Enzymkatalysierte C-C Knüpfung : Reaktionstechnische Untersuchungen zur Synthese pharmazeutischer Intermediate

In dieser Arbeit werden Möglichkeiten untersucht, durch enzymkatalysierte asymmetrische C C-Knüpfung, relevante pharmazeutische Intermediate zu synthetisieren. Das Ziel ist es die Synthese unterschiedlicher Produkte durch die Benzaldehydlyase (BAL) reaktionstechnisch zu charakterisieren. Anschließend werden die erhaltenen Daten zur Auswahl und Optimierung geeigneter Reaktorkonzepte verwendet und die Produkte im Gramm-Maßstab hergestellt. Im ersten Teil der Arbeit wird die Kopplung von teilweise substituierten aromatischen Aldehyden zu Benzoin(derivaten) untersucht. Aus der Messung der Anfangsreaktions-geschwindigkeiten werden die kinetischen Parameter der ausgewählten Reaktionen bestimmt und anschließend durch Simulation verschiedener Reaktionsverläufe ergänzt. Die Substrate und Produkte der BAL sind gering wasserlöslich. Es werden daher geeignete Reaktionsbedingungen untersucht die Löslichkeit der Reaktanden sowie die Substratverfügbarkeit zu verbessern. Die Untersuchungen werden im Hinblick auf technologische Anforderungen unter realen Bedingungen durchgeführt. Somit werden die Enzymstabilität und dafür relevante Faktoren wie Temperatur und Phasengrenzflächen untersucht und in der Simulation berücksichtigt. Für die Benzoinsynthese im Zwei-Phasensystem eignet sich das Substrat als organische Phase. Die Produktabtrennung durch gezielte Kristallisation ist für die Benzoinsynthese die Methode der Wahl [100]. Aus den Reaktoren werden mit hoher Enantioselektivität (> 99 %) (R)-Benzoin, (R)-3,3'-Dimethoxy-benzoin und (R)-3-Methoxy-2'-chlor-benzoin als Produkte im Gramm-Maßstab erhalten. Im zweiten Teil der Arbeit wird die enzymkatalysierte Synthese von 2-Hydroxy-1-phenylpropanon-derivaten (HPP) aus aromatischen und aliphatischen Aldehyden charakterisiert. Zunächst wird eine geeignete Analytik zur Untersuchung des Reaktionsverlaufes entwickelt und das als Substrat benötigte Methoxyacetaldehyd synthetisiert. Die für die Arbeit ausgewählten Reaktionen werden reaktionstechnisch charakterisiert. Die erhaltenen kinetischen Parameter werden anhand von Reaktorläufen im Satzreaktor, membrangestützten Zwei-Phasensystem und im kontinuierlichen Enzym-Membran-Reaktor überprüft. Aus den realisierten Reaktoren werden die Produkte (R)-3-Chlor-HPP, (R)-3,4'-Dimethoxy-HPP und (R)-3,3-Dimethoxy-HPP im Gramm-Maßstab erhalten. Für (R)-3-Chlor-HPP wird die Folgesynthese zum pharmazeutisch wirksamen Hydroxybupropion gezeigt. Diese Arbeit liefert ausführliche Informationen über die enantioselektive C C-Knüpfung mittels enzymatischer Katalyse. Etablierte chemische Verfahren werden für die Enzymkatalyse zu einem technologischen Konzept zusammengeführt, welches auf unterschiedlich substituierte Produkte übertragbar ist. Die Simulation der Prozessabläufe auf Basis der empirischen Modellbildung trägt entscheidend zum Verständnis des Gesamtprozesses bei.Enzyme catalysed C-C bond formation : Reaction engineering for the synthesis of pharmaceutical intermediates This thesis investigates feasibilities to synthesise relevant pharmaceutical intermediates by enzyme catalysed asymmetric C-C bond formation. Therefore the synthesis of several products formed by the Benzaldehyde lyase (BAL) are characterised by means of reaction engineering. Subsequent the resulting data are used for choosing and optimizing appropriate reactor concepts and the preparation of the products on a gram scale. The first part of this work deals with the coupling of partially substituted aromatic aldehydes to Benzoin (-derivatives). The kinetic parameters of the chosen reactions are determined by initial rate measurement and completed by simulation of various reactor experiments. The substrates and products of the BAL are low soluble in water. Hence suitable reaction conditions are studied to improve the solubility of the reactants and the availability of the substrate during the reaction. The investigations are conducted with respect to the specific technological requirements under realistic conditions. Consequently the stability of the enzyme and crucial factors like temperature and phase interfaces are investigated and included into the simulation. For the synthesis of Benzoin (-derivatives) a two-phase-system with the substrate as organic phase is shown to be a suitable approach. The separation of the product by well directed crystallisation is the method of choice [100]. From the reactor experiments (R)-Benzoine, (R)-3,3'-Dimethoxy-benzoine and (R)-3-Methoxy-2'-chloro-benzoine are obtained with high enantioselectivity (> 99 %) on a multi gram scale. In the second part of the thesis characterises the enzyme catalysed synthesis of 2-Hydroxy-1-phenylpropanone-derivatives (HPP) starting from aromatic and aliphatic aldehydes. In a first step an analytic method for the investigation of the reaction progress is developed and the required substrate Methoxyacetaldehyde is synthesised. Thereafter the selected reactions are characterised by means of reaction engineering. The obtained kinetic parameters are confirmed with experiments in batch reactor, membrane based two-phase-system and in the continuous enzyme-membrane-reactor. The realised reactors supply the products (R)-3-Chloro-HPP, (R)-3,4'-Dimethoxy-HPP ad (R)-3,3-Dimethoxy-HPP on a gram scale. For (R)-3-Chloro-HPP a follow-up synthesis to the pharmaceutical active Hydroxybupropion is shown. This dissertation contains comprehensive information on the enantioselective C-C-bond formation by enzymatic catalysis. Classical chemical processes are combined for the enzyme catalysis to a technological concept, which is applicable for the synthesis of various substituted products. The simulation of the reaction progress, based on an empirical model development, is an essential contribution to the understanding of the whole process

Quantifying Stratospheric Chlorine Chemistry by the Satellite Spectrometers GOME and SCIAMACHY

Since the mid eighties of the past century, an almost complete destruction of the ozone layer above the Antarctic in springtime (and to a lesser extent also above the Arctic) is observed, which is caused by the release of atomic chlorine and bromine from anthropogenic halogen compounds. Due to the fast phase out of the production of CFCs, the increase of the stratospheric chlorine loading could be slowed down, and the recovery of the ozone layer is expected around the year 2050. However, the impacts on atmospheric chemistry, the climate and the biosphere are considerable. By differential optical absorption spectroscopy (DOAS) various molecules with characteristic absorption spectra can be detected simultaneously in the atmosphere, among them also many halogen compounds. In this thesis, measurements of the satellite spectrometers GOME and SCIAMACHY were evaluated for the atmospheric trace gas chlorine dioxide (OClO) and analysed with respect to their relevance for the polar stratospheric ozone chemistry. OClO is an important indicator for stratospheric chlorine activation, the conversion of the mainly anthropogenic chlorine from its ozone- inert reservoir compounds into ozone destroying (active) species. This activation of chlorine takes place by heterogeneous reactions on polar stratospheric clouds, which may be formed under the extremely cold stratospheric temperatures prevailing in polar nights. In this thesis, measurements of the OClO column density by GOME and SCIAMACHY during the polar winters 1995-2005 are quantitatively related to the meteorological conditions, the effect on the ozone column and the interaction with other stratospheric trace gases. The measurements by SCIAMACHY in the limb geometry can be inverted to vertical profiles by calculation of the radiative transfer, which increases the information content with respect to a measurement of the column density essentially. In this thesis, algorithms for the retrieval of vertical profiles of OClO, ozone, BrO and NO2 were developed and first results are presented

First Results on the DOAS-Retrieval of OClO from SCIAMACHY Nadir Measurements

The Scanning Imaging Absorption Spectrometer for Atmospheric Chartography was launched successfully onboard ENVISAT on March 1, 2002. It observes solar radiation transmitted, backscattered from the atmosphere and reflected from the ground in nadir, limb and occultation viewing modes. Chlorinedioxide (OClO), an important indicator for stratospheric chlorine activation, can be measured in the UV spectral range by Differential Optical Absorption Spectroscopy (DOAS). First results of the DOAS retrieval of OClO slant column densities (SCDs) from the SCIAMACHY measurements are presented. The influence of several parameters like the wavelength range chosen as fitting window or the reference spectra included in the fit on the quality of the retrieval is examined. It is found that a proper correction of polarisation features in the spectra is essential for a good DOAS analysis of OClO. The OClO SCDs derived from SCIAMACHY are compared to measurements of the Global Ozone Monitoring Experiment (GOME) which has successfully measured OClO since 1995. SCIAMACHY flies in the same orbit, but measures approx. 30 minutes earlier than GOME. As OClO shows a strong diurnal variation, this leads to differences in the observed column densities, which may be useful to investigate the photochemistry of OClO and related compounds. Also, the spatial resolution of SCIAMACHY is higher (30*60 km^2 compared to 40*320 km^2 for GOME), which will allow a more detailed study of small scale effects like e.g. chlorine activation in mountain waves

Total body irradiation as part of conditioning regimens in childhood leukemia—long-term outcome, toxicity, and secondary malignancies

Background: Total body irradiation (TBI) is an established part of conditioning regimens prior to stem cell transplantation in childhood leukemia but is associated with long-term toxicity. We retrospectively analyzed survival, long-term toxicity, and secondary malignancies in a pooled cohort of pediatric patients (pts.) treated with the same TBI regimen. Methods: Analyzed were 109 pts. treated between September 1996 and November 2015. Conditioning treatment according to EBMT guidelines and the ALL SCTped 2012 FORUM trial consisted of chemotherapy (CT) and TBI with 2 Gy b.i.d. on 3 consecutive days to a total dose of 12 Gy. Median follow-up was 97.9 months (2-228 months). Results: Overall survival (OS) in our cohort at 2, 5, and 10 years was 86.1, 75.5, and 63.0%, respectively. Median survival was not reached. Long-term toxicity developed in 47 pts. After chronically abnormal liver and kidney parameters in 31 and 7 pts., respectively, growth retardation was the most frequent finding as seen in 13 pts. Secondary malignancies were rare (n = 3). Conclusion: TBI-containing conditioning regimens in pediatric stem cell transplantation (SCT) are highly effective. Efforts to replace TBI- with CT-containing regimens have only been successful in subgroups of pts. Although we could show long-term toxicity in 43% of pts., overall survival was 63% at 10 years. Still, long-term effects such as growth retardation can permanently impact the pts.' quality of life and functioning. Along with new substances, efforts should be undertaken to optimize TBI techniques and accompany the treatment by systematic follow-up programs beyond 5 years to improve detection of rare events

Long-term small-fiber neuropathy and pain sensitization in survivors of pediatric acute lymphoblastic leukemia after stem cell transplantation

Purpose: We aimed at describing for the first time peripheral small-fiber neurotoxicity and pain sensitization in survivors of pediatric acute lymphoblastic leukemia after stem cell transplantation (SCT). Methods: In a cross-sectional, retrospective, single-center study, we assessed 25 relapse-free long-term survivors (median age at SCT: 11 ± 4.9 years; median time between SCT and testing: 8.25 years, 19 males) using a reduced version of the pediatric-modified total neuropathy score for clinical assessment and Quantitative Sensory Testing (QST). Inclusion criteria: ≥ 6 years old at testing, ≤ 18 years old at time of SCT, ≥ 1 year between SCT and testing. Results: Nine patients (36%) had peripheral neuropathy as defined by the clinical red-pmTNS (≥ 4). The QST parameters mechanical pain sensitivity, mechanical detection threshold, thermal sensory limen, vibration detection threshold and pressure pain threshold were significantly abnormal in the survivor cohort (p < 0.0038). Except for one, all survivors showed at least one abnormal QST parameter. When using QST, signs of small and large fiber dysfunction were present in 22 (88%) and 17 (68%) survivors, respectively. More than half of all survivors were found to experience pathologic sensitization to pain. Conclusions and implications for cancer survivors: Survivors of pediatric acute lymphoblastic leukemia after SCT are at high risk for long-term peripheral neuropathy with a dominating small-fiber and pain sensitization pattern

Allogeneic hematopoietic stem cell transplantation from sibling and unrelated donors in pediatric patients with sickle cell disease—A single center experience

HSCT is curative in SCD. Patients with HLA-identical sibling donor have an excellent outcome ranging from 90%-100% overall and event-free survival. However, due to the lack of matched sibling donors this option is out of reach for 70% of patients with SCD. The pool of potential donors needs to be extended. Transplantations from HLA-matched unrelated donors were reported to be less successful with shorter event-free survival and higher incidences of complications including graft-vs-host disease, especially in patients with advanced stage SCD. Here we report transplantation outcomes for 25 children with SCD transplanted using HLA-matched grafts from related or unrelated donors. Overall survival was 100% with no severe (grade III-IV) graft-vs-host disease and a 12% rejection rate. Mixed donor chimerisms only occurred in transplantations from siblings, while transplantations from unrelated donors resulted in either complete donor chimerism or rejection. Despite the small patient number, overall and disease-free survival for unrelated donor transplantations is excellent in this cohort. The advanced disease state, higher alloreactive effect and stronger immunosuppression in unrelated donor transplantations raises patient risk, for which possible solutions could be found in optimization of transplant preparation, graft manipulation or haploidentical transplantation using T cell receptor α/β-depleted grafts

Application of 233 nm far-UVC LEDs for eradication of MRSA and MSSA and risk assessment on skin models

A newly developed UVC LED source with an emission wavelength of 233 nm was proved on bactericidal efficacy and skin tolerability. The bactericidal efficacy was qualitatively analysed using blood agar test. Subsequently, quantitative analyses were performed on germ carrier tests using the MRSA strain DSM11822, the MSSA strain DSM799, S. epidermidis DSM1798 with various soil loads. Additionally, the compatibility of the germicidal radiation doses on excised human skin and reconstructed human epidermis was proved. Cell viability, DNA damage and production of radicals were assessed in comparison to typical UVC radiation from discharge lamps (222 nm, 254 nm) and UVB (280-380 nm) radiation for clinical assessment. At a dose of 40 mJ/cm(2), the 233 nm light source reduced the viable microorganisms by a log(10) reduction (LR) of 5 log(10) levels if no soil load was present. Mucin and protein containing soil loads diminished the effect to an LR of 1.5-3.3. A salt solution representing artificial sweat (pH 8.4) had only minor effects on the reduction. The viability of the skin models was not reduced and the DNA damage was far below the damage evoked by 0.1 UVB minimal erythema dose, which can be regarded as safe. Furthermore, the induced damage vanished after 24 h. Irradiation on four consecutive days also did not evoke DNA damage. The radical formation was far lower than 20 min outdoor visible light would cause, which is classified as low radical load and can be compensated by the antioxidant defence system

Исследование влияния частоты и длительности импульсов на свойства покрытий, осаждаемых на титановые сплавы методом микродугового оксидирования

Объектом исследования являлись кальций-фосфатные покрытия, сформированные методом микродугового оксидирования, при различных специально подобранных режимах работы. Целью данной работы являлось исследование влияния частоты следования и длительности импульсов на физико-химические и механические свойства покрытий, сформированных методом микродугового оксидирования. В результате исследования были выявлены утверждения, которые могут быть полезны при подборе режимов для формирования специальных покрытий для разного рода применений.The object of the study is calcium-phosphate cotaing formed by microarc oxidation method on different parameters. The aim of the work is to study the effect of the repetition frequency and pulse duration on the physical, chemical and mechanical properties of coatings formed by microarc oxidation. As a result of the research, statements were identified that can be useful in selecting modes for forming special coatings for various types of applications

Feasibility and initial experience of assessment of mechanical dyssynchrony using cardiovascular magnetic resonance and semi-automatic border detection

<p>Abstract</p> <p>Background</p> <p>The systolic dyssynchrony index (SDI) has been introduced as a measure of mechanical dyssynchrony using three-dimensional echocardiography to select patients who may benefit from cardiac resynchronization therapy (CRT). However, three-dimensional echocardiography may be inadequate in a number of patients with suboptimal acoustic window and no single echocardiographic measure of dyssynchrony has proven to be of value in selecting patients for CRT. Thus, the aim of this study was to determine the value of cardiovascular magnetic resonance (CMR) for the assessment of the SDI in patients with reduced LV function as well as in healthy controls using semi-automatic border tracking.</p> <p>Methods</p> <p>We investigated a total of 45 patients including 35 patients (65 ± 8 years) with reduced LV function (EF 30 ± 11%) and a wide QRS complex as well as 10 control subjects (42 ± 21 years, EF 70 ± 11%). For cine imaging a standard SSFP imaging sequence was used with a temporal resolution of 40 frames per RR-interval. Quantitative analysis was performed off-line using a software prototype for semi-automatic border detection. Global volumes, ejection fraction and the SDI were calculated in each subject. SDI was compared with standard echocardiographic parameters of dyssynchrony.</p> <p>Results</p> <p>The mean SDI differed significantly between patients (14 ± 5%) and controls (5 ± 2%, p < 0.001). An exponential correlation between the EF and the SDI was observed (r = -0.84; p < 0.001). In addition, a significant association between the SDI and the standard deviation of time to peak systolic motion of 12 LV segments (Ts-SD) determined by echocardiography was observed (r = 0.66, p = 0.002).</p> <p>Conclusion</p> <p>The results of this preliminary study suggest that CMR with semi-automatic border detection may be useful for the assessment of mechanical dyssynchrony in patients with reduced LV function.</p> <p>No trial registration due to recruitment period between October 2004 and November 2006</p