Development and validation of a new bioanalytical method for simultaneous determination of ribociclib, letrozole and ibuprofen by capillary electrophoresis

Rak dojke, s dominantnim podtipom HR+ HER2−, najčešći je rak u svijetu. Jedan od načina liječenja HR+ HER2− raka dojke je kombinacijom CDK4/6 inhibitora i inhibitora aromataze, često uz lijekove ribociklib i letrozol. Kapilarna elektroforeza metoda je koja se iz dana u dan sve više razvija te bi se s odgovarajućom reproducibilnošću i osjetljivošću mogla koristiti u svrhu terapijskog praćenja lijekova. U ovom je radu razvijena i validirana metoda za određivanje ribocikliba, letrozola i ibuprofena kapilarnom elektroforezom uz ketoprofen kao unutarnji standard. Separacija je postignuta uz 75 mmol dm−3 fosfat, 8,5 mmol dm−3 sulfobutileter-β-ciklodekstrin (SBE-β-CD) i 5 % izopropanola u radnom puferu pri naponu 15 kV i temperaturi 25 °C. Uzorak je injektiran hidrodinamički pri tlaku −50 mbar u trajanju 5 sekundi, nakon injektiranja ultračiste vode 6 sekundi. Metoda je precizna, linearna u rasponu masene koncentracije od 1 do 75 µg cm−3, specifična, robusna za promjenu temperature, koncentracije SBE-β-CD te napona.Breast cancer is the most common type of cancer in the world, with subtype HR+ HER2− being most prevalent. HR+ HER2− breast cancer can be treated with hormone therapy, with ribociclib (CDK 4/6 inhibitor) and letrozole (aromatase inhibitor) being a commonly used medicine combination. Capillary electrophoresis is a rapidly developing method which could be used for therapeutic drug monitoring purposes after providing acceptable reproducibility and sensitivity. A method for analysis of ribociclib, letrozole and ibuprofen by capillary electrophoresis using ketoprofene as an internal standard was developed and validated. The separation was obtained with 75 mmol dm−3 phosphate, 8,5 mmol dm−3 sulphobutylether-β-cyclodextrin (SBE-β-CD) and 5% isopropanol. The hydrodynamic injection was performed at −50 mbar pressure for 5 seconds, after the injection of ultrapure water for 6 seconds. The method is precise, linear in a range of mass concentration of 1 to 75 µg cm−3, specific and robust for the change of temperature, SBE-β-CD concentration and voltage

The role of BPM1 protein in de novo DNA methylation mechanism during development of Arabidopsis thaliana L.

Proteini BPM iz vrste Arabidopsis thaliana primarnu ulogu ostvaruju u putu proteasomalne razgradnje ovisne o ubikvitinu, u sklopu kojeg prepoznaju specifične supstrate za razgradnju. Prethodno je utvrđena interakcija proteina BPM1 s proteinima RDM1 i DMS3, komponentama metilacijskog mehanizma RdDM. Uloga proteina BPM1 u mehanizmu RdDM u ovom je radu ispitana primjenom kromatinske imunoprecipitacije u svrhu identifikacije novih ciljnih gena ovog mehanizma u čijoj regulaciji metilacije posreduje protein BPM1. Identificirani su geni FBW2, RKP i AGL14, a kao referentni odabran je gen CML41. Analizom metilacije ovih gena metodom bisulfitnog sekvenciranja i pirosekvenciranja u linijama s prekomjernom ekspresijom proteina od interesa (oeBPM1, oeDMS3), te linijama s narušenom funkcijom mehanizma RdDM (dms3-1, nrpe1-10), potvrđen je stimulativni učinak proteina BPM1 na aktivnost mehanizma, koji je najizraženiji tijekom embriogeneze, a sam mehanizam RdDM značajniji je za zigotnu nego za somatsku embrigenezu. Ovisno o poziciji, učinci metilacijskih događaja reflektiraju se na ekspresiju svih analiziranih gena.Proteins BPM of Arabidopsis thaliana have a primary role in the ubiquitin-dependent proteasomal degradation pathway, where they recognize specific substrates for degradation. The interaction of BPM1 protein with RDM1 and DMS3, components of RdDM methylation mechanism, has previously been established. Here, the role of BPM1 protein in RdDM was analyzed using chromatin immunoprecipitation for identification of new RdDM targets whose methylation is mediated by BPM1 protein. Genes FBW2, RKP and AGL14 were identified as targets, and CML41 was chosen as a reference gene. Methylation analysis of these genes was conducted using bisulfite sequencing and pyrosequencing in lines with overexpression of BPM1 (oeBPM1) or DMS3 (oeDMS3), and lines with impaired RdDM function (dms3-1, nrpe1-10). The results confirmed a stimulative role of BPM1 protein in RdDM. This mechanism proved to be more significant in zygotic than somatic embyrogenesis. Depending on the position, methylation had a diverse effect on expression of analyzed genes

Virusi današnjice – prevencija, dijagnostika i liječenje

Ovaj dokument nastao je kroz projekt “Molekularna raznolikost virusa Epstein-Barr” (IP-2020-02-8961), financiran od Hrvatske zaklade za znanost, kao materijal za neformalno obrazovanje koje je dio cjeloživotnog obrazovanja

Stochastic calculus in financial mathematics

U ovom radu objasnili smo veliku važnost stohastičkog računa u financijskoj matematici, te pokazali neke od njegovih najbitnijih upotreba i utjecaj na razvoj tržišta. U prvom smo poglavlju dali kratak uvod u svijet opcija i portfelja, te se upoznali s ključnim pojmovima koji su nam potrebni kroz cijeli rad, kao što su Brownovo gibanje te Itôv integral i njegova formula. Oni nam daju osnovu za uvođenje jednog od najvažnijih modela za vrednovanje financijske imovine, a to je Black-Scholes-Mertonov model kojeg promatramo u drugom poglavlju. Prije samog izvoda modela definiramo pojmove ekvivalentne martingalne mjere i navodimo Girsanovljev teorem koji omogućuje promjenu mjere, kako za jednodimenzionalni tako i za višedimenzionalni slučaj. U trećem poglavlju analiziramo povezanost stohastičkih i parcijalnih diferencijalnih jednadžbi pomoću Feynman-Kac teorema i uvodimo Hull-White model. Na kraju, proučavamo opcije koje se sastoje od više osnovnih imovina te se na početku poglavlja upoznajemo s višedimenzionalnom Black-Scholesovom funkcijom pomoću koje u ostatku poglavlja računamo cijene tih opcija. Opisujemo tri opcije s više osnovnih imovina, a to su rainbow, basket i quanto opcije, a one igraju ključnu ulogu u smislu upravljanja rizicima, te diverzifikaciji i optimizaciji portfelja.This thesis explains the great importance of stochastic calculus in financial mathematics, and shows some of its most important applications and its influence on market development. In the first chapter, we gave a brief introduction to the world of options and portfolios, and got acquainted with the key concepts necessary throughout the paper, such as Brownian motion and It\^o integral along with its formula. They give us the foundation for introducing one of the most important models for the valuation of financial assets, the Black-Scholes-Merton model, which we analyze in the second chapter. Before deriving the model itself, we define the terms of equivalent martingale measure and state Girsanov's theorem, which allows a measure change, both for the one-dimensional and for the multidimensional case. In the third chapter, we analyze the connection between stochastic and partial differential equations using the Feynman-Kac theorem and introduce the Hull-White model. Finally, we study options that consist of several underlying assets, and at the beginning of the chapter we are introduced to the multidimensional Black-Scholes function, which we use to calculate the prices of these options in the rest of the chapter. We describe three types of multi-asset options: rainbow, basket and quanto options, which play a key role in terms of risk management, porfolio diversification and optimization

Hidromorphological status and perspectives of revitalisation of the river Jadro

Rječica Jadro, „hrvatski Jordan“, vodoopskrbna je žila kucavica Solina i širega splitskoga područja od antike do suvremenosti, ali i jamac opstanka stanovništva od samih početaka naseljenosti u njezinu poriječju. Ona je i ekosustav jedinstvenih hidroekoloških obilježja, sve više pritisnut urbanizacijom, turistifikacijom i onečišćenjima. Upravo i s ciljem poboljšanja njezina narušena hidroekološkoga stanja te postizanja održive ravnoteže rijeke i čovjeka u budućnosti provedeno je istraživanje kojemu je glavni cilj bilo predlaganje revitalizacijskih mjera u koritu. Uz prethodnu pripremu temeljenu na dostupnim kartografskim izvorima i literaturi dosadašnjih istraživanja, tijekom ljeta 2024. terenskim je izvidima utvrđeno i dokumentirano zatečeno hidromorfološko stanje rijeke u cijeloj dužini njezina toka. Nakon obrade i usporedbe terenskih i literaturnih podataka, metodom hidromorfološke ocjene provedeno je hidromorfološko vrednovanje Jadra po prethodno određenim dionicama. Na temelju te procjene odabrana su tri lokaliteta u vodotoku na kojima su predložene revitalizacijske mjere u smjeru oprirodnjenja toka uklanjanjem navedenih vodotehničkih zahvata i uređenjem naplavna područja.The Jadro River, known as the "Croatian Jordan", is not only the water supply artery of Solin and the wider urban area of Split since antiquity, but also an ecosystem of unique hydroecological characteristics, increasingly pressured by urbanization, tourism and pollution. It is precisely with the aim of improving its deteriorated hydroecological status and achieving a sustainable balance between the river and humans in the future that a study was conducted, the main goal of which was to propose revitalization measures in the riverbed. With prior preparation based on available cartographic sources and literature from previous research, during the summer of 2024, field surveys determined and documented the current hydromorphological state of the river along its entire length. After processing and comparing field and literature data, a hydromorphological evaluation of the Jadro was carried out using the hydromorphological assessment method along previously determined sections. Based on this assessment, threelocations in the watercourse were selected whererevitalization measures were proposed in the direction of naturalizing the flow, i.e. removing unnecessary water engineering interventions and regulating floodplains

Impact of the Jadral alumina factory (Obrovac) on the geochemical characteristics of the surrounding soils

Tijekom procesa proizvodnje glinice jedan od najopasnijih nusprodukata jest crveni mulj. Velike količine crvenog mulja odlažu se u sklopu tvornica na otvorenom u velikim, plitkim bazenima što u kontaktu s okolinom, čak i desetljećima nakon njihovog zatvaranja, može imati vrlo negativne posljedice zbog visoke alkalnosti te prisutnosti raznih (teških) metala u njegovom sastavu. Nekadašnja tvornica glinice Jadral nalazi se na obrovačkom području u čijoj geološkoj građi prevladavaju mezozojske i paleogneske sedimentne stijene dok se u neposrednoj blizini nalazi estuarij rijeke Zrmanje, a cijelo područje izloženo je jakim vjetrovima. Primjenom kiselog reagensa (pH=2,87) na uzorcima tla uzorkovanih na području tvornice simulirana je pojava kiselijeg ispiranja u okolišu te su određene ukupne koncentracije metala u tlima i pridruženim eluatima u četiri veličinske frakcije (<0,45μm, <0,2μm, <0,1μm, <0,008μm). Rezultati pokazuju kako sama tla imaju puferirajući učinak, a mobilnost metala varira u ovisnosti o kationskom/anionskom karakteru te o njihovoj prisutnosti u pojedinim mineralima. Smjer prijenosa metala definiran je dominantnim vjetrom koji prevladava - bura. U kiselim uvjetima najmobilniji su elementi s kationskim karakterom vezani za karbonate (Sr, Mn, Cd), dok elementi s anionskim karakterom u kiselom mediju nisu značajno mobilni.During the alumina production process, one of the most dangerous by-products is red sludge. Large amounts of red sludge are disposed of in factories outdoors in large, shallow pools, which in contact with the environment, even decades after their closure, can have very negative consequences due to the high alkalinity and the presence of various (heavy) metals in its composition. The former Jadral alumina factory is located in the Obrovac area, whose geological structure is dominated by Mesozoic and Paleogene sedimentary rocks. The estuary of the Zrmanja river is located in the immediate vicinity and the entire area is exposed to strong winds. By applying an acidic reagent (pH=2.87) to soil samples sampled in the factory area, the occurrence of acidic leaching in the environment was simulated, and the total concentrations of metals in soils and associated eluates were determined in four size fractions (<0.45μm, <0.2μm, <0.1μm, <0.008μm). The results show that the soils have a buffering effect and the mobility of metals varies depending on the cationic/anionic character and their presence in certain minerals. The direction of metal transfer is defined by the prevailing wind - bora. In acidic conditions, the most mobile elements are ones with a cationic character and bound to carbonates (Sr, Mn, Cd), while the elements with an anionic character are not significantly mobile in an acidic medium

O-benzyloximes as butyrylcholinesterase inhibitors

U okviru ovog rada klasičnim postupcima organske sinteze pripremljen je O-benzilni eter kinuklidin-3- on-oksima reakcijom hidroklorida kinuklidin-3-ona i O-benzilhidroksilamina bez dodatka organskih ili anorganskih baza. Kvaternizacijom kinuklidinskog atoma dušika u molekuli sintetiziranog Obenziloksima s metil-jodidom te odabranim, različito supstituiranim benzil-bromidima (metilna ili nitroskupina te atomi halogena u meta ili para položaju) pripremljena je serija od dvanaest novih kvaternih derivata. Odabrani spojevi imaju različite modifikacije u strukturi kako bi se mogli definirati strukturni parametri važni za postizanje poboljšane biološke aktivnosti spojeva u interakciji s kolinesterazama. Pripremljenim spojevima određeno je talište te je struktura potvrđena infracrvenom spektroskopijom, spektrometrijom masa te 1D i 2D spektroskopijom nuklearne magnetske rezonancije. Nadalje, svim pripremljenim spojevima određen je inhibitorni potencijal prema enzimu butirilkolinesterazi iz konjskog seruma (EC 3.1.1.8) korištenjem Ellmanove metode za kolorimetrijsko određivanje aktivnosti kolinesteraze.In this work, O-benzyl ether of quinuclidin-3-one oxime was prepared by the reaction of hydrochloride salts of quinuclidin-3-one and O-benzylhydroxylamine without the addition of organic or inorganic bases. A series of twelve new quaternary derivatives were prepared by quaternization of the quinuclidine nitrogen atom with methyl iodide and selected, differently substituted benzyl bromides (methyl or nitro group and halogen atoms in the meta or para position). The selected compounds have different modifications in the structure to define the structural parameters important for achieving improved biological activity in the interaction with cholinesterases. The melting points of the prepared compounds were determined, and the structure was confirmed by infrared spectroscopy, mass spectrometry and 1D and 2D nuclear magnetic resonance spectroscopy. Furthermore, the inhibitory potential of all prepared compounds toward the enzyme butyrylcholinesterase from horse serum (EC 3.1.1.8) was determined using the Ellman method for colorimetric determination of cholinesterase activity

Estimation of costal cliff retreat rate on the Vrgada Island

Erozija obala, uključujući klifove, predstavlja značajan izazov za obalne zajednice diljem svijeta. Ovaj rad fokusira se na istraživanje brzine erozije klifa na otoku Vrgada, koristeći digitalnu fotogrametriju i softver Agisoft Photoscan i Surfer za izradu digitalnih visinskih modela i razlikovnih digitalnih modela površine. Analiza razlika u visini i volumenu materijala na klifu omogućila je praćenje promjena tijekom zimske i ljetne sezone 2022./2023. godine. Rezultati istraživanja ukazuju na kontinuiranu eroziju klifa, koja se značajno povećala tijekom razdoblja obilnih padalina i jakih vjetrova. Jedni od glavnih čimbenika padinskih procesa na klifu otoka Vrgade su strmi nagib klifa i utjecaj vode koji dovode do osipanja i odrona materijala. Glavni odron materijala u travnju 2023. godine prouzročio je značajne promjene topografije klifa, s naglaskom na nagomilavanju materijala u podnožju klifa i smanjenju padine na vrhu. Kad pogledamo prema budućnosti, sve toplije i olujne klime, podizanje razine mora i povećanje ljudskih aktivnosti neizbježno će utjecati na obalno okružje otoka Vrgada. Ovaj se mijenjajući obalni krajolik izravno povezuje s opasnostima za sigurnost kupača koji često posjećuju područje ispod klifa. Kako erozija ostaje prisutna, postoji sve veća prijetnja padu kamenja i urušavanjima koja bi mogla ugroziti sigurnost ljudi na plaži.Coastline erosion, including cliffs, presents a significant challenge for coastal communities worldwide. This study focuses on investigating the rate of cliff erosion on Vrgada Island, using digital photogrammetry and the Agisoft Photoscan and Surfer software to create digital elevation models and digital surface models of difference. The analysis of height and volume differences in cliff materials allowed for monitoring changes during the winter and summer seasons of 2022/2023. The research results indicate continuous cliff erosion, which significantly intensified during periods of heavy rainfall and strong winds. Among the main factors contributing to slope processes on the cliffs of Vrgada Island are the steep cliff gradient and the influence of water, leading to material collapsing and shedding. The major material collapsing in April 2023 caused significant topographical changes in the cliff, with an emphasis on material accumulation at the base of the cliff and a reduction in the slope at the top. Looking toward the future, the increasingly warmer and stormy climates, rising sea levels, and heightened human activities will inevitably impact the coastal environment of Vrgada Island. This changing coastal landscape is directly linked to risks for the safety of bathers who frequently visit the area beneath the cliff. As erosion continues to be present, there is a growing threat of rockfalls and collapses that could jeopardize the safety of people on the beach

Ultraviolet radiation mutational signatures in human uveal and skin melanoma

Melanomi uvee i kože su dva različita tumorska tipa koja nastaju iz melanocita, stanica odgovornih za sintezu pigmenta melanina. Iako dijele zajedničku ishodišnu stanicu tumora, različiti egzogeni i endogeni čimbenici uzrokuju specifične obrasce mutacija koje nazivamo mutacijskim potpisima. Cilj ovog istraživanja je okarakterizirati i usporediti mutacije i mutacijske potpise prisutne u melanomima uvee i kože s naglaskom na mutacije u genima biomarkerima. Analizirane mutacije dobivene su sekvenciranjem cijelih eksoma 80 pacijenata melenoma uvee i 468 pacijenata melanoma kože, javno dostupnih u TCGA bazi podataka. Mutacijski potpisi pozvani su “refitting”-metodom potpisa COSMIC. Fisherov test primijenjen je za analizu diferencijalno mutiranih gena i biomarkera između melanoma, a Mann-Whitney U test i jednostrani jednoproporcijski Z-test za testiranje povezanosti specifičnih mutacijskih potpisa s mutacijama u genima biomarkerima. Rezultati ukazuju na diferencijalne mutacijske profile oba melanoma, kao i na diferencijalne mutacijske procese uključene u mutagenezu melanoma. Dok u melanomu kože prevladavaju UV mutacijski potpisi (SBS7b i SBS7a) oni su slabo zastupljeni u melanomu uvee. UV zračenje pokazalo se odgovornim za nastanak pokretačkih mutacija u biomarkerima BRAF, NRAS i CDKN2A melanoma kože, dok su za mutagenezu biomarkera melanoma uvee odgovorni drugi mutacijski procesi poput potpisa aristolohične kiseline SBS22 odgovornog za mutagenezu GNA11.Uveal and skin melanoma are two distinct tumor types arising from melanocytes, cells producing melanin pigment. Despite their common cell of origin, different exogenous and endogenous factors lead to specific mutational patterns known as mutational signatures. This research aims to characterize and compare the mutations and mutational signatures in uveal and skin melanoma, focusing on the mutations in the biomarker genes. The analysis examined the whole exome sequencing data of 80 uveal melanoma patients and 468 skin melanoma patients, publicly available in the TCGA database. Mutational signatures were called using the COSMIC signature refitting method. Fisher’s test was applied to analyze the differentially mutated genes and biomarkers between the melanomas, and the Mann-Whitney U test and one-tailed One-Proportion Z-Test to test the correlation between the specific mutational signatures and biomarker mutations. The results indicate differential mutational profiles of the melanomas, as well as differential mutational processes involved in melanoma mutagenesis. While UV mutational signatures (SBS7b and SBS7a) are predominant in skin melanoma, they are poorly represented in uveal melanoma. UV radiation seems to be responsible for driver mutations in the BRAF, NRAS, and CDKN2A biomarkers in skin melanoma, while other mutational processes are responsible for mutagenesis of uveal melanoma biomarkers, such as aristolochic acid signature SBS22 for GNA11 mutagenesis

Molecular-phylogenetic analysis of Palaearctic species of the genus Limnius (Coleoptera: Elmidae)

Rod Limnius Illiger, 1802 pripada porodici vodenih kornjaša Elmidae (hrv. slapoljubi). Do sada je unutar roda Limnius opisano 14 vrsta rasprostranjenih u palearktičkom području, a preliminarne molekularne analize ukazuju na složene sistematske odnose unutar ovoga roda koji zahtijevaju dodatna istraživanja. U sklopu ovog rada odredila sam filogenetske odnose između vrsta unutar roda Limnius za uzorke prikupljene u zapadno-palearktičkoj regiji upotrebom mitohondrijskog gena COI te nuklearnih gena ArgK, H3, CAD i ITS1. Za pojedinačne markere kao i za kombinacije setova markera su konstruirana filogenetska stabla raznim filogenetskim metodama. Rezultati su uglavnom usporedivi između upotrijebljenih setova podataka, no najbolju rezoluciju daje kombinirani set svih molekularnih markera. Dobiveni rezultati potvrđuju prethodne filogenetske hipoteze temeljene na morfologiji o bliskim odnosima vrsta L. volckmari / L. intermedius, L. muelleri / L. satanus, L. opacus / L. subopacus i L. perrisi / L. colchicus, no odnosi između pojedinih skupina vrsta nisu sa sigurnošću potvrđeni. Također, rezultati ukazuju na postojanje određenih kompleksa vrsta kao i neopisanih (kriptičnih) vrsta unutar ovoga roda. Ovo istraživanje daje važan doprinos poznavanju sistematike unutar roda Limnius i predstavlja osnovu za buduća sveobuhvatna taksonomska i filogenetska istraživanja upotrebom integrativnog pristupa.The genus Limnius Illiger, 1802 belongs to the family of aquatic beetles Elmidae (riffle beetles). So far, 14 species distributed in the Palearctic region have been described within the genus, and preliminary molecular analyzes indicate complex systematic relationships that require additional research. In this work I determined the phylogenetic relationships between species within the genus Limnius for samples collected in the Western Palearctic region using the mitochondrial gene COI and the nuclear genes ArgK, H3, CAD and ITS1. Phylogenetic trees were constructed using various phylogenetic methods for individual markers as well as for combinations of marker sets. The results are mostly comparable between the data sets used, but the best resolution is provided using the combined set of all molecular markers. The obtained results confirm previous phylogenetic hypotheses based on morphology about the close relationships of the species L. volckmari / L. intermedius, L. muelleri / L. satanus, L. opacus / L. subopacus and L. perrisi / L. colchicus, but the relationships between the species groups have not been confirmed with certainty. Also, the results indicate the existence of certain species complexes as well as undescribed (cryptic) species within this genus. This research makes an important contribution to the knowledge of systematics within the genus Limnius and represents the basis for future comprehensive taxonomic and phylogenetic studies using an integrative approach