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Interaction of dietary fibers with carotenoids and curcumin from plant extracts: effect on bioavailability
No full textBeta karoten, likopen i kurkumin su biljni pigmenti koji pokazuju veliki antioksidacijski potencijal. Bitni su i često korišteni u održavanju ljudskog zdravlja i sprječavanju nastanka bolesti. Zbog svog antioksidacijskog djelovanja smatraju se nutraceuticima – vrijednim sastavnicama novih oblika enteralne prehrane.
Enteralna prehrana, zbog svoje uloge dopunjavanja ljudske prehrane u stanjima i bolestima kada je to kompromitirano kroz zdravu prehranu, sadržajno je bogata makronutrijentima i mikronutrijentima. Prehrambena vlakna su bitna sastavnica koja osigurava zdravu probavu i može djelovati probiotički, no zbog svojih fizikalno-kemijskih svojstava stupaju u interakcije s karotenoidima i kurkuminom te mogu smanjiti njihovu biodostupnost.
U ovom diplomskom radu ispitana je stabilnost beta karotena, likopena i kurkumina u simuliranim uvjetima probave te biodostupnost s 10 mješavinama prehrambenih vlakana.
Beta karoten i likopen pokazali su stabilnost pri simulaciji uvjeta probave, dok se koncentracija kurkumina značajno smanjila (na oko 70%).
Likopenu je svih 10 ispitivanih mješavina prehrambenih vlakana smanjilo biodostupnost, dok je biodostupnost beta karotena i kurkumina s nekim mješavinama ostala nepromijenjena. Dvije standardne mješavine STAND-2 i STAND-3 imaju značajan utjecaj na smanjenje biodostupnosti sva tri ispitivana spoja. Inovativne mješavine imaju različiti utjecaj na spojeve, ovisno o njihovom sastavu. Sve inovativne mješavine imaju mali utjecaj na biodostupnost beta karotena. Najveća biodostupnost likopena je s mješavinom INOV-1B (98,3%). Najveća biodostupnost kurkumina je uz mješavinu STAND-1 (113,7%). Dobiveni rezultati upućuju na sličnost dviju standardnih mješavina (STAND-2 i STAND-3), a različite sastave ostalih 8 mješavina te različite afinitet prema vezanju ispitivanih spojeva ovisno o sastavu mješavina.Beta carotene, lycopene and curcumin are plant pigments that show great antioxidant potential. They are important in maintaining human health and preventing diseases. Due to their antioxidant effect, they are considered nutraceuticals - valuable components of new formulas of enteral nutrition.
Enteral nutrition, due to its role of supplementing human nutrition in conditions and diseases that compromise ingestion through a healthy diet, is rich in macronutrients and micronutrients. Dietary fibers are an essential component that ensures healthy digestion and can have a probiotic effect, but due to its physical and chemical properties, its interaction with carotenoids and curcumin can reduce their bioavailability.
The stability of beta carotene, lycopene, and curcumin in simulated digestive conditions and their bioavailability as affected by different dietary fibers mixtures were examined in this master thesis.
Beta carotene and lycopene showed stability during the simulation of digestion conditions, while the concentration of curcumin decreased significantly (to about 70%).
The bioavailability of lycopene was reduced by all 10 dietary fiber mixtures that were analyzed, while the bioavailability of beta-carotene and curcumin remained unchanged when analyzed with some of 10 mixtures. Two standard mixtures STAND-2 and STAND-3 have a significant effect on reducing the bioavailability of all three analyzed compounds. Innovative mixtures have different effects on compounds, depending on their composition. All innovative mixtures have little effect on the bioavailability of beta-carotene. The highest bioavailability of lycopene was shown with the mixture INOV-1B (98.3%). The highest bioavailability of curcumin was shown with the STAND-3 mixture (113.7%). With obtained results it can be concluded there is similarity between two standard mixtures (STAND-2 and STAND-3), and there is difference between composition of the other 8 mixtures with different affinity towards the binding of analyzed compounds
N-glycosylation of immunoglobulin A in type 1 diabetes mellitus
No full textŠećerna bolest tipa 1 (T1DM) multifaktorska je, kronična i autoimuna bolest koja nastaje kao posljedica nedostatka
endogenog lučenja inzulina iz β-stanica gušterače. Trenutno su dostupna dva biomarkera koji ukazuju na dugoročnu
kontrolu dijabetesa – HbA1c i fruktozamin, ali oba imaju svoja ograničenja. Stoga, postoji velika potreba za markerom
koji bi na precizniji način omogućio longitudinalno praćenje T1DM.
Promjene u profilu N-glikozilacije ukupnih proteina plazme i imunoglobulina G u mnogim su se studijama pokazale kao
potencijalni prognostički, dijagnostički i terapijski markeri raznih bolesti, posebice onih upalne prirode kao što je T1DM.
Glikozilacija imunoglobulina A u različitim bolestima općenito je manje proučavana, a tek prije nekoliko godina
započela su intenzivna istraživanja uloge njegove glikozilacije u T1DM.
Cilj ovog istraživanja je identificirati promjene u IgA N-glikozilaciji u populaciji odraslih bolesnika s T1DM te ih
usporediti sa zdravim osobama. Analiza glikana učinjena je HILIC-UPLC metodom. U ovom diplomskom radu je
otkriveno da kod odraslih ispitanika s dijagnosticiranim T1DM postoji statistički značajna promjena udjela glikanskih
struktura GP2, GP4, GP5, GP7, GP13, GP14, GP17, GP19, GP21, GP24, GP25, GP27 i GP29. Detaljnije proučavanje
deriviranih svojstava pokazalo je smanjenje udjela nisko razgranatih glikana, a povećanje visoko razgranatih glikana.
Nadalje, smanjenje udjela digalaktoziliranih glikana i povećanje udjela trigalaktoziliranih i trisijaliniziranih glikana
pronađeno je u bolesnika s T1DM. Udio glikana sa sržnom fukozom značajno je smanjen u bolesnika s T1DM, dok je
udio oligomanoznih glikana značajno povećan.
Ove promjene u glikozilaciji IgA upućuju na njegovu moguću ulogu u progresiji bolesti te na potrebu daljnjeg ispitivanja
njegove prognostičke vrijednosti.Type 1 diabetes mellitus (T1DM) is a multifactorial, chronic, and autoimmune disease that results from a lack of
endogenous insulin secretion from pancreatic β-cells. There are two markers currently available that indicate long-term
diabetes control – HbA1c and fructosamine, but both have their limitations. Therefore, there is a great need for a marker
that would show the longitudinal progression of T1DM in a more precise way.
Changes in the N-glycosylation profile of total plasma proteins and immunoglobulin G in many studies have proven to
be a potential prognostic, diagnostic and therapeutic marker of various diseases, especially those of an inflammatory
nature, such as T1DM. Glycosylation of immunoglobulin A in various diseases has generally been less studied, and only
a few years ago, intensive research into the role of its glycosylation in T1DM began.
The aim of this study is to identify changes in IgA N-glycosylation in the population of adult patients with T1DM and
compare them with healthy individuals. Glycan analysis was done by ultra performance liquid chromatography based
on hydrophilic interactions. In this thesis, it was discovered that in adult subjects diagnosed with T1DM there is a
statistically significant change in the proportion of glycan structures GP2, GP4, GP5, GP7, GP13, GP14, GP17, GP19,
GP21, GP24, GP25, GP27 and GP29. A more detailed study of the derived traits showed a decrease in the proportion of
low-branched glycans, and an increase in highly branched glycans. Furthermore, a reduction in the proportion of
digalactosylated glycans and an increase in the proportion of trigalactosylated and trisialinised glycans is found in T1DM
patients. The proportion of glycans with core fucose was significantly reduced in T1DM patients, while the proportion
of high mannose glycans was significantly increased.
These changes in IgA glycosylation indicate that it has a possible role in the disease progression and the need for further
examination of its prognostic value
Frequency of CYP3A4*1B, CYP3A4*22 and CYP3A5*3 polymorphisms in the Croatian population
No full textEnzimi pod-porodice citokroma P450 3A (CYP3A) sudjeluju u metabolizmu oko 50% lijekova u kliničkoj primjeni. Najvažniji su enzimi CYP3A4 i CYP3A5, za koje je karakteristična interindividualna varijabilnost. Genetičke varijante mogu objasniti dio varijabilnosti aktivnosti CYP3A. Klinički najvažnije su varijante CYP3A5*3 i CYP3A4*22, manje važan je polimorfizam CYP3A4*1B. Podaci o učestalosti polimorfizama CYP3A u hrvatskoj populaciji postoje, ali nisu iz istog skupa podataka. U ovom radu su istraživane učestalosti CYP3A4*1B, CYP3A4*22 i CYP3A5*3 u istom skupu podataka u hrvatskoj populaciji.
Uzorci DNA izolirani su iz uzoraka pune krvi tijekom 8 godina rutinskih farmakogenetičkih analiza u Kliničkom bolničkom centru Zagreb. U ovo istraživanje uključen je ukupno 1100 ispitanik. Genotipizacija CYP3A4*1B, CYP3A4*22 i CYP3A5*3 je provedena PCR metodom u stvarnom vremenu TaqMan®.
Učestalosti varijantnih alela bile su za CYP3A4*1B=0.0698 (n=258), CYP3A4*22=0.0355 (n=1100) i CYP3A5*3=0.9645 (n=440). Analiza genotipova polimorfizama pokazala je: (i) za CYP3A4*1B 86,43% ispitanika su nositelji genotipa*1/*1, 13,18% genotipa *1/*1B, a 0,39% genotipa *1B/*1B; (ii) za CYP3A4*22 93,09% su nositelji genotipa *1/*1, 6,73% genotipa *1/*22, a 0,18% genotipa *22/*22; (iii) za CYP3A5*3 83,18% ispitanika su nositelji genotipa *3/*3, 16,14% genotipa *1/*3, a 0,68%, a genotipa *1/*1.
U ovom radu su učestalosti svih ispitivanih polimorfizama su bile nešto veće od prethodnih studija provedenih u hrvatskoj populaciji, jer se radi o pacijentima koji su genotipizirani u kliničkom okružju zbog nuspojava i/ili neučinkovitosti lijekova supstrata CYP3A. Učestalost alela i genotipova varijanti CYP3A4*1B, CYP3A4*22 i CYP3A5*3 u hrvatskoj populaciji u skladu je s ostalim europskim populacijama.Cytochrome P450 3A (CYP3A) subfamily enzymes are involved in the metabolism of about 50% of drugs in clinical use. The most important are CYP3A4 and CYP3A5 enzymes, which are characterised by interindividual variability. Genetic variants can explain part of the variability of CYP3A activity. The clinically most important are variants CYP3A5*3 and CYP3A4*22, while the CYP3A4*1B is less important. Data on the frequency of CYP3A polymorphisms in the Croatian population exist, but they are not from the same dataset. In this study, the frequencies of CYP3A4*1B, CYP3A4*22 and CYP3A5*3 were investigated in the same data set in the Croatian population.
DNA samples were extracted from whole blood samples over eight years of routine pharmacogenetic testing at the University Hospital Centre Zagreb. A total of 1100 subjects were included in this study. Genotyping of CYP3A4*1B, CYP3A4*22 and CYP3A5*3 was performed by the TaqMan® real-time PCR method.
CYP3A allele frequencies were CYP3A4*1B=0.0698 (n=258), CYP3A4*22=0.0355 (n=1100) and CYP3A5*3=0.9645 (n=440). The analysis of genotypes showed: (i) for CYP3A4*1B, 86.43% of subjects were carriers of genotype *1/*1, 13.18% of genotype *1/*1B, and 0.39% of genotype *1B/*1B; (ii) for CYP3A4*22, 93.09% are carriers of the *1/*1 genotype, 6.73% of the *1/*22 genotype, and 0.18% of the *22/*22 genotype; (iii) for CYP3A5*3, 83.18% of respondents are carriers of the *3/*3 genotype, 16.14% of the *1/*3 genotype, and 0.68% of the *1/*1 genotype.
In this study, the frequencies of all investigated polymorphisms were slightly higher than in previous studies conducted on the Croatian population. The frequency of the CYP3A4*1B, CYP3A4*22 and CYP3A5*3 variant alleles and genotypes in the Croatian population is in accordance with the other European populations
Diagnostic accuracy of UK-NEQAS guidelines for cerebrospinal fluid pigment analysis in suspected subarachnoid haemorrhage
No full textOdređivanje hematogenih pigmenata u likvoru važno je za detekciju subarahnoidalnog krvarenja (SAH) kod paci-jenata s visokom kliničkom sumnjom i negativnim nalazima dobivenim slikovnim tehnikama. Cilj ovog istraživa-nja bio je ispitati dijagnostičku točnost UK-NEQAS metode i računskih formula za određivanje hematogenih pig-menata u likvoru pri sumnji na SAH i usporediti ih s vizualnom metodom očitavanja hematogenih pigmenata iz spektra. Istraživanje je provedeno u Kliničkom zavodu za laboratorijsku dijagnostiku KBC-a Zagreb na 37 uzas-topnih pacijenata sa sumnjom na SAH kojima je analiziran likvor dobiven lumbalnom punkcijom. Izmjeren je apsorpcijski spektar supernatanta centrifugiranog likvora u području 250-600 nm te je interpretiran trima meto-dama: vizualnom metodom, izračunom ukupne apsorbancije bilirubina (NBA) i oksihemoglobina (NOA) prema UK-NEQAS-u, te u slučaju pozitivnog NBA UK-NEQAS metodom uz korekciju NBA. SAH je dijagnosticiran referentnom metodom digitalnom subtrakcijskom angiografijom (DSA). Kod 3/37 pacijenata je potvrđen SAH i svim trima metodama detektiran bilirubin u likvoru s jednakom osjetljivosti, a računska metoda prema UK-NEQAS-u pokazala je statistički značajno veću specifičnost od vizualne metode. Lažno pozitivni nalazi posljedica su drugih uzroka krvarenja (prethodne traumatske punkcije, operacija, tromboza). Nije bilo lažno negativnih re-zultata. Cohenovim kappa testom pokazano je snažno podudaranje rezultata između metoda. Računska metoda prema UK-NEQAS smjernicama ima izvrsne dijagnostičke karakteristike za dijagnozu SAH-a i standardizira po-stupak interpretacije apsorpcijskih spektara. Najbolje karakteristike testa za dijagnozu SAH-a ima metoda korigi-ranog NBA. U usporedbi sa vizualnom metodom, daje manje lažno pozitivnih rezultata i pokazuje veću specifič-nost za SAH.Cerebrospinal fluid (CSF) pigment analysis is important for detection of subarachnoid haemorrhage (SAH) in patients with high clinical suspicion and negative findings with imaging techniques. The aim was to exa-mine the diagnostic accuracy of UK-NEQAS method for CSF pigment analysis in suspected SAH and to compare it with the visual assessment of bilirubin from the absorption spectrum. The study included 37 conse-cutive patients with suspected SAH who underwent lumbar puncture to obtain CSF for analysis in Department of Laboratory Diagnostics, UHC Zagreb. UV-VIS spectrophotometric scan of supernatant of centrifuged CSF was performed in the 250-600 nm range. Three methods were used for interpreting absorption curves: visual assessment, net bilirubin (NBA) and oxyhaemoglobin (NOA) absorbance calculation according to UK-NEQAS and NBA correction in case of a positive NBA by UK-NEQAS method. SAH was diagnosed using digital subtraction angiography. Bilirubin was detected by all three methods in 3/37 patients with SAH with equal sensitivity. UK-NEQAS method displayed significantly higher specificity than visual method. False positives were the result of other causes of bleeding (prior traumatic puncture, operation, thrombosis). There were no false-negative results. A strong agreement between the methods was shown using the Cohen kappa test. UK-NEQAS method has great diagnostic characteristics for SAH and standardizes the process of abs-orption curve interpretation. NBA correction method has shown the best diagnostic characteristics for SAH. Compared to the visual method it produces less false-positive results and displays higher specificity for SAH
Synthesis and characterization of amide type harmicenes
No full textβ-karbolinski alkaloidi predstavljaju veliku skupinu prirodnih i sintetskih spojeva. Široko su rasprostranjeni u prirodi te pokazuju širok spektar farmakoloških učinaka poput antimalarijskog, antimikrobnog i citostatskog. S druge strane, ferocen zbog svoje stabilnosti, slabe toksičnosti i oksidoredukcijskih svojstava čini idealnog partnera β-karbolinima u sintezi novih hibridnih spojeva, s obzirom da je dokazano da derivati ferocena pokazuju antimalarijsko i citostatsko djelovanje.
U okviru ovog rada sintetizirana su četiri nova harmicena amidnog tipa, hibridna spoja β-karbolina harmina i ferocenoctene kiseline (6, 11, 15, 18) koji se razlikuju po položaju derivatizacije β-karbolinskog prstena (1, 3, 7 i 9). Prije sinteze hibridnih spojeva, za sintezu derivata u položajima 1 i 3 bilo je potrebno sintetizirati β-karbolinske prstenove 1 i 7 pomoću Pictet-Spenglerove reakcije, dok je za sintezu derivata u položajima 7 i 9 početni spoj bio harmin. U idućim reakcijama u molekulu β-karbolina bilo je potrebno uvesti primarnu amino skupinu, koja u reakciji povezivanja s ferocenoctenom kiselinom daje konačne spojeve, harmicene amidnog tipa. U reakciji povezivanja korišteni su standardni reagensi i uvjeti (HATU/DIEA/sobna temperatura).
Sintetiziranim spojevima strukture su potvrđene standardnim analitičkim i spektroskopskim metodama (1H i 13C NMR, IR te MS), a usto su im određena i tališta. Biološki učinak sintetiziranih spojeva ispitat će se u budućim istraživanjima.Alkaloids of the β-carboline type represent a large group of natural and synthetic compounds. They are widely spread and display a wide array of pharmacological effects such as antimalarial, antimicrobial and cytostatic. On the other hand, ferrocene, owing to its stability, low toxicity and redox properties, is an ideal partner of β-carbolines in the synthesis of new hybrid compounds, since it has been shown that ferrocene derivatives exhibit antimalarial and cytostatic activities.
Here we present synthesis of four new harmicenes of the amide type, i.e. hybrid compounds composed of β-carboline and ferrocene moieties linked via amide bond, in four different positions of the β-carboline ring (1, 3, 7 i 9) (6, 11, 15, 18). Before the synthesis of hybrid compounds in the positions 1 and 3, β-carbolines 1 and 7 were prepared by the means of Pictet-Spengler reaction, whereas harmine was the starting compound for the synthesis of harmicenes prepared in the positions 7 and 9 of the b-carboline ring. Further reactions included introduction of the primary amino group in the b-carboline moiety and coupling reaction with ferroceneacetic acid, by applying standard reagents and conditions (HATU/DIEA/room temperature).
The structures of synthesized compounds were confirmed by standard analytical and spectroscopic methods (1H and 13C NMR, IR and MS), also their melting points were determined. Their biological activities will be evaluated in the future studies
Optimization of extraction and biological activity of black medick (Medicago lupulina L.) polypropylene glycol extracts
No full textStarenje kože progresivan je proces kojeg je moguće usporiti primjenom kozmetičkih pripravaka.
Biljni polifenolni ekstrakti pokazuju širok spektar bioloških učinaka na koži, dobru podnošljivost te ekološku
prihvatljivost, što ih čini idealnim kandidatom za izradu „ anti-aging " kozmetike. Cilj ovog istraživanja bio
je optimizirati ultrazvučnu ekstrakciju zeleni hmeljaste vije u polipropilenglikolu, ekološki prihvatljivom
otapalu niske toksičnosti sa svojstvima humektansa. Također su provedena in vitro biološka ispitivanja s
ciljem vrjednovanja terapijskog i kozmetičkog potencijala ekstrakata. Provođenje ekstrakcije te odabir
ekstrakcij skih parametara prethodno su planirani koristeći Box-Behnkenov dizajn. Kao ekstrakcijski
parametri odabrani su maseni udio PPG, temperatura te vrijeme ekstrakcije. Sadržaj kumestrola u ekstraktima
kvantificiran je pomoću HPLC-a, dok je sadržaj ukupnih polifenola izmjeren spektrofotometrijski.
Višestrukom regresijskom analizom dobivena je polinomna jednadžba koja opisuje utjecaj ekstrakcijskih
parametara na sadržaj kumestrola te ukupnih polifenola. Rješavanjem jednadžbe dobiveni su optimalni
ekstrakcijski uvjeti: 50 % PPG, 80 °C, 3 5 min za kumestrol te 1 0% PPG, 70 °C, 20 min za ukupne polifenole.
Naposlijetku su provedena četiri biološka testa: P-karoten linoleatna analiza, ispitivanje inhibicijskog
djelovanja na lipoksigenazu, ispitivanje inhibicijskog djelovanja na elastazu te ispitivanje inhibicijskog
djelovanja na tirozinazu. Rezultati testova upućuju na izraženo antioksidacijsko te blago protuupalno
djelovanje ekstrakta, pokazujući time potencijal za dermatološku i kozmetičku primjenu.Skin aging is a progressive process which can be delayed by applying cosmetic products. Plant
polyphenolic extracts show a wide range of biological effects on the skin, good tolerabi lity and environmental
acceptability, which makes them an ideal candidate for the preparation of anti-aging cosmetics. The aim of
this study was to optimize ultrasound-assisted extraction of black medick aerial parts using polypropylene
glycol, an environmentally friendly low-toxicity solvent with humectant properties. Furthermore, several in
vitro biological tests have been preformed in order to evaluate therapeutic and cosmetic potential of the
extracts. The extraction and selection of extraction parameters were previously planned using Box-Behnken
design. Mass fraction of PPG, temperature and time were selected as extraction parameters. Coumestrol
content in extracts was quantified using HPLC, while the total polyphenolic content was measured
spectrophotometrically. Multiple regression analysis y ielded a polinomal equation describing the influence
of extraction parameters on coumestrol and total polyphenol content. By solving the equation, optimal
extraction conditions were obtained: 50% PPG, 80 °C, 3 5 min for coumestrol and 1 0% PPG, 70 °C, 20 min
for total polyphenols. Finally, four biological tests were performed: -carotene linoleate analysis,
l ipoxygenase inhibition assay, elastase inhibition assay and tyrosinase inhibition assay. The test results
indicate a pronounced antioxidant and mild anti-inflammatory activity of the extract, thus showing the
potential for dermatological and cosmetic use
Terapijski pristup liječenju kronične opstrukcijske plućne bolesti u vrijeme pandemije COVID-19
No full textChronic obstructive pulmonary disease (COPD) is a common
lung disease that is characterized by long-term respiratory
symptoms and airflow limitation. It is important to investigate whether patients
with COPD are at the increased risk of infection with SARS-Co V -2 and what are
the clinical outcomes of coronavirus disease 2019 (COVID-19). Since both diseases
affect the respiratory tract, the purpose of this review was to determine
whether there is an interrelationship between COPD and COVID-19. Currently,
it is not clear whether COPD patients are at higher risk of SARS-Co V-2 infection;
however, it is confirmed that, if they are affected by COVID-19, there is a
higher rate of hospitalisation and death. COPD patients should continue prescribed
COPD therapy even if they have COVID-19 infection because it is not
confirmed that taking these medications changes the clinical outcomes of
COVID-19 infection. Protective measures are very important in times of higher
prevalence of COVID-19 since they reduce the risk of infection and help COPD
patients maintain their health
Impact of 2-methoxy-1,4-naphtokinone on oxidative stress parameters of radish (Raphanus sativus L.)
No full text2-metoksi-1,4-naftokinon (2-MNQ) je sekundarni metabolit biljaka, gljiva i mikroorganizama. Zbog svojih antikancerogenih, antimetastatskih, antimikrobnih i drugih ljekovitih svojstava predmet je brojnih znanstevnih istraživanja. Cilj ovog diplomskog rada bio je ispitati utjecaj 2-MNQ na fitotoksičnost i parametre oksidacijskog stresa u klijanaca obične rotkvice (Raphanus sativus L.). Sjemenke obične rotkvice bile su izložene tri dana 2-MNQ u koncentracijama 1, 5, 10, 20 i 30 μg/mL. U ispitivanje su bile uključene sjemenke rotkvice izložene tri dana destiliranoj vodi (negativna kontrola) i sjemenke rotkvice izložene tri dana 0,02 M otopini CuSO4 (pozitivna kontrola). Modelni organizam tretiran je u triplikatu. Nakon trodnevna izlaganja izvagana je masa svježega tkiva te su pripremljeni homogenati klijanaca tkiva u trikloroctenoj kiselini (100 mg tkiva/0,5 mL). U supernatantu homogenata određeni su parametri oksidacijskog stresa: glutation (GSH), antocijanini i malondialdehid (MDA). Sadržaj GSH određen je pomoću DTNB reagensa, antocijanina pH diferencijalnom metodom, a MDA pomoću TBA testa spektrofotometrijski. Dobiveni rezultati su statistički obrađeni t-testom (P ≤ 0,05). Trodnevna izloženost 2-MNQ u koncentracijskom rasponu (1-30 μg/mL) nije dovela do statistički značajnog smanjenja mase klijanaca rotkvice u odnosu na negativnu kontrolu, što znači da ispitivane koncentracije 2-MNQ nisu imale fitotoksičan učinak na modelni organizam. Izloženost 2-MNQ u koncentracijama 1-20 μg/mL kroz tri dana dovela je do statistički značajnog pad sadržaja GSH u odnosu na negativnu kontrolu (P < 0,05), dok je 2-MNQ u koncentraciji od 30 μg/mL doveo do porasta sadržaja GSH. To se može objasniti potrošnjom zaliha GSH nakon trodnevne izloženosti nižim koncentracijama 2-MNQ u svrhu obrane od ROS-ova te de novo sintezom GSH kao odgovorom na oksidacijski stres nakon izloženosti višoj koncentraciji 2-MNQ. Značajnija promjena sadržaja antocijanina nakon trodnevne izloženosti 2-MNQ u ispitivanom koncentracijskom rasponu u odnosu na negativnu kontrolu nije zabilježena. Trodnevna izloženost najvišoj koncentraciji 2-MNQ (30 μg/mL) uzrokovala je blagi porast sadržaja MDA u odnosu na negativnu kontrolu, dok je izloženost nižim koncentracijama 2-MNQ (1-20 μg/mL) uzrokovala statistički značajan pad sadržaja MDA. S obzirom da je izloženost nižim koncentracijama 2-MNQ uzrokovala pad sadržaja i GSH i MDA, a viša koncentracija rast njihovog sadržaja, može se pretpostaviti da je rotkvica koristila GSH kao obrambeni sustav te se potrošnjom sadržaja GSH smanjivao i sadržaj MDA. Iz dobivenih rezultata može se zaključiti da ispitivane koncentracije 2-MNQ (1-30 μg/mL) nisu fitotoksične za rotkvicu te da je jedan od mehanizama djelovanja 2-MNQ oksidacijski stres.2-methoxy-1,4-naphthoquinone (2-MNQ) is as secondary metabolite of plants, fungi, and microorganisms. Due to its anticancer, antimetastatic, antimicrobic and other beneficial properties, it has been subject of number of scientific studies. The aim of this study was to explore impact of 2-MNQ on phytotoxicity and parameters of oxidative stress in a plant model, radish (Raphanus sativus L.). Radish seeds were exposed to 2-MNQ at concentrations 1, 5, 10, 20 and 30 μg/mL for 3 days. In the experiment a negative control (radish seeds exposed to distilled water for 3 days) and a positive control (radish seeds exposed to 0.02 M CuSO4 solution for 3 days) were included. The model organism was treated in triplicate. After 3 days of exposure, fresh tissue weight was determined, and plant tissue homogenate was prepared using trichloroacetic acid (100 mg tissue/0.5 mL). As parameters of oxidative stress glutathione (GSH), anthocyanins and malondialdehyde (MDA) were assessed. GSH level was determined using DTNB reagent, anthocyanin level by pH differential method, and MDA with TBA assay, spectrophotometrically. The obtained results were statistically analysed by use of t-test (P <0.05). After 3-day exposure of radish seedlings to 2-MNQ (1-30 μg/mL) no significant reduction in fresh tissue weight was observed, suggesting that 2-MNQ in concentrations applied, is not phytotoxic to the model organism. A significant reduction of GSH level in comparison to the negative control was observed after exposure to 2-MNQ in concentrations 1-20 μg/mL. However, exposure to 2-MNQ at 30 μg/mL for 3-days slightly increase GSH levels. A decrease in GSH level after exposure to lower 2-MNQ concentrations can be explained by its consumption by ROS, while an increase of GSH level after exposure to higher concentrations of 2-MNQ can be explained by its de novo synthesis in response to oxidative stress. No significant change in the level anthocyanins after 3-days exposure to 2-MNQ compared to the negative control was recorded. 3-days exposure to the highest 2-MNQ concentration (30 μg/mL) slightly increased MDA level comparing to negative control, while 2-MNQ at lower concentrations (1-20 μg/mL) significantly reduced MDA level. Similar pattern in the level of GSH and MDA in the radish cells after 3-days exposure to 2-MNQ could be due to utilization of GSH as an antioxidant in order to protect cells form oxidative stress that leads to decrease of MDA level. In conclusion, 2-MNQ in tested concentration range was not phytotoxic to radish, and one of its mechanisms of toxicity is oxidative stress
Determination of arbutin and tannin content in lingonberry leaf, Vaccinium vitis idaea L.
No full textU okviru ovog diplomskog rada provedena je analiza arbutina i trjeslovina u čajevima od listova vrste Vaccinium vitis-idaea (Ericaceae), tj. brusnice. Kvantifikacija arbutina provedena je koristeći farmakopejske metode, UV-Vis spektrofotometriju i tekućinsku kromatografiju visoke djelotvornosti, za određivanje derivata hidrokinona. Čaj od lista brusnice proizvođača Suban sadržava najveći udio hidrokinonskih derivata od svih uzoraka u iznosu od 5,94 % (na 455 nm prema metodi) izraženih kao arbutin, dok čaj proizvođača Agristar omogućuje unos najviše dnevne doze arbutina u iznosu od 251,10 mg. Kvantifikacija trjeslovina provedena je farmakopejskom metodom UV-Vis spektrofotometrije za određivanje trjeslovina. Čaj od lista brusnice proizvođača Suban sadržava najveći udio trjeslovina od svih uzoraka u iznosu od 4,18 % (na 760 nm prema metodi) izraženih kao pirogalol. Rezultati upućuju na to da brusnica ima potencijal za korištenje u liječenju urinarnih infekcija s lakšim i manje učestalim gastrointestinalnim nuspojavama u odnosu na list medvjetke.As part of this diploma thesis, an analysis of arbutin and tannins in teas made from the leaves of the species Vaccinium vitis-idaea (Ericaceae), ie lingonberries, was performed. Quantification of arbutin was performed using pharmacopoeial methods, UV-Vis spectrophotometry and high performance liquid chromatography, for the determination of hydroquinone derivatives. Suban’s lingonberry leaf tea contains the highest proportion of hydroquinone derivatives of all samples in the amount of 5.94% (455 nm according to the method) expressed as arbutin, while Agristar’s tea allows the intake of the highest daily dose of arbutin in the amount of 251.10 mg. Quantification of tannins was performed by pharmacopoeial method of UV-Vis spectrophotometry for the determination of tannins. Suban’s lingonberry leaf tea contains the most tannins of all samples and this was 4.18 % (760 nm according to the method) expressed as pyrogallol. The results suggest that lingonberry has the potential for use in the treatment of urinary tract infections with milder and less frequent gastrointestinal side effects compared to bearberry leaf
Preparation, redox properties and biological activity of ferrocene-nucleobase derivatives
No full textU okviru ove doktorske disertacije pripravljeni su, te eksperimentalnim i teorijskim metodama istraženi,
novi ferocenski derivati različito supstituiranih purinskih nukleobaza s karbonilnom poveznicom između
heterocikličkog i organometalnog dijela. Ferocenoil-purinski derivati 1-31 pripravljeni su reakcijom između
purinskog aniona i ferocenoil-klorida pri čemu su, ovisno o C6 ili C2 supstituentima na nukleobazi, nastali N7 i
N9 regioizomeri, ili isključivo N9 produkti. Pripravljeni su spojevi karakterizirani spektroskopskim i
elektrokemijskim metodama (NMR, IR, UV-Vis, CV i SWV) te spektrometrijom masa (MS), ispitano je njihovo
in vitro citotoksično djelovanje na L929 staničnim linijama i na humanim tumorskim staničnim linijama (A549,
HepG2, PANC-1, MCF-7) te sposobnost stvaranja ROS-a kao mogućeg mehanizma citotoksičnosti. Teorijskim su
pristupom uz upotrebu DFT izračuna opisana fizikalno-kemijska svojstava ferocenskih derivata nukleobaza,
definirani su pogodni modeli za opis njihovih elektrokemijskih svojstva,te je objašnjen reakcijski mehanizma
njihova nastanka i opisana transacilacija u organskim otapalima. Pripravljeni su spojevi elektrokemijski aktivni i
stvaraju ROS acelularno i celularno. Spojevi 11 i 12 pokazali su izraženi citotoksični učinak na svim ispitanim
staničnim linijama, a slijede spojevi 3 i 4, dok je spoj 26 selektivno citotoksičan za HepG2 stanice. Rezultati ovog
istraživanja dobiveni povezivanjem eksperimentalnog i teorijskog pristupa, proširuju spektar pripravljenih i
ispitanih redoks aktivnih ferocenoil-purinskih spojeva kao potencijalnih citostatskih agenasa.In the framework of this doctoral dissertation, new ferrocene derivatives containing variously substituted purine
nucleobases with a carbonyl bond between the two parts were prepared and investigated by the experimental and
theoretical methods. Ferrocenoyl-purine derivatives 1-31 were prepared by nucleophilic substitution reaction on
the carbon atom of the carbonyl group of ferrocenoyl-chloride with purine nucleobase anion generated by the
deprotonation with a strong base. Regioselectivity and regiospecificity of reaction were strongly influenced by the
C6 or C6, C2 substituents of the purine ring. The compounds were characterized by spectroscopic, spectrometric,
and electrochemical methods (NMR, IR, UV-Vis, MS, CV and SWV) and screened in an in vitro cytotoxic assay
against L929 cell lines and human cancer cell lines (A549, HepG2, PANC-1, MCF-7). The ability to generate ROS
was measured as a possible mechanism of cytotoxic action. Using a theoretical approach by the means of DFT
calculations, the physicochemical properties of ferrocene-nucleobase derivatives were predicted and described,
the reaction mechanism of their formation was resolved as well as the transacylation in DMSO. Prepared
compounds showed to be electrochemically active and form ROS in both acellular and cellular media. Compounds
11 and 12 showed enhanced cytotoxic effect against all tested cell lines, followed by compounds 3 and 4, while
compound 26 showed selective cytotoxic effect against HepG2 cell line. The results of this study obtained by
combination of experimental and theoretical approaches, expand the range of prepared and tested redox active
ferrocenoyl-purine compounds as potential cytostatic agents