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IMMUNOGLOBULIN G GLYCOSYLATION IN DOWN SYNDROME
No full textGlikozilacija je jedna od najčešćih ko- i posttranslacijskih modifikacija proteina. Glikani vezani na imunoglobulin G (IgG) utječu na njegovu strukturu i funkciju i mijenjaju se s dobi te u brojnim fiziološkim i patološkim stanjima organizma. Znakovi preuranjenog starenja, kao i brojne druge bolesti, česti su kod Downova sindroma (DS). Tekućinskom kromatografijom ultravisoke učinkovitosti analizirana je glikozilacija IgG-a u tri populacije osoba s DS-om za koje su postojali klinički podaci o najčešćim komorbiditetima i uspoređena s onom njihovih zdravih vršnjaka. Proučen je utjecaj produkata gena kromosoma 21 B3GALT5, RUNX1 i DYRK1A na glikozilaciju IgG-a u DS-u digestijom glikana IgG-a egzoglikozidazama te kemijskom inhibicijom produkata gena u limfoblastoidnim staničnim linijama. Pokazano je kako glikozilacija IgG-a osoba s DS-om neovisno o učestalim komorbiditetima odgovara kronološki starijim zdravim osobama. U korištenim eksperimentalnim sustavima RUNX1 ima izravan utjecaj na glikozilaciju IgG-a u DS-u, dok B3GALT5 i DYRK1A nemaju.Glycosylation is one of the most common co- and posttranslational modifications. Glycans attached to immunoglobulin G (IgG) affect its function and change with age and in numerous physiological and pathological conditions. Signs of premature aging, along with many other diseases, are common in Down syndrome (DS). IgG glycosylation of three DS populations that had clinical data on the most common comorbidities was analyzed by ultra-high performance liquid chromatography and compared to their healthy peers. The impact of chromosome 21 gene products B3GALT5, DYRK1A and RUNX1 on IgG glycosylation was studied using enzymatic digestions of IgG glycans and IgG-producing lymphoblastoid cell lines obtained from individuals with and without DS. IgG glycosylation was revealed to be significantly shifted in the direction of aging in DS, showing a ubiquitous trend independent of comorbidities, and an insight into the effect of gene products B3GALT5, DYRK1A and RUNX1 on IgG glycosylation was provided
Effect of copper oxide nanoparticles on oxidative stress in the freshwater alga Chlorella vulgaris
No full textBakar je važan metal sve zastupljeniji u obliku nanomaterijala u svakodnevnom životu. Intenzivna proizvodnja i korištenje materijala koji sadrže nanočestice bakra dovodi do otpuštanja tih čestica u okoliš što predstavlja potencijalnu opasnost za žive organizme zbog njihove visoke reaktivnosti i toksičnosti. Zelena alga Chlorella vulgaris je uvriježeni modelni organizam u ekotoksikološkim istraživanjima. U svom diplomskom radu analizirao sam utjecaj nanočestica CuO i Cu2O te iona bakra (CuSO4) u koncentracijama koje omogućuju preživljenje stanica alge C. vulgaris od 75% nakon 72 sata. Rezultati pokazuju da su nanočestice i ioni bakra značajno povećali razine reaktivnih oblika kisika, što je narušilo integritet biomembrana, dok su nanočestice CuO dodatno izazvale oštećenje proteina u stanicama. Nanočestice bakra nisu značajno utjecale na promjenu aktivnosti antioksidacijskih enzima, dok je ionski bakar značajno inhibirao njihovu aktivnost. Sadržaj reduciranog glutationa se povećao, a razina prolina smanjila nakon svih tretmana što ukazuje na pojačanu pripremu, ali i na kontinuiranu obranu stanica od oksidacijskog stresa. Nakon svih tretmana, ekspresija proteina Hsp70 se nije promijenila, dok se ekspresija proteina Hsp90 značajno smanjila. Izražena plazmoliza i destabilizacija stanične stijenke i membrane prati sve tretmane. Nanočestice CuO i Cu2O nakupljaju se na stanicama alge, u vakuoli te na tilakoidnim membranama. Tretmani s nanočesticama bakra imaju fitotoksičniji učinak na stanice alge u odnosu na tretman ionima bakra. Nanočestice Cu2O bile su fitotoksičnije na stanice alge u odnosu na nanočestice CuO.Copper is an important metal that is increasingly present in everyday life in the form of nanomaterials. The intensive production and use of materials containing copper nanoparticles leads to the release of these particles into the environment, posing a potential risk to living organisms due to their high reactivity and toxicity. The green alga Chlorella vulgaris is an established model organism in ecotoxicological research. In my master thesis, I investigated the influence of CuO and Cu2O nanoparticles and copper ions (CuSO4) in concentrations that enable 75% of the algal cells of C. vulgaris to survive after 72 hours. The results show that copper nanoparticles and ions significantly increase the level of reactive oxygen species, that damage the integrity of biomembranes, while CuO nanoparticles additionally cause protein damage in the cells. Copper nanoparticles had no significant effect changing the activity of antioxidant enzymes, while ionic copper significantly inhibited their activity. Reduced glutathione levels increased, and proline levels decreased after all treatments, indicating improved priming but also continuous defense of cells against oxidative stress. After all treatments, the expression of the Hsp70 protein did not change, while the expression of the Hsp90 protein decreased significantly. Pronounced plasmolysis and destabilization of the cell wall and membrane were observed after all treatments. CuO and Cu2O nanoparticles accumulate on algal cells, in the vacuole and on thylakoid membranes. Copper nanoparticle treatments have a more toxic effect on algal cells compared to copper ions. Cu2O nanoparticles were more phytotoxic to algal cells compared to CuO nanoparticles
Molecular characterization of ribosomal group 16SrV phytoplasma isolates from north-western Croatia
No full textFitoplazme su biljne patogene bakterije bez stanične stijenke koje pripadaju rodu 'Candidatus Phytoplasma'. Nastanjuju floem biljaka i prenose se kukcima-vektorima koji su ujedno i njihovi domaćini. Fitoplazme predstavljaju veliku opasnost u poljoprivredi uništavanjem usjeva raznih poljoprivrednih kultura, a u Europi su poznate po uništavanju usjeva vinovih loza uzrokujući bolesti žutica vinove loze (eng. grapevine yellows; GY). S tom bolesti povezane su fitoplazme flavescence dorée (FDp; ribosomska skupina 16SrV) te bois noir (ribosomska podskupina 16SrXII-A; 'Ca. P. solani'). Cilj ovog diplomskog rada je bila molekularna karakterizacija izolata FDp dobivenih iz uzoraka vinove loze, crne johe i kupine prikupljenima na sjeverozapadnom području Hrvatske. Nakon potvrde prisustva fitoplazmi u izolatima triplex real‐time PCR‐om, napravljena je filogenetska analiza gena map i vmpA umnoženih PCR-om. Utvrđena je velika genetska varijabilnost oba gena, a otkriveno je potencijalnih šest novih genotipova FDp te je po prvi put napravljena molekularna karakterizacija fitoplazmatskih izolata iz kupine i crne johe u Hrvatskoj. Također, velik broj pozitivnih izolata iz crne johe pokazuje značajan potencijal širenja zaraze ovom fitoplazmom. Dobiveni rezultati omogućuju bolji uvid u epidemiologiju FDp na području sjeverozapadne Hrvatske.Phytoplasmas are plant-pathogenic bacteria lacking cell wall that belong to the genus ‘Candidatus Phytoplasma’. They inhabit the phloem of plants and are transmitted by insect vectors that are also their natural hosts. They pose a significant threat to agriculture by destroying crops from various cultures and are known in Europe for affecting grapevine crops by causing grapevine yellows (GY) disease. This disease is associated with flavescence dorée phytoplasmas (FDp; 16SrV ribosomal group) and bois noir phytoplasmas (16SrXII-A ribosomal subgroup; ‘Ca. P solani’). This study was focused on the molecular characterization of FDp isolates obtained from samples of grapevines, black alders and blackberries collected in north-western Croatia. After confirming the presence of phytoplasmas in isolates by using triplex real‐time PCR, a phylogenetic analysis of the map and vmpA genes amplified by PCR was performed. Significant genetic variability of both genes was determined, and six potential new FDp genotiypes were discovered, along with the first molecular characterization of phytoplasma isolates from blackberries and black alders in Croatia. Moreover, numerous positive isolates from black alders indicate a significant potential for the infection spread of this phytoplasma. The obtained results provide better insight into the epidemiology of FDp in the north‐western part of Croatia
Systems for extraction and filtration of fumes and gases in welding
No full textOdržavanje čistog zraka u industriji zavarivanja ključno je za sprječavanje ozbiljnih zdravstvenih problema/rizika među zavarivačima, povećanje produktivnosti i održavanje usklađenosti sa sigurnosnim standardima. Izloženost dimu od zavarivanja, koji je poznat kao kancerogen, zahtijeva sigurno i usklađeno radno okruženje za zaštitu zavarivača.
Bez obzira radi li se o maloj radionici za zavarivanje, zavarivanju na terenu ili velikoj automatiziranoj tvornici, treba se posvetiti pružanju rješenja za odvod dima. Bez obzira na korištenu metodu zavarivanja, bila ona robotska, cobot ili ručna, učinkovita ekstrakcija dima pri zavarivanju je imperativ za zaštitu zavarivača, održavanje produktivnosti i usklađenost s pravilima i propisima. Čimbenici koji uključuju metodu sakupljanja, dizajn industrijske ventilacije, odabir medija za filtriranje i pravilne prakse servisiranja/održavanja doprinose tome koliko ekstrakcija dima može biti energetski učinkovita.Ensuring clean air in the welding industry is critical to preventing serious health
problems/risks among welders, increasing productivity and maintaining compliance with safety
standards. Exposure to welding fumes, which is a known carcinogen, requires a safe and
compliant work environment to protect welders.
Whether it is a small welding shop, field welding or a large, automated factory, it should be
committed to providing fume extraction solutions. Regardless of the welding method used, be
it robotic, cobot or manual, effective welding fume extraction is imperative to protect welders,
maintain productivity and comply with rules and regulations. Factors including collection
method, industrial ventilation design, filter media selection, and proper service/maintenance
practices contribute to how energy efficient fume extraction can be
Microclimatic features of the cave Golubnjača in Plitvice Lakes National Park
No full textU diplomskom radu su prikupljeni i analizirani podaci o mikroklimi špilje Golubnjače u NP Plitvička jezera. Do 1991. godine Golubnjača je bila turistička špilja. Podaci su prikupljeni memorijskim termohidrografima, anemometrom te logerom za koncentraciju ugljikovog dioksida. Mikroklimatski parametri su mjereni 2022. i 2023. godine. Analizirani su i podaci o antropogenom utjecaju u špilji tijekom radionice restauriranja. Svrha mjerenja bila je odrediti na koji način se mikroklima u podzemlju mijenja i zašto te što i kako utječe na promjene koje se događaju.The thesis collects and analyses data on the microclimate of the Golubinjača cave, which is located in the Plitvice Lakes National Park. Until 1991, the cave was used for tourism. Data were collected with memory thermohydrographs, anemometer, and carbon dioxide concentration logger. Microclimatic parameters were measured in 2022 and 2023. Moreover, the data on the anthropogenic impact on the cave during the restoration workshop was also analysed. The purpose is to determine how the microclimate of Golubnjača is changing and why as well as what affects the changes that occur and how
Control of lubrication system in a steel wire manufacturing plant
No full textU ovome radu opisan je postupak projektiranja sustava za podmazivanje stroja u čeličani. Ukratko je opisano cijelo postrojenje s glavnim dijelovima. Dan je opis procesa podmazivanja, odnosno svi parametri koji moraju biti zadovoljeni. Korištenjem Siemens TIA portala napisan je PLC upravljački program procesa podmazivanja TMB-a. Proces podmazivanje sa svim fizikalnim veličinama (temperatura, tlak ulja) vidljiv je na korisničkom sučelju koje je napravljeno u programu WinCC Explorer V7.0. Cijeli proces je simuliran pomoću Siemens PLCSIM Advanced programa. Na kraju rada opisana je implementacija podmazivanja na TMB stroju, gdje su prikazana sva dopuštenja koja je potrebno zadovoljiti prilikom rada sustava i dan je prikaz alarma koji se javljaju prilikom kvara nekih dijelova sustava.This work describes the process of designing a lubrication system for one of the machines in the steel mill. The entire plant with its main components is briefly described. The lubrication process is described, including all parameters that must be met. Using the Siemens TIA Portal, a PLC control program for the lubrication process of the TMB machine was written. The lubrication process with all physical quantities (temperature, oil pressure) is visible on the user interface created in WinCC Explorer V7.0. The entire process is simulated using the Siemens PLCSIM Advanced program. At the end of the work, the implementation of lubrication on the TMB machine is shown, including all permissions that need to be satisfied during system operation, and alarm displays that occur in case of failure of some system parts are provided
Trends in newspaper criticism of Croatian feature fiction films from 2001 to 2019
No full textU središtu je ove disertacije znanstveno istraživanje suvremene novinske filmske kritike hrvatskih dugometražnih igranih filmova. Primjenom različitih metoda analiza medijskog teksta (prvenstveno kvantitativno-kvalitativnom analizom sadržaja) i dubinskim intervjuima s kritičarima dnevnih novina analizirani su različiti aspekti novinskih kritika i hrvatskih filmova. Cilj je bio ovim istraživanjem ukazati i na osobitosti suvremene hrvatske novinske filmske kritike. Istraženi su, nadalje, trendovi i promjene u načinu praćenja, analiziranja i kritičkog vrednovanja hrvatskog dugometražnog igranog filma u dnevnom tisku u kontekstu hrvatskog sociokulturnog okruženja u 21. stoljeću. Pokazalo se kako se u razdoblju od 2001. do 2019. godine u dnevnim novinama kritička recepcija hrvatskog filma promijenila iz neutralne u pretežno pozitivnu, te se utvrdilo povezivanje zaključaka kritika i širih društvenih pitanja, veća razina afirmativnosti kritike prema filmovima za koje se smatra da ukazuju na aktualne društvene probleme, te mala razina senzacionalizma uredničkih naslova kritika. Također su predstavljeni, sintetizirani, analizirani i komentirani rezultati pripremnog istraživanja, a kojim se utvrdilo, uz analizu kritičarske argumentacije i društvenog konteksta u navedenom razdoblju, koji su kritički najpozitivnije, a koji najnegativnije ocijenjeni hrvatski igrani filmovi, redatelji i žanrovi, na koje načine hrvatski igrani film pridonosi pluralizmu predstavljanja društvenih tema. Rezultati ovog istraživanja pružaju znanstvene uvide u osobitosti i trendove suvremene hrvatske novinske filmske kritike, a što predstavlja novost u nacionalnoj znanstvenoj literaturi i znanstveni doprinos istraživanju hrvatskog novinarstva. Utvrđivanjem osobitosti suvremenog hrvatskog dugometražnog igranog filma kao masovnog medija današnjice iz perspektive dnevne novinske filmske kritike, u kontekstu informacijskih i komunikacijskih znanosti (iz grana novinarstva i masovnih medija) i znanosti o umjetnosti (iz grane filmologije), ova je disertacija dala i relevantan multidisciplinarni doprinos proučavanju suvremenog hrvatskog filma.The focus of this doctoral dissertation is the scientific study of contemporary Croatian newspaper film criticism of Croatian feature films. Different aspects of newspaper criticism and Croatian cinema were analysed using different methods of media text analysis (primarily quantitative and qualitative content analysis) and in-depth interviews with daily newspaper critics. The main aim of the research was to show the specificities of contemporary Croatian newspaper film criticism. Furthermore, trends and changes in the way of covering, analysing, and critically evaluating Croatian feature films in the daily press in the context of the Croatian socio-cultural environment in the 21st century were explored. The research shows how in the period from 2001 to 2019, the critical reception of Croatian films in the daily newspapers changed from neutral to predominantly positive, the conclusions of critics were connected to broader social issues, there was a higher level of affirmative criticism towards films that were considered to indicate on current social problems, and there was a low level of sensationalism in editorial headlines. It also presents, synthesizes, analyses, and comments on the results of the preliminary research that determined, always along with the analysis of critical arguments and the social context in the
mentioned period, which are the most positive and most negatively evaluated Croatian feature films, directors and genres, and in which ways Croatian cinema contributes to the pluralism of the representation of social topics.
The results of this research provide scientific insights into the specificities and trends of contemporary Croatian newspaper film criticism, which is a novelty in the national scientific literature and a scientific contribution to the research of Croatian journalism. By establishing the specificities of the contemporary Croatian feature film as contemporary mass media from the perspective of daily newspaper film criticism, in the context of Information and Communication
sciences (Journalism and Mass Media) and Art Science (Filmology), this dissertation also made a relevant multidisciplinary contribution to the study of contemporary Croatian cinema. The dissertation is divided into nine chapters. In the introductory chapter, the subject of the dissertation, the research objectives and hypotheses, the structure of the dissertation, a review of the literature important for the theoretical treatment of the topic and the scientific contribution of
the work are presented. The second chapter describes the basic concepts important for understanding the content of the dissertation and their main features: journalistic genres, criticism and newspaper criticism, film criticism and newspaper film criticism. The third chapter deals with the origin and development of film criticism in the world and in Croatia, and then with the Croatian newspaper film criticism and Croatian feature film in the 21st century: critical reception of Croatian film directors, genres of Croatian feature film and analysis of social themes and subjects in Croatian film. The fourth chapter describes the research methodology: all hypotheses were tested by quantitative-qualitative content analysis and in-depth interviews, and the dissertation also uses inferential statistics in order to formally test hypothesis H3. At the end of the chapter, the results of the research are presented, i.e. the analysis and comparison of the results of the conducted
research, which prove each of the 4 hypotheses. The fifth chapter is the discussion, and the sixth is the conclusion of the entire dissertation, especially the research results. An overview of scientific literature, is listed in the seventh chapter, appendices (code sheet, in-depth interview protocol, and list of newspaper reviews from printed editions and online editions) in the eighth, and the author's biography and published works in the ninth. The dissertation determined how the critical reception of the Croatian feature films changed from neutral to predominantly positive during the period. Critical reception can be described by a neutral (average) average rating of 2.88 in the first year of research, 2.89 in the first three and four years, and 2.90 in the first five years of research, while at the end of the period covered by the research,
critical reception is predominantly positive, as assumed in the hypothesis, with a score of 3.66 in the last year of research, 3.51 in the last three years and 3.5 in the last 4 years. The difference in the earlier and later periods of the analysed period is also visible in the result pointing that, according to critics' evaluations, extremely bad films are produced less often in later periods, and that in later periods there are more films rated moderately and extremely positively. Critics point out that they link their negative critical reception of Croatian films (and that of their colleagues
from other daily newspapers) to the 1990s, and not to films from the beginning of the 21st century, for which they give a relatively low, but still average, rating. It was established that newspaper reviews of Croatian feature films are characterized by drawing conclusions in connection with wider social issues, thus confirming the theoretical assumptions of newspaper film criticism that Žigo and other theorists of criticism highlighted in their works. 29% of reviews were characterized by a strong level of conclusions regarding social issues, 51% fair, 20% moderate, and 0.68% low. The average recorded level of drawing conclusions in connection with broader social issues in all three analysed lists is 4.08. During the years covered by the research, there were no significant deviations: the lowest level of 3.81 was recorded in 2003 and 2018, and the highest 4.32 was recorded in 2009, so in all the researched years the average level was considerable. Critics point out that the reason for drawing conclusions in connection with broader social issues, apart from their critical interests, are the topics that Croatian films deal with, because Croatian cinematography less often films escapist genre productions, and more often dramas from Croatian everyday life. It was also established that Croatian newspaper film critics predominantly wrote more affirmatively about Croatian feature films that they considered to indicate current social problems, than about those that did not. Films rated with extremely and moderately positive reviews had a
fair level of pointing out social problems (4.34 and 4.06), as well as neutral (average) rated ones (3.54), while moderately and extremely negatively rated films had a moderate level of pointing out to social problems (3.04 and 2.78). The research showed that critics mostly emphasize that they value socially relevant films, but only if they are of high quality, which was also confirmed by the research of their reviews. There are several examples according to which it can be noted
that social criticism by itself cannot bring positive criticism to a film that does not have the appropriate qualities. Thus, socially critical films received generally more positive reviews than socially non-critical ones due to the greater overall quality that most of them were distinguished by, and not only because of film critics' affinity towards social criticism. The editorial titles of criticism of Croatian feature films in Croatian daily newspapers were to a small extent sensationalistic, and mostly objectively reflected the views of the authors. The titles studied were, depending on the critics and editors, interesting, informative, funny, intelligent, ambiguous, likeable, intriguing or provocative, and the level of sensationalism ranged from the level of not at all (36%), very little (51%) to moderate (13 %), with an average low level of 1.78, without quite or extremely sensational headlines. Titles that are moderately sensational are often characterized by exaggeration, they often refer to popular culture (songs, sayings, books and films), but mostly objectively reflect the critic's attitude. Editors generally readily accepted the most intriguing theses, quips or comparisons offered by critics in their texts. Critics point out that newspaper headlines can be sensationalistic and reflect the author's position, just as they can falsify the author's position without being sensationalist at all, and the analysis of their criticisms shows that there were all the cases mentioned, so the sensationalism of the headlines should not be equated with the non-objective presentation of the critic's position. The most positively evaluated directors by critics among those with three or more films from 2001. till 2019. were Rajko Grlić, Ognjen Sviličić, Zrinko Ogresta, Vinko Brešan, Antonio Nuić, Arsen Anton Ostojić, Dalibor Matanić and Kristijan Milić, with a note that according to critics' ratings, filsm made by Grlić, Brešan and Ogresta had the most consistent quality. The most negatively evaluated directors among those with two or more films in this period are Vicko Ruić, Zdravko Mustać, Filip Šovagović, Jakov Sedlar, Fadil Hadžić and Krsto Papić. No female director made 3 feature films in this period, and only Snježana Tribuson made 2. Nevertheless, women directed some of the highest-rated films in this period. At the top of the list is Hana Jušić, Dana Budisavljević and Ivona Juka are among the highly respected, followed by Snježana Tribuson. The most positively rated Croatian films by newspaper film critics are Hana Jušić's Quit Staring at my Plate and Dalibor Matanić's High Noon, while the third on the list is A stranger by Bobo Jelčić. The most negatively rated Croatian films by newspaper film critics are The Recollection Thief and Seraphim, the Lighthouse Keeper's Son by Vicko Ruić, followed by Remembrance of Georgia by Jakov Sedlar and The Enchanting Porkers by Ivan Livaković. Croatian films in the 21st century that received the most positive reviews most often belong to the genre of social-psychological drama contemporary drama, the second genre is action, while the third is war drama. Among the genres whose representatives received the most negative reviews in the 21st century, road films, erotic films and biographical historical dramas are leading the way. Thematically speaking, global themes were definitely never the focus of Croatian directors, who mostly dealt with domestic problems and motives. Croatian cinematography dealt with the subject of violence and torture in many ways, intensively and, according to critics, competently, from historical (from the Second World War to the Croatian Homeland War) to contemporary subjects (terror against sexual minorities in Croatian society, fan hooligan violence), and the treatment of these subjects created some of the most appreciated film productions of that period. Regarding national minorities, the Serbian minority in Croatia was the most common film inspiration. Croatian films undoubtedly point to the activities of economic and political power holders, but cautiously and somewhat abstractly. Even the bravest directors did not pass their critical judgment on some specific political names from Croatian politics through their films, and with the exception of Vinko Brešan, almost no one dealt more seriously with the characters of Croatian prime ministers and presidents, but even in his films we can see general characters of politicians who cannot be related for some concrete Croatian political figures. There are very few films that have caused wider social controversies. Analysing the diversity of the angle of view of individual directors, i.e. their approach in the presentation of key social themes in Croatian feature films, we come to the conclusion that the most socially critical director was Vinko Brešan (with the themes of war crimes of Croatian soldiers, sexual abuse of children by priests and political manipulation), and then Branko Schmidt (with topics about young thugs and drug addicts, crimes within the healthcare system and
pedophilia in the Church). This dissertation provides a relevant scientific contribution to the study of contemporary
newspaper film criticism, as well as contemporary Croatian journalism in general. The results obtained in this doctoral dissertation provide scientific insight into the specificities of contemporary Croatian newspaper film criticism, but they also contribute to the positioning of newspaper film criticism in the context of journalistic discourse, pointing to the specifics of
newspaper film criticism as a journalistic genre which, in this sense, should be understood beyond the narrow links with literature and filmology, in the context of daily newspapers in which reviews are published and by connecting the conclusions with socially relevant topics. It should be emphasized that the proposed research model, taking into account the specifics of theatrical and literary works, could be applied to newspaper literary and theatrical criticism
Promiscuous aldol additions of phosphoenolpyruvate to aliphatic and aromatic aldehydes catalysed by PEP-dependent synthases
No full textStvaranje C-C veze ključna je u sintetskoj organskoj kemiji jer povezuje manje podjedinice u složenije molekule. Klasična sinteza zahtijeva sofisticirane asimetrične metode i zaštitu funkcionalnih skupina. Biokatalitička kondenzacija postaje sve zanimljivija jer omogućava dobivanje sterički čistih molekula bez potrebe za zaštitom funkcionalnih skupina. Ovaj rad istražuje neprirodne aldolne adicije fosfoenolpiruvata (PEP) na alifatske i aromatske aldehide, katalizirane PEP-ovisnim aldolazama, fokusirajući se na enzime DHAPH i KDOH iz metagenomske zbirke. Cilj rada bio je optimizirati reakcijske uvjete i ispitati konverziju i enantioselektivnost ovih enzima kako bi se utvrdila njihova primjenjivost u formiranju C-C veze. U eksperimentalnom dijelu, istraženo je 38 enzima iz metagenomske zbirke s različitim aldehidima. Najbolji rezultati postignuti su s enzimom KDOH i aldehidom fenilacetaldehidom, gdje su konverzije dosegle do 92,7 %. Posebno su se istaknuli karbamatni spojevi, s konverzijama koje su prelazile 56,6 %. Optimizacija udjela dimetilformamida (DMF) pokazala je da smanjenje DMF-a povećava enzimske aktivnosti, ali je njegovo potpuno uklanjanje nepoželjno zbog netopivosti aldehida u vodi. DMF udio od 5 % naspram 20 % doveo je do značajnog povećanja konverzije, čak do 44,9 %. Daljnja ispitivanja pročišćenih enzima pokazala su niže stope konverzije u usporedbi s početnim probirom, a neki od razloga mogu biti gubitk stabilizirajućih faktora ili parcijalne denaturacije enzima tijekom pročišćavanja. Također, enantioselektivnost se nije mogla jasno odrediti zbog niske aktivnosti enzima i formiranja nusprodukata umjesto ciljanih produkata reakcije. DHAPH i KDOH enzimi imaju potencijal za daljnju primjenu u formiranju C-C veze, ali su potrebna dodatna istraživanja i optimizacija reakcijskih uvjeta kako bi se postigli zadovoljavajući rezultati.The formation of C-C bonds is crucial in synthetic organic chemistry as it connects smaller subunits into more complex molecules. Sophisticated asymmetric methods and protection of functional groups are required in classical synthesis. Biocatalytic condensation is becoming increasingly interesting as it allows the production of sterically pure molecules without the need for functional group protection. Unnatural aldol additions of phosphoenolpyruvate (PEP) to aliphatic and aromatic aldehydes, catalyzed by PEP-dependent aldolases, are investigated in this study, focusing on DHAPH and KDOH enzymes from a metagenomic library. The aim was to optimize reaction conditions and examine the conversion and enantioselectivity of these enzymes to determine their applicability in C-C bond formation.
In the experimental part, 38 enzymes from the metagenomic library were tested with different aldehydes. The best results were achieved with the KDOH enzyme and phenylacetaldehyde, with conversions reaching up to 92,7 %. Carbamate compounds were particularly notable, with conversions exceeding 56,6 %. Optimization of dimethylformamide (DMF) content showed that reducing DMF increases enzyme activities, but its complete removal is undesirable due to the insolubility of aldehydes in water. A DMF content of 5 % compared to 20% led to a significant increase in conversion, up to 44,9 %. Further testing of purified enzymes showed lower conversion rates compared to initial screening, possibly due to the loss of stabilizing factors or partial denaturation of enzymes during purification. Additionally, enantioselectivity could not be clearly determined due to low enzyme activity and the formation of by-products instead of target reaction products. DHAPH and KDOH enzymes have potential for further application in C-C bond formation, but additional research and optimization of reaction conditions are needed to achieve satisfactory results
Validation of the MEKC-MS/MS method for quantitative analysis of anastrozole and letrozole in human plasma
No full textInhibitori aromataze anastrozol i letrozol koriste se u liječenju o estrogenu ovisnog i HER2
negativnog raka dojke kod postmenopauzalnih pacijentica, kao monoterapija ili u kombinaciji s
drugim lijekovima poput CDK4/6 inhibitora. Praćenje pridržavanja pacijentica ovako dugotrajnoj
terapiji ključno je za postizanje željenih ishoda liječenja.
Glavni cilj ovog diplomskog rada bio je validirati brzu i jednostavnu novorazvijenu MEKC-MS/MS
metodu za analizu inhibitora aromataze anastrozola i letrozola u biološkim uzorcima, što je važno
za terapijsko praćenje lijekova. Ispitivani validacijski parametri bili su linearnost i kalibracijska
krivulja, točnost i preciznost, stabilnost uzoraka u autosampleru te utjecaj matrice.
Validacija metode pokazala je da matrica ne predstavlja analitički problem prilikom ispitivanja
anastrozola (ME<1,3%) i letrozola (ME<17,3%). Parametar linearnosti ove analitičke metode
zadovoljen je za anastrozol (r>0,9945) u zadanom koncentracijskom rasponu od 20 do 200 ng/mL
kao i za letrozol (r>0,9802) u zadanom koncentracijskom rasponu od 30 do 300 ng/mL. Točnost i
preciznost (odstupanje, RSD<15%, odnosno <20% za LLOQ) zadovoljavajuće su za oba analita.
Također, stabilnost anastrozola i letrozola u autosampleru potvrđena je (odstupanje, RSD<15%), što
doprinosi pouzdanosti same metode.Aromatase inhibitors anastrozole and letrozole are used in the treatment of estrogen-dependent and
HER2-negative breast cancer in postmenopausal patients, either as monotherapy or in combination
with other drugs such as CDK4/6 inhibitors. Monitoring patient adherence to such long-term therapy
is crucial for achieving desired treatment outcomes.
The main objective of this thesis was to validate a fast and simple newly developed MEKC-MS/MS
method for the analysis of aromatase inhibitors anastrozole and letrozole in biological samples,
which is important for therapeutic drug monitoring. The validation parameters tested included
linearity and calibration curve, accuracy and precision, sample stability in the autosampler, and
matrix effect.
The method validation demonstrated that the matrix does not pose an analytical problem for the
analysis of anastrozole (ME<1.3%) and letrozole (ME<17.3%). The linearity parameter of this
analytical method was met for anastrozole (r>0.9945) in the specified concentration range of 20 to
200 ng/mL, as well as for letrozole (r>0.9802) in the specified concentration range of 30 to 300
ng/mL. Accuracy and precision (bias, RSD<15%, and <20% for LLOQ) were satisfactory for both
analytes. Additionally, the stability of anastrozole and letrozole in the autosampler was confirmed
(bias, RSD<15%), contributing to the reliability of the method itself
Quality control of Verbasci flos
No full textU okviru ovog diplomskog rada uspoređena su dva uzorka droge Verbasci flos sakupljena na dvije različite lokacije te je provedena kontrola kakvoće tih uzoraka. Određivanjem broj bubrenja i gubitka sušenjem ustanovljeno je da uzorci odgovaraju farmakopejskim zahtjevima, dok je u uzorku 2 pronađena nešto veća količina fragmenta čaške i ostalih stranih tvari. Mikroskopskim pregledom pronađeni su svi dijelovi koje Farmakopeja propisuje. Tankoslojnom kromatografijom (TLC) dokazana je prisutnost flavonoida i fenolnih kiselina koje su potom određene UV-Vis spektrofotometrijskom metodom te je sadržaj iznosio 0,053-0,075% i 0,08- 0,24%. Također određen je i sadržaj trjeslovina koji je iznosio 0,172-0,24%.As part of this thesis, two types of mullein flower from two different locations were compared and the quality control of these samples was carried out. By determining the swelling index and loss on drying, it was found that the samples correspond to pharmacopoeial requirements, while in the sample 2 there was slightly larger amount of fragments of the calyx, and other foreign substances was found. By microscopic examination we found all the parts prescribed by the Pharmacopoeia. Thin-layer chromatography (TLC) showed the presence of flavonoids and phenolic acids, which were then quantified by UV-vis spectrometric method, with the content ranging from 0.053% to 0.075% and 0.08% to 0.24%. The content of tannins was also determined, ranging from 0.172% to 0.24%