TY - JOUR A1 - Endrullat, Christoph A1 - Glökler, Jörn A1 - Franke, Philipp A1 - Frohme, Marcus T1 - Standardization and quality management in next-generation sequencing JF - Applied & Translational Genomics N2 - DNA sequencing continues to evolve quickly even after > 30 years. Many new platforms suddenly appeared and former established systems have vanished in almost the same manner. Since establishment of next-generation sequencing devices, this progress gains momentum due to the continually growing demand for higher throughput, lower costs and better quality of data. In consequence of this rapid development, standardized procedures and data formats as well as comprehensive quality management considerations are still scarce. Here, we listed and summarized current standardization efforts and quality management initiatives from companies, organizations and societies in form of published studies and ongoing projects. These comprise on the one hand quality documentation issues like technical notes, accreditation checklists and guidelines for validation of sequencing workflows. On the other hand, general standard proposals and quality metrics are developed and applied to the sequencing workflow steps with the main focus on upstream processes. Finally, certain standard developments for downstream pipeline data handling, processing and storage are discussed in brief. These standardization approaches represent a first basis for continuing work in order to prospectively implement next-generation sequencing in important areas such as clinical diagnostics, where reliable results and fast processing is crucial. Additionally, these efforts will exert a decisive influence on traceability and reproducibility of sequence data. KW - next-generation sequencing KW - quality management KW - data quality KW - standardization KW - validation KW - guideline Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-6637 SN - 2212-0661 VL - 10 SP - 2 EP - 9 ER - TY - JOUR A1 - Tillich, Ulrich M. A1 - Wolter, Nick A1 - Schulze, Katja A1 - Kramer, Dan A1 - Brödel, Oliver A1 - Frohme, Marcus T1 - High-throughput cultivation and screening platform for unicellular phototrophs JF - BMC Microbiology N2 - In this work we describe and test high-throughput methods with the model organism Synechocystis sp. PCC6803. The required technical automation for these processes was achieved with a Tecan Freedom Evo 200 pipetting robot. The cultivation was performed in 2.2 ml deepwell microtiter plates within a cultivation chamber outfitted with programmable shaking conditions, variable illumination, variable temperature, and an adjustable CO2 atmosphere. Each microtiter-well within the chamber functions as a separate cultivation vessel with reproducible conditions. The automated measurement of various parameters such as growth, full absorption spectrum, chlorophyll concentration, MALDI-TOF-MS, as well as a novel vitality measurement protocol, have already been established and can be monitored during cultivation. Measurement of growth parameters can be used as inputs for the system to allow for periodic automatic dilutions and therefore a semi-continuous cultivation of hundreds of cultures in parallel. The system also allows the automatic generation of mid and long term backups of cultures to repeat experiments or to retrieve strains of interest. KW - cyanobacteria KW - synechocystis KW - HTS KW - high throughput KW - automated cultivation Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-6007 SN - 1471-2180 VL - 14 IS - 239 ER - TY - JOUR A1 - Kipkorir, R. A1 - Muhoho, S. A1 - Muliro, P. A1 - Mugendi, Beatrice A1 - Frohme, Marcus A1 - Brödel, Oliver T1 - Effects of Coffee Processing Technologies on Aroma Profiles and Sensory Quality of Ruiru 11 and SL 28 Kenyan Coffee Varieties JF - Asian Journal of Agriculture and Food Sciences N2 - The study aimed at comparing the effects of three coffee processing methods on aroma profiles and sensory quality of Ruiru 11 and SL 28 coffee varieties. The processing methods varied on stages of processing and method of mucilage removal. The green coffee beans obtained from the three processing methods were graded and roasted, ground and analyzed for the aroma profiles and sensory quality. Headspace Solid phase Microextraction fibre (SPME) technique were used for the extraction of aroma compounds from coffee samples and characterization of the compounds with use of gas chromatography mass spectrometry (GC-MS). Sensory quality were analyzed by an expert panelist. Various volatile aroma compounds were identified in roasted coffee and classified into their chemical classes involving furans, ketones, pyrazines ketones pyridines, pyrroles and acids. The intensity of aroma compounds were compared in terms of their peak areas and variations were noted between the processing methods with the ecopulper showing higher levels of pyrazines such as 2-methylpyrazine, 2-ethyl-6-methylpyrazine, and wet pulper showing higher levels of furans such as furfuryl formate and furfuryl alcohol, acetate. The hand pulper was high in the level of acids and esters such as acetic acid and propanoic acid, ethyl ester. Similar aroma compounds were identified in headspace of Ruiru 11 and SL 28 and there were variations in the intensities of aroma groups such as pyrroles and pyridines. It was concluded that the eco-pulper and hand pulper methods gives better aroma quality than the wet pulper while SL 28 variety gives higher sensory quality than Ruiru 11. KW - coffee KW - processing KW - aroma KW - sensory quality Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-6482 UR - https://ajouronline.com/index.php/AJAFS/article/view/2551 SN - 2321-1571 VL - 3 IS - 2 SP - 178 EP - 188 ER - TY - JOUR A1 - Radivoievych, Aleksandar A1 - Prylutska, Svitlana A1 - Zolk, Oliver A1 - Ritter, Uwe A1 - Frohme, Marcus A1 - Grebinyk, Anna T1 - Comparison of Sonodynamic Treatment Set-Ups for Cancer Cells with Organic Sonosensitizers and Nanosonosensitizers JF - Pharmaceutics N2 - Cancer sonodynamic therapy (SDT) is the therapeutic strategy of a high-frequency ultrasound (US) combined with a special sonosensitizer that becomes cytotoxic upon US exposure. The growing number of newly discovered sonosensitizers and custom US in vitro treatment solutions push the SDT field into a need for systemic studies and reproducible in vitro experimental set-ups. In the current research, we aimed to compare two of the most used and suitable SDT in vitro set-ups—“sealed well” and “transducer in well”—in one systematic study. We assessed US pressure, intensity, and temperature distribution in wells under US irradiation. Treatment efficacy was evaluated for both set-ups towards cancer cell lines of different origins, treated with two promising sonosensitizer candidates—carbon nanoparticle C60 fullerene (C60) and herbal alkaloid berberine. C60 was found to exhibit higher sonotoxicity toward cancer cells than berberine. The higher efficacy of sonodynamic treatment with a “transducer in well” set-up than a “sealed well” set-up underlined its promising application for SDT in vitro studies. The “transducer in well” set-up is recommended for in vitro US treatment investigations based on its US-field homogeneity and pronounced cellular effects. Moreover, SDT with C60 and berberine could be exploited as a promising combinative approach for cancer treatment. KW - ultrasound KW - C60 fullerene KW - berberine KW - sonodynamic therapy KW - apoptosis Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-18223 SN - 1999-4923 VL - 15 IS - 11 PB - MDPI ER - TY - JOUR A1 - Kagel, Heike A1 - Bier, Frank Fabian A1 - Frohme, Marcus A1 - Glökler, Jörn T1 - A Novel Optical Method To Reversibly Control Enzymatic Activity Based On Photoacids JF - Scientific Reports N2 - Most biochemical reactions depend on the pH value of the aqueous environment and some are strongly favoured to occur in an acidic environment. A non-invasive control of pH to tightly regulate such reactions with defined start and end points is a highly desirable feature in certain applications, but has proven difficult to achieve so far. We report a novel optical approach to reversibly control a typical biochemical reaction by changing the pH and using acid phosphatase as a model enzyme. The reversible photoacid G-acid functions as a proton donor, changing the pH rapidly and reversibly by using high power UV LEDs as an illumination source in our experimental setup. The reaction can be tightly controlled by simply switching the light on and off and should be applicable to a wide range of other enzymatic reactions, thus enabling miniaturization and parallelization through non-invasive optical means. Y1 - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-12685 SN - 2045-2322 VL - 9 ER - TY - JOUR A1 - Amro, Ahmad A1 - Moskalenko, Olga A1 - Hamarsheh, Omar A1 - Frohme, Marcus T1 - Spatiotemporal analysis of cutaneous leishmaniasis in Palestine and foresight study by projections modelling until 2060 based on climate change prediction JF - PLoS ONE N2 - Background Cutaneous leishmaniasis (CL) is a vector-borne parasitic diseases of public health importance that is prevalent in the West Bank but not in the Gaza Strip. The disease caused by parasitic protozoans from the genus Leishmania and it is transmitted by infected phlebotomine sand flies. The aim of our study is to investigate the eco-epidemiological parameters and spatiotemporal projections of CL in Palestine over a 30-years period from 1990 through 2020 and to explore future projections until 2060. Methodology/Principal findings This long-term descriptive epidemiological study includes investigation of demographic characteristics of reported patients by the Palestinian Ministry of Health (PMoH). Moreover, we explored spatiotemporal distribution of CL including future projection based on climate change scenarios. The number of CL patients reported during this period was 5855 cases, and the average annual incidence rate (AAIR) was 18.5 cases/105 population. The male to female ratio was 1.25:1. Patients-age ranged from 2 months to 89 years (mean = 22.5, std 18.67, and the median was 18 years). More than 65% of the cases came from three governates in the West Bank; Jenin 29% (1617 cases), Jericho 25% (1403), and Tubas 12% (658) with no cases reported in the Gaza Strip. Seasonal occurrence of CL starts to increase in December and peaked during March and April of the following year. Current distribution of CL indicate that Jericho, Tubas, Jenin and Nablus have the most suitable climatic settings for the sandfly vectors. Future projections until 2060 suggest an increasing incidence from northwest of Jenin down to the southwest of Ramallah, disappearance of the foci in Jericho and Tubas throughout the Jordan Vally, and possible emergence of new foci in Gaza Strip. Conclusions/Significance The future projection of CL in Palestine until 2060 show a tendency of increasing incidence in the north western parts of the West Bank, disappearance from Jericho and Tubas throughout the Jordan Vally, and emergence of new CL endemic foci in the Gaza Strip. These results should be considered to implement effective control and surveillance systems to counteract spatial expansion of CL vectors. Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-16166 SN - 1932-6203 VL - 17 IS - 6 PB - Public Library of Science (PLoS) ER - TY - JOUR A1 - Grebinyk, Anna A1 - Yashchuk, Valeriy A1 - Bashmakova, Nataliya A1 - Gryn, Dmytro A1 - Hagemann, Tobias A1 - Naumenko, Antonina A1 - Kutsevol, Nataliya A1 - Dandekar, Thomas A1 - Frohme, Marcus T1 - A new triple system DNA-Nanosilver-Berberine for cancer therapy JF - Applied Nanoscience N2 - The isoquinoline quaternary alkaloid Berberine possesses a variety of pharmacological properties that suggests its promising application for an anticancer delivery system design utilizing its ability to intercalate DNA. In the current work, we have investigated the effects of Berberine on the human T cell leukemia cell line in vitro. Fluorescent microscopy of leukemic cells revealed Berberine nuclear localization. The results showed that Berberine inhibited leukemic cell growth in a time- and dose-dependent manner, that was associated with reactive oxygen species production intensification and caspase 3/7 activity increase with followed apoptosis induction. Berberine was used as a toxic and phototoxic agent for triple system synthesis along with DNA as a carrier and nanosilver as a plasmonic accelerator of Berberine electronic transitions and high energy emission absorbent centers. The proposed method allows to obtain the complex of DNA with Berberine molecules and silver nanoparticles. The optical properties of free components as well as their various combinations, including the final triple system DNA-Nanosilver-Berberine, were investigated. Obtained results support the possibility to use the triple system DNA-Nanosilver-Berberine as an alternative therapeutic agent for cancer treatment. Y1 - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-16105 SN - 2190-5517 VL - 9 SP - 945 EP - 956 PB - Springer ER - TY - JOUR A1 - Grebinyk, Anna A1 - Prylutska, Svitlana A1 - Grebinyk, Sergii A1 - Evstigneev, Maxim A1 - Krysiuk, Iryna A1 - Skaterna, Tetiana A1 - Horak, Iryna A1 - Sun, Yanfang A1 - Drobot, Liudmyla A1 - Matyshevska, Olga A1 - Prylutskyy, Yuriy A1 - Ritter, Uwe A1 - Frohme, Marcus T1 - Antitumor efficiency of the natural alkaloid berberine complexed with C60 fullerene in Lewis lung carcinoma in vitro and in vivo JF - Cancer Nanotechnology N2 - Background Berberine (Ber) is a herbal alkaloid with pharmacological activity in general and a high anticancer potency in particular. However, due to its low bioavailability, the difficulty in reaching a target and choosing the right dose, there is a need to improve approaches of Ber use in anticancer therapy. In this study, Ber, noncovalently bound to a carbon nanostructure C60 fullerene (C60) at various molar ratios of the components, was explored against Lewis lung carcinoma (LLC). Methods C60–Ber noncovalent nanocomplexes were synthesized in 1:2, 1:1 and 2:1 molar ratios. Ber release from the nanocomplexes was studied after prolonged incubation at different pH with the liquid chromatography–mass spectrometry analysis of free Ber content. Biological effects of the free and C60-complaxated Ber were studied in vitro towards LLC cells with phase-contrast and fluorescence microscopy, flow cytometry, MTT reduction, caspase activity and wound closure assays. The treatment with C60–Ber nanocomplex was evaluated in vivo with the LLC-tumored C57Bl mice. The mice body weight, tumor size, tumor weight and tumor weight index were assessed for four groups, treated with saline, 15 mg C60/kg, 7.5 mg Ber/kg or 2:1 C60-Ber nanocomplex (15 mg C60/kg, 7.5 mg Ber/kg). Results Ber release from C60–Ber nanocomplexes was promoted with medium acidification. LLC cells treatment with C60–Ber nanocomplexes was followed by enhanced Ber intracellular uptake as compared to free Ber. The cytotoxicity of the studied agents followed the order: free Ber < 1:2 < 1:1 < 2:1 C60–Ber nanocomplex. The potency of cytotoxic effect of 2:1 C60–Ber nanocomplex was confirmed by 21.3-fold decrease of IC50 value (0.8 ± 0.3 µM) compared to IC50 for free Ber (17 ± 2 µM). C60–Ber nanocomplexes induced caspase 3/7 activation and suppressed the migration activity of LLC cells. The therapeutic potency of 2:1 C60–Ber nanocomplex was confirmed in a mouse model of LLC. The tumor growth in the group treated with 2:1 C60–Ber nanocomplex is suppressed by approximately 50% at the end of experiment, while in the tumor-bearing group treated with free Ber no therapeutic effect was detected. Conclusions This study indicates that complexation of natural alkaloid Ber with C60 may be a novel therapeutic strategy against lung carcinoma. Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-14749 SN - 1868-6966 VL - 12 SP - 24 PB - BioMed Central ER - TY - JOUR A1 - Grebinyk, Anna A1 - Prylutska, Svitlana A1 - Grebinyk, Sergii A1 - Ponomarenko, Stanislav A1 - Virych, Pavlo A1 - Chumachenko, Vasyl A1 - Kutsevol, Nataliya A1 - Prylutskyy, Yuriy A1 - Ritter, Uwe A1 - Frohme, Marcus T1 - Drug delivery with a pH-sensitive star-like dextran-graft polyacrylamide copolymer JF - Nanoscale Advances N2 - The development of precision cancer medicine relies on novel formulation strategies for targeted drug delivery to increase the therapeutic outcome. Biocompatible polymer nanoparticles, namely dextran-graft-polyacrylamide (D-g-PAA) copolymers, represent one of the innovative non-invasive approaches for drug delivery applications in cancer therapy. In this study, the star-like D-g-PAA copolymer in anionic form (D-g-PAAan) was developed for pH-triggered targeted drug delivery of the common chemotherapeutic drugs – doxorubicin (Dox) and cisplatin (Cis). The initial D-g-PAA copolymer was synthesized by the radical graft polymerization method, and then alkaline-hydrolyzed to get this polymer in anionic form for further use for drug encapsulation. The acidification of the buffer promoted the release of loaded drugs. D-g-PAAan nanoparticles increased the toxic potential of the drugs against human and mouse lung carcinoma cells (A549 and LLC), but not against normal human lung cells (HEL299). The drug-loaded D-g-PAAan-nanoparticles promoted further oxidative stress and apoptosis induction in LLC cells. D-g-PAAan-nanoparticles improved Dox accumulation and drugs’ toxicity in a 3D LLC multi-cellular spheroid model. The data obtained indicate that the strategy of chemotherapeutic drug encapsulation within the branched D-g-PAAan nanoparticle allows not only to realize pH-triggered drug release but also to potentiate its cytotoxic, prooxidant and proapoptotic effects against lung carcinoma cells. Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-16749 VL - 4 IS - 23 SP - 5077 EP - 5088 PB - Royal Society of Chemistry (RSC) ER - TY - JOUR A1 - Grebinyk, Anna A1 - Prylutska, Svitlana A1 - Grebinyk, Sergii A1 - Prylutskyy, Yuriy A1 - Ritter, Uwe A1 - Matyshevska, Olga A1 - Dandekar, Thomas A1 - Frohme, Marcus T1 - Complexation with C60 Fullerene Increases Doxorubicin Efficiency against Leukemic Cells In Vitro JF - Nanoscale Research Letters N2 - Conventional anticancer chemotherapy is limited because of severe side effects as well as a quickly evolving multidrug resistance of the tumor cells. To address this problem, we have explored a C60 fullerene-based nanosized system as a carrier for anticancer drugs for an optimized drug delivery to leukemic cells. Here, we studied the physicochemical properties and anticancer activity of C60 fullerene noncovalent complexes with the commonly used anticancer drug doxorubicin. C60-Doxorubicin complexes in a ratio 1:1 and 2:1 were characterized with UV/Vis spectrometry, dynamic light scattering, and high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The obtained analytical data indicated that the 140-nm complexes were stable and could be used for biological applications. In leukemic cell lines (CCRF-CEM, Jurkat, THP1 and Molt-16), the nanocomplexes revealed ≤ 3.5 higher cytotoxic potential in comparison with the free drug in a range of nanomolar concentrations. Also, the intracellular drug’s level evidenced C60 fullerene considerable nanocarrier function. The results of this study indicated that C60 fullerene-based delivery nanocomplexes had a potential value for optimization of doxorubicin efficiency against leukemic cells. Y1 - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:526-opus4-11415 SN - 1556-276X VL - 14 ER -