University of Zagreb
Repository of Faculty of Pharmacy and Biochemistry University of ZgrebNot a member yet
2946 research outputs found
Sort by
Antinociceptive effect of botulinum toxin type A in an animal model of formalin-induced pain
No full textBotulinum toksin tipa A (BT-A) neurotoksin je kojeg sintetizira bakterija Clostridium botulinum. Njegov specifični
mehanizam djelovanja, koji uključuje proteolitičko kidanje SNAP-25 proteina odgovornog za egzocitozu
neurotransmitera, prepoznat je kao moguće farmakološko rješenje nekih poremećaja autonomnog živčanog sustava,
stanja povezanih s hiperkontraktilnošću mišića i određenih vrsta boli. Antinociceptivni učinak BT-A dugo se vremena
smatrao posljedicom inhibicije perifernog lučenja neurotransmitera, no danas se vjeruje da su u pozadini složeni i još
uvijek nerazjašnjeni mehanizmi unutar središnjeg živčanog sustava (SŽS). Cilj ovog istraživanja bio je ispitati
mogućnost transcitoze kao jednog od mehanizama koji doprinosi kompleksnom središnjem djelovanju BT-A na bol.
BT-A (7 i.j./kg) je unilateralno primijenjen u područje brka Wistar štakora jedan dan prije primjene neutralizirajućeg
antitoksina za BT-A (anti-BT-A) (5 i.j./10 μL) u veliku moždanu komoru. Nakon sedam dana od primjene BT-A,
uzrokovana je bol primjenom formalina (2,5 %) te se kroz period od 45 minuta mjerila učestalost ponašanja povezanih
s boli. Periferno primijenjen BT-A smanjio je bol unutar druge faze formalinskog testa koja odgovara ranoj upalnoj
boli. Anti-BT-A poništio je antinociceptivno djelovanje BT-A, što sugerira središnje učinke BT-A na bol posredovane
transcitozom. Budući da primjena kolhicina, blokatora aksonalnog transporta, u trigeminalni ganglij s kontralateralne
strane nije utjecala na antinociceptivni učinak BT-A, isključena je periferna difuzija toksina i sistemsko širenje na
kontralateralnu stranu. Dodatno, primjenom BT-A sa suprotne strane od mjesta indukcije boli pokazan je
antinociceptivni učinak u upalnoj fazi formalinskog testa, što ukazuje da je kontralateralni učinak BT-A na bol
neovisan o mjestu ozljede i djelovanju toksina na strani primjene. Zaključno, rezultati prikazani u ovom diplomskom
radu doprinose razjašnjenju kompleksnog središnjeg djelovanja BT-A na bol. Po prvi puta se predlaže transcitoza u
senzornom sustavu kao važan čimbenik u antinociceptivnom učinku BT-A, što zahtijeva daljnja detaljna istraživanja.Botulinum toxin type A (BoNT-A) is a neurotoxin synthesised by the bacterium Clostridium botulinum. Its specific
mechanism of action, which involves proteolytic cleavage of SNAP-25, a protein responsible for exocytosis of various
neurotransmitters, was recognized as a possible pharmacological solution for some autonomic disorders, conditions
accompanied by muscle hypercontractility and certain types of pain. Antinociceptive effect of BoNT-A was long
considered to be the consequence of solely peripheral inhibition of neurotransmitter exocytosis, but nowadays it is
believed that more complex and still unresolved mechanisms are at play. The aim of this study was to investigate the
possibility of transcytosis as a mechanism underlying the complex central action of BoNT-A on pain. BoNT-A (7
IU/kg) was unilaterally administered in the facial vibrissae of Wistar rats a day before the injection of antibody against
BoNT-A (5 IU/10μL) inside the cisterna magna. After seven days, pain was caused by formalin application (2.5%) and
the frequency of pain-associated behaviours was measured at 5-minute intervals over a period of 45 minutes.
Peripherally applied BoNT-A reduced pain during the second phase of formalin test, corresponding to early
inflammatory pain. Anti-BoNT-A prevented antinociceptive effect of BoNT-A, proposing central effects, mediated by
transcytosis. Since the application of colchicine, an axonal transport blocker, in the contralateral trigeminal ganglion
did not affect the antinociceptive effect of BoNT-A, possibility of peripheral diffusion of toxin and systemic spread to
the contralateral side was excluded. Additionally, when BT-A was applied on the opposite side of the pain induction
site, an antinociceptive effect was observed during the inflammatory phase of the formalin test, indicating that the
contralateral effect of BT-A on pain is independent of the site of injury and the action of toxin on the site of
application. In conclusion, the results presented in this master’s thesis contribute to the elucidation of the complex
central action of BoNT-A on pain. For the first time, transcytosis in the sensory system is proposed as an important
factor in the antinociceptive effect of BT-A, which requires further detailed investigation
Interaction of dietary fibers with beta carotene: effect on in vitro bioavailability
No full textBiodostupnost beta-karotena vrlo je varijabilna te uvelike ovisi o uvjetima okoline i interakcijama s ostalim spojevima
prisutnim u matriksu namirnice. Značajan utjecaj na biodostupnost mogu imati prehrambena vlakna zbog mogućnosti
stvaranja kompleksa s beta-karotenom i drugih interakcija.
Osnovna hipoteza ovog rada je bila da način interakcije prehrambenih vlakana s beta-karotenom i posljedični učinak na
biodostupnost značajno ovise o fizikalno-kemijskim svojstvima prehrambenih vlakana. Zbog toga je u okviru rada
istražen kapacitet vezanja prehrambenih vlakana različite topljivosti (za beta-karoten) te utjecaj smjesa vlakana
(različitog omjera topljivih i netopljivih vlakana) na in vitro biodostupnost cis- i trans- izomera beta-karotena iz realnog
matriksa namirnice. Biodostupnost beta-karotena određena je in vitro statičkom simulacijom probave prema
standardiziranoj metodi, a koncentracije beta-karotena su određene spektrofotometrijski odnosno primjenom visokorazdjelne tekućinske kromatografije (HPLC).
Rezultati su pokazali da fizikalno-kemijska svojstva prehrambenih vlakana značajno utječu na sposobnost vezanja betakarotena, pri čemu netopljiva vlakna tipa celuloze imaju najizraženije negativne učinke. Smjese prehrambenih vlakana
s pretežno netopljivom komponentom imaju negativan učinak na in vitro biodostupnost beta-karotena dok taj učinak
kod smjesa, koje se sastoje od pretežno topljivih vlakana, izostaje. Utjecaj je izraženiji na trans- izomere
beta-karotena.
Navedeni rezultati značajno doprinose vrlo ograničenim saznanjima o utjecaju prehrambenih vlakana na biodostupnost
beta-karotena, a osobito su relevantni u kontekstu pametnog dizajna funkcionalne hrane koja zahtijeva visok sadržaj
prehrambenih vlakana uz istovremeno zadržavanje zadovoljavajuće biodostupnosti bioaktivnih sastavnica kao što je
beta-karoten.The bioavailability of beta-carotene is highly variable and largely depends on environmental conditions and interactions
with other compounds present in the food matrix. Dietary fibers can have a significant impact on bioavailability due to
the possibility of formation of complex compounds with beta-carotene and other interactions.
The basic hypothesis of this work was that the way dietary fibers interact with beta-carotene (and the consequent effect
on bioavailability) significantly depends on the physico-chemical properties of dietary fibers. For this reason, the binding
capacity of different solubility (for beta-carotene) and the influence of fiber mixtures (with different ratios of soluble
and insoluble fibers) on the in vitro bioavailability of cis- and trans- isomers of beta carotene from the real food matrix
were investigated in this work. The bioavailability of beta-carotene was determined by in vitro static digestion simulation
according to a standardized method, and beta-carotene concentrations were determined spectrophotometrically, i.e.
using high-resolution liquid chromatography (HPLC).
The results showed that the physico-chemical properties of dietary fibers significantly affect the ability to bind betacarotene, with insoluble cellulose-type fibers having the most pronounced negative effects. Dietary fiber mixtures with
a predominantly insoluble component have a negative effect on the in vitro bioavailability of beta-carotene, while this
effect is absent in mixtures consisting of predominantly soluble fibers. The effect is more pronounced on the transisomers of beta-carotene.
The above results significantly contribute to the very limited knowledge about the influence of dietary fibers on the
bioavailability of beta-carotene and are particularly relevant in the context of the smart design of functional food that
requires a high content of dietary fiber while maintaining satisfactory bioavailability of bioactive components such as
beta-carotene
Analytical verification of the latex-enhanced immunoturbidimetric D-dimer assay on the Alinity C clinical chemistry analyzer
No full textD-dimeri su razgradni produkti umreženog fibrina koji nastaju procesom fibrinolize. Njihovo određivanje iznimno je
važno za isključivanje venske tromboembolije (VTE) koja čini treću vodeću kardiovaskularnu dijagnozu. Testovi za
određivanje D-dimera temelje se na imunokemijskim metodama koje koriste monoklonska protutijela za specifične
epitope na fragmentima D-dimera. Zbog potrebe za pouzdanim rezultatima u što kraćem vremenu, na tržištu je dostupan
sve veći broj potpuno automatiziranih testova za određivanje D-dimera pri čemu većinu tih testova čine lateks
imunoturbidimetrijski testovi (LPIA). Cilj ovog rada bio je provesti analitičku verifikaciju lateks imunoturbidimetrijske
metode, Abbotova testa za D-dimere, na biokemijskom analizatoru Alinity c. Verifikacija je obuhvatila ispitivanje
preciznosti te usporedbu s INNOVANCE testom za D-dimere koji se koristi u rutinskom radu na koagulacijskom
analizatoru. Koeficijenti varijacije dobiveni ispitivanjem preciznosti metode zadovoljili su optimalne kriterije
prihvatljivosti prema EFLM bazi biološke varijabilnosti. Između Abbottova testa za D-dimere i INNOVANCE testa za
D-dimere utvrđena je snažna pozitivna korelacija (ρ = 0,975) bez značajnog odstupanja u linearnosti. Metode su pokazale
visoku usporedivost te je utvrđeno postojanje statistički značajnog konstantnog i proporcionalnog odstupanja koje nije
klinički značajno. Na temelju provedenih analiza i dobivenih rezultata zaključeno je da je ispitivana lateks
imunoturbidimetrijska metoda za određivanje D-dimera prikladna za uvođenje u rutinski rad laboratorija.D-dimers are degradation products of cross-linked fibrin formed during the process of fibrinolysis. Their determination
is crucial for excluding venous thromboembolism (VTE), which is the third leading cardiovascular diagnosis. D-dimer
assays are based on immunochemical methods utilizing monoclonal antibodies targeting specific epitopes on D-dimer
fragments. Due to the need for reliable results in the shortest possible time, an increasing number of fully automated Ddimer assays are available on the market, the majority of which are latex-enhanced immunoturbidimetric assays (LPIA).
The aim of this study was to conduct an analytical verification of the latex-enhanced immunoturbidimetric method,
Abbott D-dimer assay, on the Alinity c clinical chemistry analyzer. The verification included precision testing and
comparison with the INNOVANCE D-dimer assay used in routine work on the coagulation analyzer. The coefficients
of variation obtained from precision testing met the optimal acceptance criteria according to the EFLM database of
biological variability. A strong positive correlation (ρ = 0.975) was found between Abbott's D-dimer assay and the
INNOVANCE D-dimer assay, with no significant deviation in linearity. Both methods demonstrated high comparability,
and a statistically significant constant and proportional deviation was observed, though it was not clinically significant.
Based on the conducted analyses and obtained results, it was concluded that the tested latex-enhanced
immunoturbidimetric method for D-dimer determination is suitable for implementation in routine laboratory practice
Detection of JAK2 V617F mutation in liquid biopsy samples of patients with colorectal cancer using digital polymerase chain reaction method
No full textKolorektalni karcinom (CRC) zloćudni je tumor debelog i završnog crijeva koji prema statističkim podatcima
predstavlja jedan od vodećih uzroka smrti od malignih oboljenja u svijetu. Bolest se često dijagnosticira tek
u kasnijim stadijima, a rano otkrivanje značajno utječe na poboljšanje izgleda za uspješno izlječenje. Tekuća
biopsija omogućila je detekciju specifičnih tumorskih biljega na neinvazivan način, a kao najkorisniji biljeg
pokazala se slobodna cirkulirajuća DNA (ccfDNA). U pacijenata s malignim oboljenjem dolazi do
oslobađanja cirkulirajuće tumorske DNA s očuvanim genskim i epigenskim karakteristikama tumora što
omogućuje praćenje dinamike bolesti praćenjem koncentracije i molekularnog profila ccfDNA. Signalni put
JAK2/STAT3 ima značajan utjecaj na proliferaciju tumorskih stanica i na metastatski potencijal tumora i
korisna je terapijska meta u liječenju malignih oboljenja. S obzirom da je najčešća mutacija JAK2 V617F
karakterizirana hiperaktivacijom JAK2, a posljedično i STAT3, cilj ovoga rada bio je utvrditi prisutnost
navedene mutacije u uzorcima ccfDNA 31 ispitanika s dijagnozom CRC-a. Zbog niskih koncentracija
ccfDNA u uzorcima tijekom izrade rada korištena je metoda digitalne lančane reakcije polimerazom (dPCR)
na sustavu QuantStudio™ 3D Digital PCR System (Applied Biosystems, Foster City, Kalifornija, SAD).
Rezultati su očitani mjerenjem fluorescencijskih signala VIC™ i FAM™ s obilježenih sondi gdje je VIC™
vezana za sondu specifičnu za alel divljeg tipa, a FAM™ za mutirani alel. Obradom i pregledom rezultata u
računalnom programu QuantStudio™ 3D AnalysisSuite Cloud Software (Applied Biosystems, Foster City,
Kalifornija, SAD) utvrđeno je da mutacija JAK2 V617F u uzorcima ccfDNA ispitanika s dijagnozom CRCa
nije prisutna.Colorectal cancer (CRC) is a malignant tumour of the colon and rectum that, according to statistical data, is
one of the leading causes of cancer-related deaths worldwide. The disease is often diagnosed at later stages,
while early detection significantly improves the chances of successful treatment. Liquid biopsy has enabled
the detection of specific tumour markers in a non-invasive manner, with cell-free circulating DNA (ccfDNA)
emerging as the most useful marker. In patients with malignancies, circulating tumour DNA is released,
maintaining the genetic and epigenetic characteristics of the tumour, which allows for disease monitoring by
tracking the concentration and molecular profile of ccfDNA. The JAK2/STAT3 signaling pathway has a
significant impact on the tumour cell proliferation and the metastatic potential of the tumour, making it a
valuable therapeutic target in the treatment of malignancies. Since the most common mutation, JAK2 V617F,
is characterized by hyperactivation of JAK2 and, consequently, STAT3, the aim of this study was to
determine the presence of this mutation in ccfDNA samples from 31 CRC patient. Due to the low
concentrations of ccfDNA in the samples, digital polymerase chain reaction (dPCR) was used, performed on
the QuantStudio™ 3D Digital PCR System (Applied Biosystems, Foster City, California, USA). The results
were analysed by measuring the fluorescence signals of VIC™ and FAM™ dyes bound to the labeled probes,
where VIC™ is bound to the probe specific for the wild-type allele and FAM™ to the probe specific for the
mutated allele. The results were processed and analysed using the QuantStudio™ 3D AnalysisSuite Cloud
Software (Applied Biosystems, Foster City, California, USA) and it has been concluded that the JAK2 V617F
mutation is not present in the ccfDNA samples of the CRC patients
Sigurna primjena i učestalost nuspojava beta-blokatora u liječenju dojenačkih hemangioma
No full textCilj istraživanja: Rad istražuje primjenu beta-blokatora, s naglaskom na propranolol i timolol, u liječenju
dojenačkih hemangioma (IH). Propranolol je lijek prvog izbora u terapiji dojenačkih hemangioma zbog svoje
sposobnosti brzog smanjenja njihove veličine i intenziteta boje, dok je timolol pokazao visoku učinkovitost u
liječenju manjih, površinskih hemangioma. Primjena ovih lijekova rezultirala je uglavnom dobrim ili odličnim
terapijskim odgovorom, uz minimalne nuspojave. Rad obuhvaća procjenu sigurnosnog profila ovih lijekova i
analizu nuspojava prijavljenih u EudraVigilance bazi podataka te naglašava ulogu kliničkog farmaceuta u
prevenciji nuspojava, edukaciji roditelja i prilagodbi terapije.
Materijal: Rad se oslanja na podatke iz sustavnog pregleda studija o pacijentima liječenim propranololom i
timololom za dojenačke hemangiome. Proveden je i pregled dosadašnjih prijavljenih nuspojava u EudraVigilance
bazi podataka, uz usporedbu s dostupnim literaturnim izvorima. Naglasak je stavljen na doziranje propranolola,
praćenje nuspojava te individualizirani pristup terapiji, osobito kod pacijenata s komorbiditetima.
Rezultati: Propranolol je pokazao visoku učinkovitost u brzom smanjenju veličine i boje hemangioma, dok je
timolol bio uspješan u liječenju manjih, površinskih dojenačkih hemangioma. Nuspojave su uglavnom bile
minimalne i nisu zahtijevale prekid terapije, ali su prijavljeni i slučajevi ozbiljnijih nuspojava poput hipotenzije,
bradikardije i hipoglikemije. Pregledom studija pokazana je važnost postupne prilagodbe doze, koja se pokazala
učinkovitom u svrhu boljeg podnošenja terapije i smanjenja nuspojava.
Zaključak: Beta-blokatori, posebno propranolol i timolol, učinkoviti su u liječenju dojenačkih hemangioma, s
dobrim sigurnosnim profilom. Preporučuje se kontinuirano praćenje pacijenata i prilagodba terapije, osobito kod
pacijenata s komorbiditetima. Potrebna su daljnja istraživanja kako bi se ispitali dugoročni učinci terapije i
usporedila primjena beta-blokatora s drugim terapijskim opcijama.Objectives: This paper investigates the use of beta-blockers, with an emphasis on propranolol and timolol, in the
treatment of infantile hemangiomas (IH). Propranolol is the drug of first choice in the treatment of infantile
hemangiomas due to it's ability to rapidly reduce their size and color intensity, while timolol has shown high
efficacy in the treatment of smaller, superficial hemangiomas. The use of these drugs resulted in mostly good or
excellent therapeutic response, with minimal side effects. The paper includes an assessment of the safety profile
of these drugs and an analysis of side effects reported in the EudraVigilance database, and emphasizes the role of
the clinical pharmacist in the prevention of side effects, parent education and therapy adjustments.
Material: The paper used data from a systematic review of relevant studies that included patients treated with
propranolol and timolol for infantile hemangiomas. An overview of previously reported side effects in the
EudraVigilance database was also conducted, with a comparison with available literature sources. Emphasis is
placed on the dosing of propranolol, the monitoring of side effects and the individualized approach to therapy,
especially in patients with comorbidities.
Results: Propranolol showed high efficacy in rapidly reducing the size and color of hemangiomas, while timolol
was successful in the treatment of smaller, superficial infantile hemangiomas. Side effects were mostly minimal
and did not require discontinuation of therapy, but cases of more serious side effects such as hypotension,
bradycardia, and hypoglycemia were also reported. A review of studies showed the importance of gradual dose
adjustment, which proved to be effective in order to better tolerate therapy and reduce side effects.
Conclusion: Beta-blockers, especially propranolol and timolol, are effective in the treatment of infantile
hemangiomas, with a good safety profile. Continuous monitoring of patients and adjustment of therapy is
recommended, especially in patients with comorbidities. Further studies are needed to examine the long-term
effects of therapy and to compare the use of beta-blockers with other therapeutic options
In vitro mucoadhesion studies of nasal pectin gels
No full textNazalna dostava lijekova je brzo rastuće područje. Mukoadhezivni i/ili in situ gelirajući polimeri omogućuju nazalnim formulacijama prevladavanje učinka mukocilijarnog čišćenja, produljenim zadržavanjem lijeka na sluznici. Procjena mukoadhezivnosti stoga je važan korak u razvoju nazalnih farmaceutskih oblika. In vitro/ex vivo metode razvijaju se kao prikladnije alternative in vivo ispitivanjima te danas postoji velik broj različitih metoda za tu namjenu, uključujući metode vlačnog naprezanja i nagnute ploče. Složeni proces mukoadhezije ograničava predviđanja in vitro ispitivanjima, a nedostatak standardizacije metoda otežava usporedbu rezultata između različitih studija. Prema tome, bolje poznavanje postojećih metoda i mehanizama mukoadhezije potrebno je za unaprjeđenje ispitivanja i optimizaciju nazalnih formulacija.
Cilj ovog rada bio je evaluirati metodu nagnute ploče za ispitivanje mukoadhezivnosti pektinskih gelova. Tumačenje je upotpunjeno testom vlačnog naprezanja na analizatoru teksture, kojim je ispitan i utjecaj uklapanja lijeka na mukoadhezivnost gelova, te rotacijskim reološkim testom. U tu svrhu pripremljeni su pektinski in situ gelirajući sustavi, koncentracije 1,0 i 1,5 % (m/m), bez i sa uklopljenim prednizolonom (0,2 %, m/m), i kitozanski gel (2,5 %, m/m) kao kontrolni uzorak. Geliranje pektinskih sustava potaknuto je miješanjem sa SNF-om u volumnom omjeru 1:1. Metoda nagnute ploče pokazala se prikladnom za ispitivanje mukoadhezivnosti pektinskih gelova. Omogućila je diferencijaciju pektinskih gelova s obzirom na mukoadhezivnost, pri čemu se mukoadhezivnijim pokazao gel pripravljen iz koncentriranije otopine pektina. S druge je strane potvrđena njena ograničena primjena u ispitivanju mukoadhezivnosti viskoznih uzoraka, kao što je kitozanski gel, zbog značajnog utjecaja reoloških svojstava na profil zadržavanja na nagnutoj ploči. Istaknuta je i potreba za optimiranjem izvedbenih parametara metode, poput kuta nagiba i količine uzorka, prema ispitivanoj formulaciji. Testom vlačnog naprezanja potvrđena je mukoadhezivnost ispitivanih gelova, a razlike u rezultatima u odnosu na metodu nagnute ploče pripisane su različitim temperaturama ispitivanja (redom 25°C i 34°C). Utjecaj uklapanja lijeka na mukoadhezivnost pektinskih gelova ovisio je o koncentraciji pektina u geliranom sustavu. Reološkim ispitivanjem potvrđeno je pseudoplastično ponašanje gelova i veća osjetljivost pektinskih gelova na deformaciju u odnosu na kitozanski gel. Pad temperature imao je izraženiji utjecaj na povećanje viskoznosti koncentriranijeg pektinskog gela. Kombinacija metode nagnute ploče, vlačnog naprezanja i reološkog ispitivanja omogućila je bolje razumijevanje mukoadhezivnog karaktera gelova i čimbenika koji na nju utječu.Nasal drug delivery is a rapidly growing field. Mucoadhesive and/or in situ gelling polymers help nasal formulations overcome the mucociliary clearance by prolonging drug retention on the mucosa. Assessing mucoadhesion is, therefore, crucial in nasal dosage form development. In vitro/ex vivo methods are being developed as more suitable alternatives to in vivo testing, and today, a large number of different methods exist for this purpose, including tensile stress and inclined plane methods. The complexity of mucoadhesion limits the predictive ability of in vitro tests, and the lack of standardization among methods makes it difficult to compare results across different studies. Thus, a better understanding of existing methods and the mechanisms of mucoadhesion is necessary to improve testing and optimize nasal formulations.
The aim of this study was to evaluate the inclined plane method for assessing the mucoadhesiveness of pectin gels. The interpretation was supplemented with a tensile stress test using a texture analyzer, which also examined the effect of drug incorporation on gel mucoadhesiveness, as well as a rotational rheological test. For this purpose, in situ gelling pectin systems were prepared at concentrations of 1.0 % and 1.5 % (w/w), with and without incorporated prednisolone (0.2 %, w/w), along with a chitosan gel (2.5 %, w/w) as a control sample. Gelation of the pectin systems was induced by mixing with simulated nasal fluid (SNF) in a 1:1 volume ratio. The inclined plane method proved to be suitable for assessing the mucoadhesiveness of pectin gels. It enabled differentiation of pectin gels based on their mucoadhesiveness, with the gel prepared from the more concentrated pectin solution exhibiting greater adhesion. However, its limited applicability to highly viscous samples, such as chitosan gel, was confirmed due to the significant influence of rheological properties on the retention profile on the inclined plane. The need for optimizing methodological parameters, such as the angle of inclination and sample volume, according to the tested formulation, was also highlighted. The tensile stress test confirmed the mucoadhesiveness of the tested gels, while differences in results compared to the inclined plane method were attributed to different testing temperatures (25°C and 34°C, respectively). The effect of drug incorporation on the mucoadhesiveness of pectin gels depended on the pectin concentration in the gelled system. Rheological test confirmed the pseudoplastic behavior of the gels and showed greater sensitivity of pectin gels to deformation compared to the chitosan gel. A decrease in temperature had a more pronounced effect on increasing the viscosity of the more concentrated pectin gel. The combination of the inclined plane method, tensile stress testing, and rheological test provided a better understanding of the mucoadhesive properties of the gels and the factors influencing them
Evaluation of computational methods in mass spectrometry
No full textTekućinska kromatografija visoke djelotvornosti povezana s masenom spektrometrijom je danas najčešće korištena tehnika za identifikaciju i određivanje aktivne farmaceutske tvari i njenih onečišćenja u sirovinama, farmaceutskim formulacijama i gotovim proizvodima. Rad na ovoj naprednoj tehnici zahtjeva posebnu edukaciju analitičara te bogato iskustvo rada u području kromatografije i masene spektrometrije. Kako bi se analitičarima olakšao rad na instrumentu kao i tumačenje dobivenih rezultata posljednjih godina razvijen je niz računalnih programa koji simuliraju masene spektre te predlažu strukture nastalih iona kao i puteve fragmentacije.
Cilj ovoga rada bio je koristeći se HPLC-ESI-MS/MS tehnikom provesti analizu lijeka koji se koristi u liječenju upalnih bolesti crijeva sulfasalazina. Nadalje, pomoću računalnih programa prikupljeni su simulirani maseni spektri te predložene strukture molekula koje nastanu fragmentacijom. Istraživanje je obuhvatilo detaljnu usporedbu eksperimentalno dobivenih masenih spektara te istih dobivenih simulacijom računalnim programima.High performance liquid chromatography coupled with mass spectrometry is today the most commonly used technique for the identification and determination of active pharmaceutical substances and their contamination in raw materials, pharmaceutical formulations and finished products. Working on this advanced technique requires special training of the analyst and extensive work experience in the field of chromatography and mass spectrometry. In order to make it easier for analysts to work on the instrument as well as to interpret the obtained results, a number of computer programs have been developed in recent years that simulate mass spectra and present the structure of the occurred ions as well as the fragmentation pathways.
The aim of this work was to use the HPLC-ESI-MS/MS technique to analyze the drug sulfasalazine, which is used in the treatment of inflammatory bowel diseases. Furthermore, computer programs were used to collect simulated mass spectra and proposed structures of molecules resulting from fragmentation. The research included a detailed comparison of the experimentally obtained mass spectra and the same obtained by simulation with computer programs
Determination of metal content in immortelle flower and stem using EDXRF, TXRF AND ICP-MS techniques
No full textSmilje (Helichrysum italicum (Roth) G. Don) je mediteranska cvjetnica koja se zbog svojih ljekovitih i aromatičnih svojstava koristi za izradu farmaceutskih i kozmetičkih pripravaka. Razvoj brze i osjetljive tehnike elementne analize biljnog materijala mogao bi značajno doprinijeti kakvoći i stabilnosti pripravaka te smanjiti nepotrebno izlaganje potencijalno toksičnim teškim metalima. Konvencionalne tehnike elementne analize skupe su, dugotrajne i zahtijevaju prethodnu razgradnju složenog biljnog matriksa. TXRF je mlada analitička tehnika koja vodi brigu o načelima ,,zelene kemije“, zahtjeva male količine uzorka i reagensa, a omogućava brzu i osjetljivu analizu stoga bi se mogla primjenjivati i u rutinskim analizama. Cilj ovog rada bila je usporedba rezultata dobivenih određivanjem sadržaja elemenata (Fe, Mn, Ti, Cr, Ni, Cu, Zn, Pb, Br, Se, Rb i Sr) prisutnih u uzorcima smilja primjenjujući različite analitičke metode pripreme uzoraka (suspenzija, mikrovalna digestija i pelet) i analitičke tehnike (TXRF, EDXRF i ICP-MS). Dobiveni rezultati pokazali su kako primijenjene tehnike kao ni različite metode pripreme uzoraka ne utječu značajno na rezultate analize. Pritom se TXRF pokazala kao dobra alternativna analitička tehnika budući da omogućava identifikaciju i kvantifikaciju vrlo niskih koncentracija elemenata prisutnih u uzorku bez prethodne složene kemijske obrade.Immortelle (Helichrysum italicum (Roth) G. Don) is a Mediterranean flowering plant that, due to its medicinal and aromatic properties, is used to make pharmaceutical and cosmetic preparations. The development of a fast and sensitive technique for elemental analysis of plant material could contribute significantly to the quality and stability of preparations and reduce unnecessary exposure to potentially toxic heavy metals. Conventional elemental analysis techniques are expensive, time-consuming and require prior decomposition of the complex plant matrix. TXRF is a young analytical technique that incorporates the principles of "green chemistry", requires small amounts of samples and reagents, and provides fast and sensitive analysis, so that it could be used in routine analyses. The aim of this work was to compare the results obtained by determining the content of elements (Fe, Mn, Ti, Cr, Ni, Cu, Zn, Pb, Br, Se, Rb and Sr) present in immortelle samples using different analytical methods of sample preparation (suspension, microwave digestion and pellet) and analytical techniques (TXRF, EDXRF and ICP-MS). The obtained results showed that the techniques used as well as the different sample preparation methods did not significantly affect the analytical results. Thus, TXRF proved to be a good alternative analytical technique, since it allows the identification and quantification of very low concentrations of the elements present in the sample without prior complex chemical processing
Development and validation of a self-reported medical adherence tool
No full textUnatoč dostupnosti raznih upitnika za mjerenje adherencije prema uzimanju lijekova, učinkovito
identificiranje stupnja neadherncije i njenih uzroka u praksi i dalje predstavlja izazov. Postojećim upitnicima
često nedostaje jednostavnost primjene koja je nužna za upotrebu u kliničkoj praksi. Cilj ovog diplomskog
rada bio je razviti i validirati korsnicima pristupačan upitnik koji zdravstvenim radnicima daje koncizno, a
sveobuhvatno sredstvo za identifikaciju ponašanja vezanih uz adherenciju. Metodologija se sastojala od dvije
faze: upitnik je prvo izrađen korištenjem metode nominalne skupine u kojoj su bili zdravstveni radnici, nakon
čega je uslijedilo presječno istraživanje koje je uključivalo punoljetne stanovnike Hrvatske. Analiza
valjanosti ukazala na prihvatljivu izravnu i sadržajnu valjanost te zadovoljavajuću kriterijsku valjanost, pri
čemu su rezultati „Attitudes towards meDication adHerence self-Reported questionnairE“ (ADHERE-7)
korelirali i sa rezultatima „Medication Adherence Report Scale“ (MARS-5 upitnik) (ϱ = 0.765; p < 0.001) i
sa objektivnom mjerom udjela dana pokrivenih terapijom (engl. proportion of days covered, PDC)
izračunatim pomoću povijesti podizanja lijekova iz kartica pacijenata iz ljekarni (G = 0,586; p = 0,015).
Analiza konstruktne valjanosti otkrila je tri faktora: Neadherenciju uzrokovanu obojnošću, Neadherenciju
uslijed lagode i Praktičnu neadherenciju. Prosječni rezultat ADHERE-7 upitnika je bio 26,27 ± 2,41 (raspon
od 17 do 28). Ovaj robusni proces validacije potvrdio je da je ADHERE-7 upitnik pouzdan instrument za
procjenu adherencije prema uzimanju lijekova, procjenu problema vezanih uz odbojnost, lagodu i praktičnost
te za procjenu namjerne i nenamjerne neadherencije.Despite the availability of various tools for measuring medication adherence, efficiently identifying nonadherence
levels and reasons at the point of care remains challenging. Existing tools often lack the ease of
use needed for practical clinical application. This study aimed to develop and validate a user-friendly tool to
provide healthcare professionals with a concise yet comprehensive means of identifying adherence behaviors.
The methodology consisted of two phases: tool items were first developed using the nominal group technique
with healthcare professionals, followed by a cross-sectional pilot study involving community-dwelling adults
in Croatia. Validation analysis indicated acceptable face and content validity and satisfactory criterion
validity, with Attitudes towards meDication adHErence self-Reported questionnairE (ADHERE-7) scores
correlating with both the self-reported five-item Medication Adherence Report Scale (MARS-5 tool) (ϱ =
0.765; p < 0.001) and an objective measure of the proportion of days covered (PDC) from pharmacy
prescription claims data (G = 0.586; p = 0.015). Construct validity revealed three factors: Aversion, Comfort,
and Practical Non-Adherence, with Cronbach’s alpha values of 0.617 for Aversion and 0.714 for Comfort
Non-Adherence. The mean total score for ADHERE-7 was 26.27 ± 2.41 (range 17 to 28). This robust
validation process confirms the ADHERE-7 tool as a reliable instrument for assessing medication adherence,
addressing aversion, comfort, practical issues, and both intentional and unintentional nonadherence
Antimicrobial effect of carvacrol and oregano essential oil (O. compactum) on the bacterium Staphylococcus aureus
No full textZbog sve većeg razvoja antimikrobne rezistencije (AMR) intenzivno se istražuju različite alternative antibioticima ili
potencijalne kombinacije s prirodnim tvarima koje bi smanjile korištenje antibiotika. Eterična ulja i njihove sastavnice
pokazale su se u brojnim istraživanjima kao potentne aktivne tvari svojim antimikrobnim, ali i antioksidativnim i protuupalnim
djelovanjem. Mupirocin je jedan od najčešće korištenih topikalnih antibiotika pogotovo za infekcije kože uzrokovane S.
aureusom. Razvojem posebnih formulacija antibiotika zajedno s eteričnim uljima ili njihovim sastavnicama mogao bi se postići
sinergistički učinak u vidu korištenja manjih koncentracija antibiotika i pojačanog antimikrobnog djelovanja takvih
formulacija. Dok antibiotici ciljaju specifične bakterijske procese, eterična ulja narušavaju integritet bakterijskih stanica i
utječu na smanjenje produkcije biofilma, čime se pojačava ukupni antimikrobni učinak. Kako bi se uopće započelo s razvojem
takvih formulacija, potrebno je utvrditi učinkovitost djelovanja eteričnog ulja i njegovih sastavnica te kombinacija s
antibiotikom. Stoga je cilj ovoga rada bio utvrditi antimikrobni potencijal eteričnog ulja origana (O. compactum) i njegove
glavne sastavnice karvakrola pojedinačno i u kombinacijama s antibiotikom mupirocinom. U radu su ispitivane MIC
vrijednosti mupirocina (MUP), karvakrola (KAR) i eteričnog ulja origana (ET.U.) pojedinačno i u kombinacijama. MIC
mupirocina za S. aureus sojeve kretao se u rasponu 0,076 – 0,141 μg/mL, dok su MIC-ovi karvakrola i eteričnog ulja origana
bili nekoliko redova veći: karvakrol 525 – 700 μg/mL, eterično ulje origana 625 – 750 μg/mL. U ispitivanju kombinacija MUP
i KAR/ET.U. na soju S. aureus NCTC 12493 dokazan je aditivni učinak dok je na S. aureus ATCC 2592 rezultat bio
indiferentan. Prilikom ispitivanja inhibicije nastanka biofilma u pojedinačnim ispitivanjima dokazana je statistički značajna
razlika u smanjenju produkcije biofilma u odnosu na pozitivnu kontrolu, ali nije zabilježena statistički značajna razlika u
povećanju koncentracija pojedinih aktivnih tvari. Kombinacija MUP + KAR/ET.U. pokazala je statistički značajnu razliku u
smanjenju produkcije biofilma u odnosu na identične koncentracije mupirocina. Najjači učinak imala je kombinacija
koncentracija od 0,0625 μg/mL MUP + 150 μg/mL KAR i identična kombinacija od 0,0625 μg/mL MUP + 150 μg/mL ET.U.
Ovime je dokazano da kombinacija manjih koncentracija mupirocina i eteričnog ulja origana, odnosno njegove glavne
sastavnice karvakrola nije djelovala na značajno smanjenje MIC-a mupirocina, ali može potentno djelovati na smanjenju
produkcije biofilma S. aureus.Due to the increasing development of antimicrobial resistance (AMR), various alternatives to antibiotics or potential
combinations with natural substances that could reduce antibiotic use are being intensively researched. Essential oils and their components have demonstrated potent active properties in numerous studies, exhibiting not only antimicrobial effects but also antioxidant and anti-inflammatory activities. Mupirocin is one of the most commonly used topical antibiotics, especially for skin infections caused by S. aureus. The development of specific antibiotic formulations together with essential oils or their components could achieve a synergistic effect, allowing the use of lower antibiotic concentrations and enhancing the antimicrobial action of such formulations. While antibiotics target specific bacterial processes, essential oils disrupt bacterial cell integrity and reduce biofilm production, thereby enhancing the overall antimicrobial effect. To initiate the development of such formulations, it is necessary to establish the efficacy of essential oil and its components, as well as their combinations with antibiotics. Therefore, the aim of this study was to determine the antimicrobial potential of oregano essential oil (O. compactum) and its main component, carvacrol, both individually and in combination with the antibiotic mupirocin. The study investigated the minimum inhibitory concentration (MIC) values for mupirocin (MUP), carvacrol (CAR), and oregano essential oil (EO) both individually and in combination. The MIC of mupirocin for S. aureus strains ranged from 0.076 to 0.141 μg/mL, whereas the MICs for carvacrol and oregano essential oil were several orders higher: carvacrol 525–700 μg/mL and oregano essential oil at 625–750 μg/mL. In tests of MUP and CAR/EO combinations on the S. aureus NCTC 12493 strain, an additive effect was observed, while the results for S. aureus ATCC 2592 were indifferent. When examining inhibition of biofilm formation in individual tests, a statistically significant reduction in biofilm production was noted compared to the positive control; however, increasing the concentrations of individual active substances did not produce a statistically
significant difference. The combination of MUP + CAR/EO showed a statistically significant reduction in biofilm production
compared to identical mupirocin concentrations. The strongest effect was seen with the combination of 0.0625 μg/mL MUP + 150 μg/mL CAR, and 0.0625 μg/mL MUP + 150 μg/mL EO. This demonstrated that while combining lower concentrations of mupirocin with oregano essential oil or its main component (carvacrol) did not significantly reduce mupirocin’s MIC, it could effectively reduce S. aureus biofilm production