Intact chloroplasts were isolated from Euglena gracilis variety bacillaris, and aliquots were exposed to several different chemical cross-linking reagents. The reagents penetrated the triple membrane of Euglena chloroplasts. This was shown by gradient acrylamide gel electrophoresis under denaturing conditions. The activity of the nonaggregated fatty acid synthetase of Euglena was located within the chloroplast stroma, and the effects of dimethylsuberimidate cross-linking on the activity of the enzyme system were examined. The acyl-carrier protein concentration in the chloroplast was measured at about 0.24 mM.

No description available.

Bio-based monomers are a more sustainable alternative to conventional oil-based monomers [1]. The bis-epoxide limonene dioxide from the epoxidation of the terpene limonene has shown potential for different applications [2]. One of those applications is the use of limonene dioxide as a crosslinking agent to improve the solvent resistance of nanofiltration membranes. Epoxidation of terpenes is conventionally done using meta-chloroperoxybenzoic acid (m-CPBA), using metal complexes with metals such as Tungsten, Titanium, and Cobalt, or different hydroperoxides. A greener method of epoxidation explored is the use of in situ generated dimethyldioxirane from the reaction of acetone and potassium peroxy-monosulfate (Oxone) [3]. The reaction uses sodium bicarbonate buffer in aqueous solution with a mixture of limonene and acetone. This project aims to synthesize different bis-, and tris-epoxides from different bio-derived terpenes including limonene, gamma-terpinene, geraniol, farnesol, and nerol using the reported method using Oxone and ultrasonication. Epoxidation using m-CPBA is also investigated to compare it to the Oxone method. In general, epoxidation using m-CPBA results in higher amount of epoxide, but the Oxone method presents a more sustainable alternative with good results. Successfully synthesized epoxides are used to crosslink polybenzimidazole nanofiltration membranes. Solvent testing in dimethylacetamide is used to inspect whether crosslinking is successful. Polyethylene glycol diglycidyl ether is a commercial bis-epoxide that was used to validate the crosslinking method. Crosslinking was successful, as confirmed by solvent testing and FT-IR analysis. Filtration testing showed that the permeance of the membrane was not affected by crosslinking, while the membrane’s rejection was increased from 10.29 ± 1.01 % to 17.23 ± 2.49 % after crosslinking using polyethylene glycol diglycidyl ether. Nerol and limonene bis-epoxides were successfully synthesized with high purity and were tested as crosslinkers. However, crosslinking was unsuccessful, as demonstrated by solvent testing. This project successfully synthesized bis-epoxides from different terpenes using a greener method of epoxidation. The possibility of successful crosslinking using the terpene-based crosslinkers should be further investigated.

3D bioprinting is an emerging field with the potential to reform the process of organ transplantation. The ability to 3D print new organs and tissues would supplement the organ donor shortage and decrease the risk associated with organ rejection. One of the current areas of research focuses on printing cells using hydrogels composed of methacrylated compounds as a scaffolding. One of the chemical means of crosslinking the hydrogels is using the photoinitiator lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) to crosslink with light. The 3D bioprinter in the lab currently has an attachment for a 365nm lamp, however this is cytotoxic to cells. A 405nm laser was designed to mount on the hot tool of the BioAssemblyBot by Advanced Solutions and flash at a specific frequency when sent a signal from the bioprinter. This tool was then tested to determine effective flash frequencies for crosslinking hydrogels.

No / The azinomycins are potent antitumour antibiotics that are able to crosslink DNA, but are relatively unstable and unlikely to progress as therapeutic candidates. A prototype analogue 4 with more clinical potential has been designed and synthesised and incorporates the epoxide function of the azinomycins and a nitrogen mustard. Two further analogues 5 and 6 that can alkylate DNA but cannot crosslink the duplex have also been synthesised. Compound 4 crosslinks DNA efficiently at nM concentrations. Compounds 4¿6 were submitted to the NCI 60 cell line screen and have similar antitumour activity, although 4 is slightly less active than the non-crosslinking compounds. These observations will be important in the design of further azinomycin analogues with antitumour activity.

Ziel der Studie: Evaluation der Effektivität und Sicherheit des Kollagen-Crosslinkings der Hornhaut mit Riboflavin und UV-A (CXL) bei progressivem Keratokonus über einen Nachbeobachtungszeitraum von bis zu 10 Jahren. Design: Retrospektive klinische Längsschnittstudie Methoden: 131 Augen von 131 Patienten (männlich:weiblich = 95:36) erhielten an der Universitäts-Augenklinik Würzburg zwischen 2006 und 2016 ein Kollagen-Crosslinking (CXL) nach dem Dresdner Standardprotokoll, bestehend aus einer Abrasio, Applikation von iso-osmolaren Riboflavin/Dextran-Augentropfen für 30 min und anschließender UV-A-Bestrahlung mit 3 mW/cm2 für 30 min. Die retrospektive Nachbeobachtung betrug 1 (n=103 Augen) bis maximal 10 Jahre (n=44 Augen). Einschlusskriterien waren eine Zunahme des maximalen Hornhautkrümmungsradius (Kmax, gemessen mittels Pentacam HR) >1 dpt und eine Hornhautdicke >400 µm nach Abrasio. Für parametrische bzw. nichtparametrische Daten wurde der T-Test bzw. der Wilcoxon-Vorzeichen-Rangsummentest durchgeführt. Ergebnisse: 1 bis 3 Jahre präoperativ nahm der Median von Kmax und K2 signifikant um 1,5 dpt (p=0,001) und 1,1 dpt (p<0,001) bis zur Behandlung zu. Die apikale Hornhautdicke nahm nach 1-3 Jahren präoperativ um 12 µm ab (p=0,003). Postoperativ stieg der Median von K2 zunächst bis nach 1 Jahr um 0,1 dpt, nahm dann aber über den weiteren Nachbeobachtungszeitraum kontinuierlich ab, nach 10 Jahren lag er um 0,85 dpt (p=0,021) unter dem Ausgangswert. Die mittlere apikale Hornhautdicke nahm nach 3, 7 bzw. 10 Jahren um 11 µm (p<0,001), 9 µm (p=0,014) und 3 µm (p=0,358) ab. Der mediane Kmax zeigte Schwankungen ohne signifikante Veränderung. Der mittlere bestkorrigierte Visus (logMAR) nahm nach 5 Jahren signifikant um 0,13 und nach 10 Jahren um 0,08 ab (p=0,013 und p=0,010). Der Anteil der Non-Responder, definiert durch einen postoperativen Anstieg von Kmax>2 dpt, nahm von 16% nach 5 auf 33% nach 10 Jahren zu. Risikofaktoren waren ein junges Alter, hoher Astigmatismus, eine dünne Hornhaut und atopische Dermatitis. 4 Augen erhielten im Verlauf komplikationslos eine Revernetzung, woraufhin sich keine weitere Krankheitsprogression zeigte. Fazit: Die CXL-Behandlung kann die Progression des Keratokonus verlangsamen oder stoppen. Allerdings war ab 5 Jahren nach dem Eingriff eine Abnahme des Anteils der Responder zu beobachten. Regelmäßige Nachkontrollen sollten daher besonders auch im Langzeitverlauf durchgeführt werden, um eine erneute Progression frühzeitig erkennen und behandeln zu können. / Purpose: This study analyses long-term efficacy and safety of corneal collagen crosslinking with riboflavin and UV-A irradiation (CXL) for progressive keratoconus. Design: Retrospective longitudinal study of consecutive patients. Methods: 131 eyes of 131 patients (male:female = 95:36) were treated at the university hospital Wuerzburg between 2006 and 2016 with standard CXL, comprising abrasion, application of iso-osmolar riboflavin/dextran eye drops for 30 min, and application of UV-A irradiation at 3 mW/cm2 for 30 min. Retrospective follow-up was 1 (n=103 eyes) to maximum 10 years (n=44 eyes). Only one eye per patient was included. Inclusion criteria were increase of maximum surface radius of curvature (Kmax measured with Pentacam HR) >1 D and corneal thickness >400 µm after abrasion. Paired t test or Wilcoxon matched-pairs signed rank test were conducted for parametric or nonparametric data, respectively. Results: 1 to 3 years preoperatively, median of Kmax and K2 significantly increased by 1.5 D (p=0.001) and 1.1 D (p<0.001). Apical corneal thickness diminished by 12 µm at 1-3 years preop (p=0.003). After CXL, median K2 increased by 0.1 D after 1 year, but then decreased over the remaining postoperative period by 0.85 D (p=0.021) after 10 years. Mean apical corneal thickness decreased by 11 µm (p<0.001), 9 µm (p=0.014) and 3 µm (p=0.358) after 3, 7 and 10 years, respectively. Median Kmax showed high variation without significant change. Mean best corrected visual acuity (logMAR) significantly decreased by 0.13 after 5 and 0,08 after 10 years (p=0.013 and p=0.010). CXL-non-responders, defined by postoperative increase of Kmax>2 D, increased from 16% after 5 to 33% after 10 years. Risk factors were young age, high astigmatism, thin cornea, and atopic dermatitis. 4 eyes were re-treated after first CXL without any complications and keratoconus stabilized thereafter. Conclusion: CXL can slow down or stop the progression of keratoconus. However, the declining responder rate beyond 5 years after treatment indicates that patients who have risk factors for being a non-responder might require re-treatment. So patients should be examined at regular intervals especially after 5 years post CXL, to recognize and re-treat a progression early.

El queratocono (QC) se trata de una enfermedad corneal caracterizada por un adelgazamiento progresivo de la córnea y ectasia, que conduce a un astigmatismo irregular y finalmente, a una pérdida de la agudeza visual. Aunque existen diferentes modalidades de tratamiento del QC, el único tratamiento que ha demostrado retrasar la progresión del QC es el crosslinking corneal (CXL), con el objetivo de detener la progresión del QC, fortaleciendo las propiedades biomecánicas de la córnea. A pesar de que el CXL estándar o convencional (CXL-S), sigue siendo el "gold standard", con las posibles complicaciones secundarias relacionadas con esta técnica, en la actual tesis se ha demostrado, que la capacidad de la técnica de CXL epi-on (epithelium-on, por sus siglas en inglés) de detener la progresión del QC es similar y comparable, a la técnica de CXL epi-off (epithelium off, por sus siglas en inglés). Además se ha observado que la mayoría de los artículos publicados, comparan diferenes técnicas de CXL en QC. Sin embargo, para optimizar el tratamiento de estos pacientes, es importante establecer una seria de factores predictivos preoperatorios de los cambios visuales tras el CXL, para así elegir la mejor técnica. De esta manera, en la actual tesis se ha comprobado los factores predictivos del cambio visual a tener en cuenta a priori en las córneas sometidas a CXL, epi-on y epi-off, y se ha comprobado que los factores difieren para ambas técnicas, sugiriendo que el mecanismo de acción es distinto entre ellas. Otra opción de tratamiento en pacientes con QC, son la implantación de los anillos corneales intraestromales (ACI), que reducen las aberraciones de alto orden, y secundariamente mejoran la calidad visual. Sin embargo, los estudios disponibles presentan una gran variabilidad, que podría ser explicada por diferentes nomogramas disponibles o los distintos criterios de implantación de los ACI. En la actual tesis se ha desarrollado un nomograma de implantación de ACI optimizado, para lograr mejores resultados y más predecibles. En los últimos años, el uso de las lentes de contacto esclerales (LCE) han adquirido gran importancia en el manejo del QC, para una adecuada rehabilitación visual. Sin embargo, existe una gran variabilidad entre pacientes por varios factores, como puede ser el vault de la LCE. En la actual tesis, se ha demostrado una buena eficacia con la LCE ICD 16.50, en lo que respecta a la rehabilitación visual y lo más importante es que ésta mejora en la AV, se ha podido predecir a partir de varios factores. En definitiva, la actual tesis ha sido clave para lograr una optimización del manejo del QC. Las técnicas de CXL epi-on y epi-off, presentan una capacidad similar para detener la progresión del QC; se han obtenido los factores predictivos del efecto del tratamiento del CXL y uso de LCE en el QC; y se ha desarrollado un nomograma optimizado para la implantación de los ACI.

No description available.

No description available.

Both the thermal and copper(I) catalyzed azide-alkyne Huisgen cycloadditions were explored as strategies to be used for silicone functionalization and crosslinking. The generality of these reactions was demonstrated (Chapter 2) using 1,3-bis(azidopropyl)tetramethyldisiloxane (BAPTMDS) as a model compound. The ligation of this compound with several alkyne-containing molecules, especially the copper(I) catalyzed process or “click” proved to be easy to perform, high yielding, and gave the 1,4-triazole regioisomer as the sole product. Thermal, metal catalyst-free, azide-alkyne cross-linking (Chapter 3) using a poly(azidopropylmethylsiloxane)-co-dimethylsiloxane as the base polymer and several polysubstituted alkyne molecules as crosslinkers was efficient. The reaction of the base polymer with an ethynyl-terminated disiloxane demonstrates that a silicone elastomer can be synthesized by simple heating and that the resulting material is stable, decomposing only at temperatures higher than 230 °C. Finally, direct bioconjugation of silicones to biotin using propargylamide and BAPTMDS was examined (Chapter 4). The result of the copper(I) catalyzed Huisgen ligation of biotin onto silicones was as efficient as the reactions in the previous chapters, revealing that the “click” process can successfully be applied to a broad range of silicones. / Thesis / Master of Science (MSc)