Isolation and structure determination of oxidative degradation products of atorvastatin

Methods were developed for the preparation and isolation of four oxidative degradation products of atorvastatin. ATV-FX1 was prepared in the alkaline acetonitrile solution of atorvastatin with the addition of hydrogen peroxide. The exposition of aqueous acetonitrile solution of atorvastatin to sunlight for several hours followed by the alkalization of the solution with potassium hydroxide to pH 8–9 gave ATV-FXA. By the acidification of the solution with phosphoric acid to pH 3 ATV-FXA1 and FXA2 were prepared. The isolation of oxidative degradation products was carried out on a reversed-phase chromatographic column Luna prep C18(2) 10 μm applying several separation steps. The liquid chromatography coupled with a mass spectrometer (LC-MS), high resolution MS (HR-MS), 1D and 2D NMR spectroscopy methods were applied for the structure elucidation. All degradants are due to the oxidation of the pyrrole ring. The most probable reaction mechanism is intermediate endoperoxide formation with subsequent rearrangement and nucleophilic attack by the 5-hydroxy group of the heptanoic fragment. ATV-FX1 is 4-[1b-(4-Fluoro-phenyl)-6-hydroxy-6-isopropyl-1a-phenyl-6a-phenylcarbamoyl-hexahydro-1,2-dioxa-5a-aza-cyclopropa[a]inden-3-yl]-3-(R)-hydroxy-butyric acid and has a molecular mass increased by two oxygen atoms with regard to atorvastatin. ATV-FXA is the regioisomeric compound, 4-[6-(4-Fluoro-phenyl)-6-hydroxy-1b-isopropyl-6a-phenyl-1a-phenylcarbamoyl-hexahydro-1,2-dioxa-5a-aza-cyclopropa[a]inden-3-yl]-3-(R)-hydroxy-butyric acid. Its descendants ATV-FXA1 and FXA2 appeared without the atorvastatin heptanoic fragment and are 3-(4-Fluoro-benzoyl)-2-isobutyryl-3-phenyl-oxirane-2-carboxylic acid phenylamide and 4-(4-Fluoro-phenyl)-2,4-dihydroxy-2-isopropyl-5-phenyl-3,6-dioxa-bicyclo[3.1.0]hexane-1-carboxylic acid phenylamide, respectively. Quantitative NMR spectroscopy was employed for the assay determination of isolated oxidative degradation products. The results obtained were used for the determination of the UV response factors relative to atorvastatin.     

Effects of ghrelin on the feeding behavior of Lymantria dispar L. (Lymantriidae) caterpillars

Ghrelin is a 28-amino acid peptide which has significant effects on animal appetite, thus influencing body mass. The aim of our study was to examine the effects of ghrelin on the feeding behavior and physiology 4th instar caterpillars of the pest insect, Lymantria dispar L. Treatment of 4th instar caterpillars with four subpicomolar amounts of ghrelin had a positive influence on daily food intake, frass elimination, body mass. Also, locomotor activity increased, while stadium duration decreased in treated caterpillars. The similarity between the effects of ghrelin on caterpillar physiology and those in mammals suggests that using this model system for further studies of neuroendocrinological processes underlying feeding could lead to essential information about more complex organisms.     

Standard citrate versus sequential citrate/anticoagulant-free anticoagulation during hemodialysis: a randomized trial

Abstract:  In a randomized study, sequential anticoagulation for hemodialysis (citrate for the first 3.5 h, switching to 30-min anticoagulation-free hemodialysis) was compared to standard citrate anticoagulation. Fifty-two hemodialysis procedures were randomized either to sequential ( n  = 27) or standard citrate group ( n  = 25). The antithrombotic effect in the circuit was visually assessed after hemodialysis using a score from 1 (total clotting) to 5 (no clotting). The antithrombotic score for sequential versus standard group was as follows: dialyzer, 4.0 ± 1.1 versus 4.8 ± 0.4 ( P  < 0.01); arterial bubble trap, 4.0 ± 1.2 versus 4.7 ± 0.6 ( P  = 0.013); venous bubble trap, 4.0 ± 1.3 versus 4.8 ± 0.6 ( P  < 0.01). Serum citrate levels during sequential versus standard citrate anticoagulation (µmol/L) were as follows: at the beginning, 143 ± 65 versus 148 ± 77 (not significant [NS]); after 2 h, 317 ± 157 versus 354 ± 111 (NS); at the end, 125 ± 81 versus 405 ± 133 ( P  < 0.01). Sequential anticoagulation reduces the final serum citrate concentration to predialysis level. It can be a good anticoagulation strategy for patients in whom the reduction of citrate load is desired.     

A quantitative PCR screening method for adeno-associated viral vector 2-mediated gene doping

Gene therapy is currently prohibited in human and equine athletes and novel analytical methods are needed for its detection. Most in vivo products use non-integrating, recombinant viral vectors derived from adeno-associated virus (AAV) to deliver transgenes into cells, where they are transcribed and translated into functional proteins. Although the majority of wild-type AAV (WTAAV) DNA is removed from recombinant AAV (rAAV) vectors, some sequences are conserved. The goal of this study was to develop a quantitative polymerase chain reaction (QPCR) screening test targeting conserved AAV sequences to enable theoretical detection of all rAAV gene therapy products, regardless of encoded transgenes while excluding the presence of WTAAV DNA in horses. Primer sets were developed and validated to target an AAV2 sequence highly conserved across rAAV viral vectors and a sequence only found in wild type AAV2 (WTAAV2). Six horses were administered an intra-articular injection of rAAV. Plasma and synovial fluid were collected on days 0, 1, 2, 4, 7, 14, 28, 56, and 84. Using QPCR, rAAV was detected in plasma for up to 2–4 days in all horses. rAAV DNA was detected for 28 days in synovial fluid from two horses for which synovial fluid samples were available. No WTAAV2 DNA was detected in any sample. This is the first study to develop a QPCR test capable of screening for rAAV vectors that may be used for gene doping in horses.     

Countertransference in the therapy of suicidal patients - an important part of integrative treatment

One of the central issues in the psychotherapy of suicidal patients is the countertransference. Key concepts in countertransference include projective identification, role-responsiveness and countertransference enactment. It is important to recognize that countertransference can be effective in understanding the emotional intensity of the suicidal person's internal world. There is a significant relationship between treatment outcome and the different countertransference feelings among the therapists. In this paper we have illustrated important factors for understanding psychiatrist's countertransference reactions when working with suicidal patients.     

Depression in dermatology: an integrative perspective

An example which confirmes the adequacy of the integrative perspective in medicine is depression in dermatology because of the sufficient number of arguments linking depression and some chronic skin conditions in depth more than simply comorbidity. In recent literature, the authors found, how depression in dermatology occurs significantly more frequently than in the general population. In dermatological patients the prevalence of depression is around 30% which is more in comparison to patients in general practice where prevalence of depression is 22%. The authors found a considerable similarity in the various characteristics between depression and psoriasis, based upon recent research findings in immunity disorder and elevated concentrations of proinflammatory cytokines as well as acute phase proteins in both disorders, indicating that both disorders can be considered as immunologicaly mediated, inflammatory states with repetitive chronic progress and similar comorbidity. The afore mentioned theoretical settings evoke the integrative aspect and the integrative interdisciplinary approach placing in the center of attention not only the diseased person with his fears, needs, preocupations and expectations during treatment, but the person who is at risk of becoming burdened with these disorders.     

Predicting carotid restenosis by comparison of plaque MCP-1 mRNA expression and serum levels

As cardiovascular pathology grows in numbers, research into the discovery of new chemokine biomarkers should not be neglected, as they seem to be paramount in atherosclerosis prevention and its early detection. Chemokines attract and activate leukocytes and are well recognized in the environment of inflammatory response. MCP-1 is a valuable chemokine whose potential to become a new crucial atherosclerosis marker is surely worth investigating. Since quantities of MCP-1 found in lesions are as low as immeasurable, we propose the use of an immunohistochemical method for the quantification of MCP-1 levels in atherosclerotic lesions. Additionally, serum levels of MCP-1 can be measured by commercially available immunoassays. Proposed MCP-1 concentration increase may explain the acceleration in lesion’s atherosclerosis progression as chemokine activation occurs once they bind to specific ligands. If proven, this hypothesis would indicate the need for further studies in order to objectively link the increased MCP-1 expression with carotid restenosis.