mg/kg in dogs.