Supercritical fluid extraction of bioavailable amino acids from soils and their liquid chromatographic determination with fluorometric detection

A new procedure with supercritical CO2 modified with 0.5 mL of water and 0.75 mL of 0.1 M HCl in situ and 0.75 mL of water on-line at 15 MPa and 50 degrees C for 45 min was applied for the extraction of bioavailable amino acids from soil samples. Total extraction time was 60 min, but more favorable conditions are even possible for selected groups of amino acids. All analytes were trapped into 20 mL of methanol with satisfactory recovery (94-104%) and determined using high-performance liquid chromatography with fluorometric detection on a Zorbax Eclipse column (4.6 x 75 mm, 3.5 microm) with Na2HPO4 and acetonitrile/methanol/water as a mobile phase. Linear calibration curves were obtained (r > 0.999 except 0.99823 for Ile) with lower limits of detection (S/N = 3) in the range from 1.54 pg (Gly) to 13.5 pg (Cy2) or from 18.6 fmol (Ser) to 64.8 fmol (Lys). Validation and repeatability data are also given. Comparable results were obtained with a robust, commonly used extraction method (0.5 M ammonium acetate, 60 min in shaker, followed by filtration and lyophilization). Limiting values of artificial release of amino acids were also determined for each soil sample to eliminate any false results to ensure that all extracted amino acids originate from soil solution and exchangeable bound positions of soil samples.     

Effect of medetomidine-butorphanol and dexmedetomidine-butorphanol combinations on intraocular pressure in healthy dogs

ObjectiveTo assess the effects of intravenous (IV) medetomidine-butorphanol and IV dexmedetomidine-butorphanol on intraocular pressure (IOP).Study designProspective, randomized, blinded clinical study.AnimalsForty healthy dogs. Mean ± SD body mass 37.6 ± 6.6 kg and age 1.9 ± 1.3 years.MethodsDogs were allocated randomly to receive an IV combination of dexmedetomidine, 0.3 mg m^?2, combined with butorphanol, 6 mg m^?2, (group DEX) or medetomidine 0.3 mg m^?2, combined with butorphanol 6 mg m^?2, (group MED). IOP and pulse (PR) and respiratory (f_R) rates were measured prior to (baseline) and at 10 (T10), 20 (T20), 30 (T30) and 40 (T40) minutes after drug administration. Oxygen saturation of hemoglobin (SpO₂) was monitored following sedation. Data were analyzed by anova followed by Dunnett's tests for multiple comparisons. Changes were considered significant when p < 0.05.ResultsFollowing drug administration, PR and f_R were decreased significantly at all time points but did not differ significantly between groups. Baseline IOP in mmHg was 14 ± 2 for DEX and 13 ± 2 for MED. With both treatments, at T10, IOP increased significantly (p < 0.001), reaching 20 ± 3 and 17 ± 2 for DEX and MED respectively. This value for DEX was significantly higher than for MED. There were no significant differences in IOP values between groups at any other time points. At T30 and T40, IOP in both groups was below baseline (DEX, 12 ± 2 and 11 ± 2: MED 12 ± 2 and 11 ± 2) and this was statistically significant, for DEX.Conclusions and clinical relevanceAt the documented doses, both sedative combinations induced a transient increase and subsequent decrease of IOP relative to baseline, which must be taken into consideration when planning sedation of animals in which marked changes in IOP would be undesirable.     

家禽精液冷冻保存技术研究进展

家鸡精液冷冻保护是物种资源保存最廉价的方法。本文从家鸡精液的稀释液、冷冻保护剂、冻精形式、平衡时间、冷冻速率、解冻速率、精液质量检测方法等几个方面,综述了国内外研究进展,并对目前家鸡精液冷冻技术应用存在的问题进行了分析,提出了初步研究思路。     

Influence of Cow Age on Grazing Distribution in a Mixed-Conifer Forest

Optimal distribution of cattle on forested rangelands has long been a subject of concern specifically related to uniform and sustainable use of forage resources. Our objective was to determine if cow age influenced distribution and resource use on forested rangelands. This study was conducted from 1991 to 2001 at the US Department of Agriculture Starkey Experimental Forest and Range, northeastern Oregon, a mixed-conifer forested rangeland. We used 43 039 locations of cattle taken from 1 h prior to sunrise until 4 h after sunrise and 4 h prior to sundown until 1 h after sundown from 15 July to 30 August to evaluate cattle distribution patterns during peak foraging time. Cattle were grouped into four age classes: 2- and 3-yr-old cattle, 4- and 5-yr-old cattle, 6- and 7-yr-old cattle, and cattle ≥ 8 yr old. All age classes preferred areas with gentler slopes (P &spilt; 0.05), westerly aspects (P &spilt; 0.05), farther from water (P &spilt; 0.05), and with greater forage production (P &spilt; 0.05) than pasture averages. Cattle older than 3 yr of age selected areas with less canopy closure (P &spilt; 0.05) than the mean value for the pasture. Young cows (&spilt; 4 yr old) selected lower elevations and steeper slopes than the oldest cows (P &spilt; 0.05). In summary, cow age and correspondingly its experience directly influences distribution patterns and forage resource use of cattle at the Starkey Experimental Forest and Range.     

Incidence of seizures associated with iopamidol or iomeprol myelography in dogs with intervertebral disk disease: 161 cases (2000–2002)

Objective – To compare the incidence of seizures in dogs with intervertebral disk disease after iopamidol or iomeprol myelography, and to assess whether the incidence of seizures differed between the 2 agents when severity of neurological deficits, location of cord compression, duration of anesthesia, site of myelogram, volume of contrast, and concentration of contrast were evaluated. Design – Retrospective study. Setting – Veterinary teaching hospital. Animals – One hundred and sixty‐one client‐owned dogs with intervertebral disk disease. Interventions – Subarachnoid injection of contrast medium. Measurements and Main Results – One hundred and sixty‐one dogs with intervertebral disk disease were subjected to myelography using iopamidol (n=74) or iomeprol (n=87). Cranial myelography was performed in 31 dogs, caudal myelography in 125 and both cranial and caudal myelography in 5. Seizures occurred in 23 of 161 (14%) dogs. There was no significant difference overall between iopamidol and iomeprol myelography. However, in dogs with thoracolumbar disk extrusion and paraplegia, seizures occurred more frequently after caudal myelography using iopamidol compared with iomeprol. Conclusions – Both iomeprol and iopamidol are suitable for myelography in dogs. Iomeprol is recommended for caudal myelography in paraplegic dogs with thoracolumbar disk extrusion due to the higher incidence of seizures in this group when iopamidol was used.     

Production of UV-light-detectable faeces from house mice (Mus musculus domesticus) after consumption of encapsulated fluorescent pigment in monitoring bait

BACKGROUND: The authors investigated whether fluorescent pigment in thermoset melamine microcapsules incorporated into monitoring baits would be excreted in the faeces of wild house mice in a quantity and intensity that would be detectable by a human observer. RESULTS: Experimental mice produced 24–116 UV‐visible faecal pellets per 24 h; the mean dry weight was 582 mg. The number and weight of the faeces was independent of mouse sex and weight. The defecation of UV‐visible faeces began at 2–3 h, peaked at 5–8 h and was complete at 17 h after bait ingestion. The detectability of the highly fluorescent faecal pellets using a small UV flashlight approached 100%, and no false positives were recorded. CONCLUSION: The tested formulation is of significant value for rodent pest monitoring because faeces that are highly visible by UV light are produced for 15 h by mice after ingestion, and their detection is easy and unambiguous. Copyright © 2011 Society of Chemical Industry     

Efficacy and Mechanism of Action of Marine Alkaloid 3,10-Dibromofascaplysin in Drug-Resistant Prostate Cancer Cells

Efficacy and mechanism of action of marine alkaloid 3,10-dibromofascaplysin (DBF) were investigated in human prostate cancer (PCa) cells harboring different levels of drug resistance. Anticancer activity was observed across all cell lines examined without signs of cross-resistance to androgen receptor targeting agents (ARTA) or taxane based chemotherapy. Kinome analysis followed by functional investigation identified JNK1/2 to be one of the molecular targets of DBF in 22Rv1 cells. In contrast, no activation of p38 and ERK1/2 MAPKs was observed. Inhibition of the drug-induced JNK1/2 activation or of the basal p38 activity resulted in increased cytotoxicity of DBF, whereas an active ERK1/2 was identified to be important for anticancer activity of the alkaloid. Synergistic effects of DBF were observed in combination with PARP-inhibitor olaparib most likely due to the induction of ROS production by the marine alkaloid. In addition, DBF intensified effects of platinum-based drugs cisplatin and carboplatin, and taxane derivatives docetaxel and cabazitaxel. Finally, DBF inhibited AR-signaling and resensitized AR-V7-positive 22Rv1 prostate cancer cells to enzalutamide, presumably due to AR-V7 down-regulation. These findings propose DBF to be a promising novel drug candidate for the treatment of human PCa regardless of resistance to standard therapy.     

Structure Elucidation and Functional Studies of a Novel β-hairpin Antimicrobial Peptide from the Marine Polychaeta Capitella teleta

Endogenous antimicrobial peptides (AMPs) are evolutionary ancient molecular factors of innate immunity that play a key role in host defense. Among the most active and stable under physiological conditions AMPs are the peptides of animal origin that adopt a β-hairpin conformation stabilized by disulfide bridges. In this study, a novel BRICHOS-domain related AMP from the marine polychaeta Capitella teleta, named capitellacin, was produced as the recombinant analogue and investigated. The mature capitellacin exhibits high homology with the known β-hairpin AMP family—tachyplesins and polyphemusins from the horseshoe crabs. The β-hairpin structure of the recombinant capitellacin was proved by CD and NMR spectroscopy. In aqueous solution the peptide exists as monomeric right-handed twisted β-hairpin and its structure does not reveal significant amphipathicity. Moreover, the peptide retains this conformation in membrane environment and incorporates into lipid bilayer. Capitellacin exhibits a strong antimicrobial activity in vitro against a wide panel of bacteria including extensively drug-resistant strains. In contrast to other known β-hairpin AMPs, this peptide acts apparently via non-lytic mechanism at concentrations inhibiting bacterial growth. The molecular mechanism of the peptide antimicrobial action does not seem to be related to the inhibition of bacterial translation therefore other molecular targets may be assumed. The reduced cytotoxicity against human cells and high antibacterial cell selectivity as compared to tachyplesin-1 make it an attractive candidate compound for an anti-infective drug design.     

Study of Structure–Activity Relationships of the Marine Alkaloid Fascaplysin and Its Derivatives as Potent Anticancer Agents

Marine alkaloid fascaplysin and its derivatives are known to exhibit promising anticancer properties in vitro and in vivo. However, toxicity of these molecules to non-cancer cells was identified as a main limitation for their clinical use. Here, for the very first time, we synthesized a library of fascaplysin derivatives covering all possible substituent introduction sites, i.e., cycles A, C and E of the 12H-pyrido[1-2-a:3,4-b’]diindole system. Their selectivity towards human prostate cancer versus non-cancer cells, as well as the effects on cellular metabolism, membrane integrity, cell cycle progression, apoptosis induction and their ability to intercalate into DNA were investigated. A pronounced selectivity for cancer cells was observed for the family of di- and trisubstituted halogen derivatives (modification of cycles A and E), while a modification of cycle C resulted in a stronger activity in therapy-resistant PC-3 cells. Among others, 3,10-dibromofascaplysin exhibited the highest selectivity, presumably due to the cytostatic effects executed via the targeting of cellular metabolism. Moreover, an introduction of radical substituents at C-9, C-10 or C-10 plus C-3 resulted in a notable reduction in DNA intercalating activity and improved selectivity. Taken together, our research contributes to understanding the structure–activity relationships of fascaplysin alkaloids and defines further directions of the structural optimization.