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Application of electrospun membranes in "lung-on-a-chip" models for simulating the alveolar barrier
No full textRazvoj funkcionalnog "lung-on-a-chip" modela, koji na vjerodostojan način oponaša kompleksnu strukturu i funkciju alveolarno-kapilarne barijere, središnja je tema ovoga rada. Inspiriran fiziologijom ljudskih pluća, model je osmišljen kao multifunkcionalna platforma za istraživanje prijenosa tvari kroz plućno tkivo, ispitivanje djelovanja inhalacijskih pripravaka te evaluaciju toksičnosti aerosola i drugih inhalacijskih agensa. Ključni korak u njegovoj izradi bila je primjena metode elektroispredanja za dobivanje tankih, biokompatibilnih membrana od polikaprolaktona, pri čemu je posebna pažnja posvećena optimizaciji uvjeta izrade kako bi se što bolje reproducirale mehaničke i morfološke karakteristike plućnog tkiva. Osim toga, u sustav su integrirani hidrogelovi na bazi natrijeva alginata, koji omogućuju kontroliranu difuziju i upotpunjuju model pluća. Cilj istraživanja je izraditi dinamički model koji „diše“, odnosno simulira mehaničke sile disanja, čime se dobiva realistični prikaz respiratorne funkcije u in vitro uvjetima. Takav sustav otvara mogućnosti za unapređenje inhalacijskih terapija, provedbu toksikoloških ispitivanja te razvoj alternativnih metoda za analizu plućnih funkcija bez upotrebe životinjskih modela. Model predstavlja značajan napredak u području biomedicine i kemijskog inženjerstva, jer omogućuje ubrzanje procesa razvoja lijekova i terapija. Njegova primjena omogućuje bržu i etički prihvatljiviju evaluaciju novih terapija, osobito u područjima pulmologije i farmacije, čime postaje vrijedan alat doprinoseći razvoju sigurnijih i učinkovitijih terapijskih rješenja za respiratorne bolesti.The development of a functional lung-on-a-chip model that accurately mimics the complex structure and function of the alveolar-capillary barrier is the central focus of this work. Inspired by the physiology of the human lung, the model is designed as a multifunctional platform for studying substance transport across pulmonary tissue, evaluating the effects of inhaled formulations, and assessing the toxicity of aerosols and other inhalable agents. A key step in the fabrication of the model involved the application of electrospinning to produce thin, biocompatible membranes made of poly(ε-caprolactone), with particular attention given to optimizing fabrication conditions to best replicate the mechanical and morphological properties of lung tissue. Additionally, the system incorporates sodium alginate-based hydrogels, which enable controlled diffusion and complement the lung model. The aim of the research is to develop a dynamic, “breathing” model that simulates the mechanical forces of respiration, thereby providing a realistic representation of respiratory function under in vitro conditions. Such a system offers new opportunities for improving inhalation therapies, conducting toxicological testing, and developing alternative methods for analyzing lung function without the use of animal models. This model represents a significant advancement in the fields of biomedicine and chemical engineering, as it accelerates the process of drug and therapy development. Its application enables faster and more ethically acceptable evaluation of new therapies, particularly in the fields of pulmonology and pharmaceutical sciences, making it a valuable tool in the pursuit of safer and more effective therapeutic solutions for respiratory diseases
Monitoring the migration of additives from multiple recycled polyethylene microplastics
No full textPolimerni materijali nalaze se svuda oko nas i koriste se u raznim proizvodima prehrambene, kozmetičke, građevinske i drugih industrija, pri čemu je polietilen jedan od najčešće korištenih polimera. Dodavanjem aditiva poboljšavaju se određena svojstva polimera koja im omogućuju primjenu u različitim industrijama. S porastom globalne proizvodnje i uporabe polimernih materijala povećava se i količina plastičnog otpada. Mehaničko recikliranje često se smatra efikasnim rješenjem za njegovo smanjenje. Iako u literaturi postoji puno istraživanja koja se temelje na proučavanju polimera i njihovih svojstava, malo se njih koncentrira na to kako višestruko recikliranje utječe na otpuštanje aditiva iz plastike. Cilj ovoga rada bio je proučavanje utjecaja višestrukog recikliranja na migraciju aditiva iz polietilenske mikroplastike. U tu svrhu pripremljena su dva uzorka. Prvi uzorak se sastojao od polietilena (PE) i antioksidansa, dok se drugi uzorak sastojao od PE-a u koji su dodani antioksidansi i plastifikator. Nakon pripreme uzoraka u laboratorijskom ekstruderu provedena je simulacija pet ciklusa mehaničkog recikliranja. Nakon svakog ciklusa uzorci su usitnjeni do veličine mikroplastike. Kako bi se ustanovile promjene u strukturi, toplinskoj stabilnosti i stupnju kristalnosti uzoraka uzrokovane aditivima, provedena je karakterizacija nakon 1., 3. i 5. ciklusa recikliranja. Metode karakterizacije uključivale su infracrvenu spektroskopiju s Fourierovom transformacijom (FTIR), termogravimetrijsku analizu (TGA) i diferencijalnu pretražnu kalorimetriju (DSC). Isto tako, praćena je i migracija aditiva iz mikroplastike u vodeni medij, prilikom čega je određen sadržaj ukupnog organskog ugljika, omjer biorazgradivosti i toksičnost na organizmima Vibrio fischeri i Daphnia magna.Polymeric materials are all around us and are used in various products such as food, cosmetics, construction and other industries, whereby polyethylene is one of the most used polymers. The addition of additives is essential for enhancing certain properties of polymers, allowing their application in various industries. With increasing global production and usage of polymer materials, the amount of plastic waste is also increasing. Mechanical recycling is considered an effective way of solving this problem. Although there are many studies about polymers and their properties, not many of them are focused on the effect of multiple recycling cycles on the migration of additives from plastic. The aim of this thesis was to examine the effect of multiple recycling cycles on migration of additives from polyethylene microplastics. For this purpose, two samples were prepared. One of them was polyethylene (PE) with antioxidants, while the second one was PE with antioxidants and plasticizer. After preparation, a simulation of five recycling cycles was conducted in a laboratory extruder. After each cycle, the samples were ground to microplastic size. To establish changes in structure, thermal stability and degree of crystallinity of the samples, the characterization was done after 1st, 3rd and 5th cycle of recycling. These methods of characterization included Fourier-Transform Infrared Spectroscopy (FTIR), Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry (DSC). The migration of additives from microplastic to water was also monitored, and total organic carbon (TOC), ratio of biodegradability and toxicity on organisms Vibrio fischeri and Daphnia magna were determined
Irrigation of basil and beans with RO retentate
No full textOtpadne vode iz tekstilne industrije predstavljaju veliku opasnost za okoliš te su u pravilu zagađene širokim spektrom tvari čije uklanjanje nije jednostavno. Cilj ovog rada bio je simulirati postrojenje za obradu tekstilnih otpadnih voda (TOV) koristeći hibridni ultrafiltracijsko (UF) – reverzno osmotski (RO) proces, ali s naglaskom na zatvaranje vodenog kruga, odnosno da se sve struje iz membranskog procesa oporabe ili koriste za određene svrhe. Realni uzorci TOV-a prvo su podvrgnuti ultrafiltraciji te je skupljan permeat. Taj permeat je potom obrađivan reverznom osmozom te je retentat korišten za navodnjavanje biljaka bosiljka i graha. Rezultati pokazuju da je UF, kao prva membranska operacija, uspješno smanjila vrijednosti mutnoće, kemijske potrošnje kisika te spektralnog apsorpcijskog koeficijenta. S druge strane, RO je neophodna membranska operacija ukoliko se želi značajno smanjiti vrijednost električne vodljivosti te koncentracija raznih iona u obrađivanim vodama. Analiziranjem voda, biljnog materijala i tla, zaključuje se da su RO retentati te RO retentati razrijeđeni s vodovodnom vodom u omjeru 50:50 sigurni za razvoj biljaka u pogledu koncentracija klorida, nitrata, amonijevih kationa, sulfata, željeza i mangana te pH vrijednosti. U biljkama navodnjavanim TOV-om uočene su veće koncentracije dušika, fosfora i kalija u odnosu na iste biljke navodnjavane vodovodnom vodom, no primjenom razrijeđenih RO retentata u biljkama je utvrđena slična koncentracija navedenih tvari kao u biljkama navodnjavanih vodovodnom vodom. U tlu navodnjavanom TOV-om uočene su značajno veće koncentracije sumpora i fosfora u odnosu na tlo navodnjavano vodovodnom vodom. Primjenom razrijeđenih i nerazrijeđenih RO retentata vrijednosti koncentracija navedenih elemenata niže su nego kod uzorka tla navodnjavanog TOV-om.Textile wastewater (TWW) poses a major threat to the environment and is usually contaminated with a variety of substances that are difficult to remove. The aim of this work was to simulate a textile wastewater treatment plant using a hybrid ultrafiltration (UF) – reverse osmosis (RO) process, with the primary objective of closing the water cycle so that all process streams from the membrane process can be reused or utilised for other purposes. Real samples of TWW were first treated with the UF process and the UF permeate was collected. This permeate was then treated with the RO and the RO retentate was collected as it was used as irrigation water for basil and beans. The results show that UF was the first membrane operation to successfully reduce the values for turbidity, chemical oxygen demand, and spectral apsorption coefficient. On the other hand, RO is the necessary membrane operation when it comes to significantly reducing the values of electrical conductivity and the concentration of various ions in the treated water. Analysis of the water, plant material and soil showed that all RO retentates and all RO retentates diluted 50:50 with tap water were harmless to plant development in terms of chloride, nitrate, ammonia, sulphate, iron, and manganese concentrations and pH. Plants irrigated with TWW were found to have higher concentrations of nitrogen, phosphorus and potassium than plants irrigated with tap water, but when diluted RO retentates were used, these plants had similar concentrations of these substances to the plants irrigated with tap water. In the soil irrigated with TWW, sulphur and phosphorus concentrations were found to be much higher than in the soil irrigated with tap water. When using diluted and undiluted RO retentates, the concentration values of the mentioned elements were lower compared to the soil samples irrigated with TWW
Screening of N-acetylneuraminic acid synthase analogues on different sugars
No full textEnzimi su biokatalizatori koji pokazuju brojne prednosti nad kemijskim katalizatorima. Unatoč tome, enzimi nailaze na ograničenja u industrijskoj primjeni zbog čega se nastoje modificirati i poboljšati tehnikama enzimskog inženjerstva. Jedna od ključnih reakcija za koje se nastoje modificirati enzimi jest sinteza ugljik-ugljik veze. Aldolna kondenzacija najčešća je metoda za sintezu navedene veze, a tu reakciju kataliziraju enzimi zvani aldolaze. Sintaza
N-acetilneuraminske kiseline (NeuS) je aldolaza ovisna o fosfoenolpiruvatu koja katalizira sintezu N-acetilneuraminske kiseline (Neu5Ac). Neu5Ac pripada obitelji sijalinskih kiselina koje imaju raznovrsne funkcije u fiziološkim procesima. U ovom radu ispitivala se aktivnost analoga sintaza N-acetilneuraminske kiseline na šećere L-arabinozu, D-galaktozu, D-laktozu, D-liksozu, D-(+)-manozu i D-(+)-ksilozu kao supstrate. Kao analozi sintaza
N-acetilneuraminske kiseline korišteni su umjetno dobiveni PNH3, PNH6, PNH11 i PNH17 enzimi te NmNeuS enzim iz bakterije Neisseria meningitidis (Nm). Aktivnost tih enzima ispitivana je pomoću početnih reakcijskih brzina praćenjem koncentracija reaktanata i produkata reakcije u vremenu pomoću tekućinske kromatografije visoke djelotvornosti (HPLC), a iz prikupljenih eksperimentalnih podataka izračunate su konverzije supstrata i iskorištenja reakcije. Na temelju prikupljenih rezultata, zaključeno je da PNH3 i PNH6 enzimi imaju veću aktivnost prema heksozama (D-galaktoza, D-(+)-manoza) kao supstratima nego pentozama (L-arabinoza, D-liksoza). U slučaju staničnog ekstrakta NmNeuS je obrnuto. NmNeuS je pokazao najveće vrijednosti konverzije i iskorištenja za sve supstrate. Usporedbom iskorištenja reakcija kataliziranim sa staničnim ekstraktom NmNeuS enzima i s njegovim pročišćenim oblikom, zaključeno je da je pročišćeni enzim specifičniji prema supstratima nego stanični ekstrakt NmNeuS enzima.Enzymes are biocatalysts that exhibit numerous advantages over chemical catalysts. Despite this, enzymes face limitations in industrial applications, which is why efforts are made to modify and improve them using enzyme engineering techniques. One of the key reactions for which enzymes are being modified is the synthesis of carbon-carbon bonds. Aldol condensation is the most common method for the synthesis of these bonds, and this reaction is catalyzed by enzymes called aldolases. N-acetylneuraminic acid synthase (NeuS) is a phosphoenolpyruvate-dependent aldolase that catalyzes the synthesis of N-acetylneuraminic acid (Neu5Ac). Neu5Ac belongs to the family of sialic acids, which have various functions in physiological processes. In this study, the activity of analogs of N-acetylneuraminic acid synthase was tested on sugars L-arabinose, D-galactose, D-lactose, D-lyxose,
D-(+)-mannose, and D-(+)-xylose as substrates. The analogs of N-acetylneuraminic acid synthase used were the artificially obtained PNH3, PNH6, PNH11 and PNH17 enzymes, as well as the NmNeuS enzyme from the bacterium Neisseria meningitidis (Nm). The activity of these enzymes was tested using initial reaction rates by monitoring the concentrations of reactants and reaction products over time using high-performance liquid chromatography (HPLC), and the substrate conversions and reaction yields were calculated from the collected experimental data. Based on the results obtained, it was concluded that the PNH3 and PNH6 enzymes have greater activity towards hexoses (D-galactose, D-(+)-mannose) as substrates than towards pentoses (L-arabinose, D-lyxose). In the case of the NmNeuS cell extract, the reverse was true. NmNeuS showed the highest values of conversion and yield for all substrates. By comparing the yields of reactions catalyzed by the NmNeuS cell extract enzyme with its purified form, it was concluded that the purified enzyme is more specific towards substrates than the NmNeuS cell extract enzyme
Preparation and characterization of polysaccharide films via layer-by-layer method
No full textTanki filmovi istražuju se kao sustavi za dostavu lijekova, poput kemoterapeutika. Njihova primjena u obliku premaza za implantate ili samostalnih sustava za dostavu lijekova omogućuje kontrolirano i ciljano otpuštanje kemoterapeutika izravno na mjestu tumora, čime se potencijalno smanjuje toksičnost i poboljšava terapijska učinkovitost. Fitinska kiselina, prirodni antioksidans, prepoznata je po svojoj ulozi u prevenciji i liječenju različitih patoloških stanja i karcinoma. Međutim, potrebno je pripremiti pogodne sustave za njezinu dostavu. Zbog velikog broja hidroksilnih skupina fitiska kiselina posjeduje mogućnost stvaranja stabilnih kompleksa s prirodnim polimerima koji mogu poslužiti kao sustavi za dostavu fitinske kiseline. Biorazgradljivost, biokompatibilnost, antimikrobna svojstva i stabilnost polielektrolitnih kompleksa kitozana i alginata (CA) čine ih pogodnima za biomedicinske primjene. Fizikalnim umreživanjem CA polielektrolitnih kompleksa fitinskom kiselinom moguće je postići antitumorska svojstva tih materijala. Stoga je cilj ovoga rada razviti stabilne tanke filmove na osnovi polielektrolitnih kompleksa kitozan-alginat umreženih fitinskom kiselinom primjenom metode sloj-po-sloj. U ovom radu, ispitan je utjecaj koncentracije otopine fitinske kiseline (0,1, 0,5 i 1,0 mas.%) i vremena umreživanja (3,5 i 7 min) na režim rasta i svojstva dobivenih tankih filmova. Tanki filmovi karakterizirani su ATR-FTIR spektroskopijom kako bi se potvrdilo umreživanje fitinskom kiselinom i ispitala učinkovitost procesa ispiranja tankih filmova nakon pripreme. Morfologija površine i presjeka nakon loma ispitane su SEM analizom. Kapaciteti apsorpcije tankih filmova ispitani su u otopinama fosfatnog pufera (pH 6 i 8) te u fosfatom puferiranoj otopini soli (pH 7,4) tijekom 24 sata. ATR-FTIR spektroskopija potvrdila je umreživanje polielektrolitnog kompleksa CA fitinskom kiselinom, a izostanak apsorpcijskih vrpci otapala ukazao je na dostatnost procesa ispiranja. Iako su svi slojevi pokazali strukturnu homogenost, razlike u koncentraciji fitinske kiseline i vremenu umreživanja dovele su do razlike u mehanizmima rasta i posljedično u morfologiji presjeka tankih filmova. Ispitivanjem fizikalnih svojstava pri fiziološkim uvjetima potvrđena je stabilnost tankih filmova koji su maksimalni kapacitet apsorpcije postigli unutar tri sata.Thin films are being investigated as drug delivery systems, particularly for chemotherapeutic agents. Their application as implant coatings or standalone drug delivery systems enables controlled and targeted release of chemotherapeutics directly at the tumor site, thereby potentially reducing systemic toxicity and enhancing therapeutic efficacy. Phytic acid, a natural antioxidant, has been recognized for its role in the prevention and treatment of various pathological conditions and cancers. However, the development of suitable delivery systems is required to realize its full therapeutic potential. Due to its high density of hydroxyl groups, phytic acid has the capacity to form stable complexes with natural polymers, which can serve as carriers for its delivery. The biodegradability, biocompatibility, antimicrobial properties, and stability of chitosan–alginate (CA) polyelectrolyte complexes make them suitable for biomedical applications. Physical cross-linking of CA polyelectrolyte complexes with phytic acid may result in antitumor properties of the resulting materials. The aim of this study was to develop stable thin films based on chitosan–alginate polyelectrolyte complexes cross-linked with phytic acid using layer-by-layer assembly (LbL). The effect of phytic acid solution concentration (0.1, 0.5, and 1.0 wt%) and cross-linking time (3,5 and7 minutes) on the growth regime and properties of the resulting thin films was investigated. Thin films were characterized by ATR-FTIR spectroscopy to confirm cross-linking with phytic acid and to assess the effectiveness of the rinsing process after film preparation. Surface and cross-sectional morphology following fracture were examined using SEM analysis. The absorption capacity of the thin films was evaluated in phosphate buffer solutions (pH 6 and 8) and in phosphate-buffered saline (PBS, pH 7.4) over a period of 24 hours. ATR-FTIR spectroscopy confirmed successful cross-linking of the CA polyelectrolyte complex with phytic acid, and the absence of solvent absorption bands indicated the adequacy of the rinsing process. While all films exhibited structural homogeneity, variations in phytic acid concentration and cross-linking time resulted in differences in growth mechanisms, thereby affecting the morphology of the film cross-sections. Evaluation of film physical properties under physiological conditions confirmed their stability, with maximum apsorption capacity reached within three hours
Design of metal-free catalysts for CO2 reduction
No full textPrateći sve intenzivnije napore znanstvene zajednice u razvoju nemetalnih katalizatora za redukciju ugljikova dioksida (CO2), potaknute rastućim izazovima klimatskih promjena, ovaj rad istražuje potencijal fosfina kao donora hidrida i istodobno kao katalizatora za reakciju redukcije CO2 u mravlju kiselinu. Koristeći računalne kvantno-kemijske metode (DFT), ispitana je sposobnost doniranja hidrida kod protoniranih difosfina supstituiranih elektron-donorskim skupinama: tris(dimetilamino)fosfazenilnom, tris(pirolidino)fosfazenilnom i tetrametilgvanidinskom. Ključan aspekt dizajna ovih difosfina je mogućnost formiranja dativne P–P veze nakon doniranja hidridnog iona, koja značajno pridonosi hidridnom karakteru i katalitičkom potencijalu spojeva. Ispitana je hidričnost ukupno 18 difosfina, a za dva od njih (spojevi 1aa i 4aa) izračunati su reakcijski putevi konverzije CO2 u mravlju kiselinu. Pri tome je utvrđeno da spoj 4aa posjeduje dobar potencijal za katalitičko djelovanje jer je cijeli reakcijski ciklus, uključujući i regeneraciju katalizatora, termodinamički i kinetički ostvariv. Ovim istraživanjem otvara se novi smjer u razvoju nemetalnih katalizatora temeljenih na fosfinima, koji su dosad bili slabo istraženi u kontekstu redukcije CO2.In response to the growing need for metal-free catalyst design for the reduction of carbon dioxide (CO₂), driven by the escalating impact of climate change, this study explores the potential of protonated phosphine compounds to act as both hydride donors and catalysts for reduction of CO2 to formic acid. Using computational quantum-chemical methods (DFT), the hydride-donating ability of protonated diphosphines substituted with electron-donating groups — tris(dimethylamino)phosphazenyl, tris(pyrrolidino)phosphazenyl, and tetramethylguanidino — was investigated. The design of diphosphines capable of forming a dative P–P bond upon hydride donation has proven crucial, as this interaction significantly enhances both the hydride-donating ability and the catalytic potential of these compounds. The hydricity of a total of 18 diphosphines was investigated, and for two of them (compounds 1aa and 4aa), reaction pathways for the conversion of CO2 to formic acid were calculated. It was found that compound 4aa has good potential for catalytic activity, as the entire reaction cycle, including catalyst regeneration, is both thermodynamically and kinetically feasible. This study opens a new direction in the development of non-metallic catalysts based on phosphines, which have so far been scarcely explored in the context of CO2 reduction
Synthesis and spectroscopic characterization of novel iminocoumarine benzoxazole derivatives as potential sensors for pH in solutions
No full textU ovom radu provedena je sinteza novih iminokumarinskih derivata benzoksazola 9-12. Polazni spoj, etil-2-cijano-acetamidat-hidroklorid 3, sintetiziran je aktivacijom malonitrila 1 u prisutnost trimetilsilil-klorida (TMSCl) i etanola. Ključni međuprodukt,
2-cijanometilbenzoksazol 4, dobiven je reakcijom spoja 2 i 2-aminofenola 3, primjenom dviju sintetskih metoda: konvencionalne te mikrovalovima potpomognute sinteze. Ciljani iminokumarinski derivati 9-12 dobiveni su kondenzacijom 2-cijanometilbenzoksazola 4 s odgovarajućim supstituiranim 2-hidroksibenzaldehidima 5-8 u prisutnosti piperidina i etanola u reaktoru za paralelnu sintezu. Strukture spojeva 9-12 potvrđene su 1H i 13C NMR spektroskopijom. Radi ispitivanja njihove potencijalne primjene kao optičkih pH senzora u otopinama, spojevi 9, 10 i 12 spektroskopski su okarakterizirani UV/Vis spektroskopijom u šest organskih otapala različitih polarnosti. Provedene su i UV/Vis pH titracije u puferskim otopinama različitih pH vrijednosti. Promjene u apsorpcijskim spektrima, ovisno o polarnosti otapala i pH vrijednosti medija, ukazuju na potencijalnu primjenjivost spojeva 9, 10 i 12 kao optičkih pH senzora.In this work, the synthesis of new iminocoumarine derivatives of benzoxazoles 9-12 was carried out. The starting compound, ethyl-2-cyano-acetamidate hydrochloride 60, was synthesized by activation of malonitrile 1 in the presence of trimethylsilyl chloride (TMSCl) and ethanol. The key intermediate, 2-cyanomethylbenzoxazole 4, was obtained by reaction of compound 2 with 2-aminophenol 3, using two synthetic methods: conventional and microwave-assisted synthesis. The targeted iminocoumarine derivatives 66, 67 and 69 were obtained by condensation of 2-cyanomethylbenzoxazole 4 with the corresponding substituted 2- hydroxybenzaldehydes 5-8 in the presence of piperidine and ethanol in a parallel synthesis reactor. The structures of compounds 9-12 were confirmed by 1H and 13C NMR spectroscopy. To investigate their potential application as optical pH sensors in solutions, compounds 9, 10 and 12 were spectroscopically characterizedby UV/Vis spectroscopy in six organic solvents of different polarities, as well as by UV/Vis pH titrations in a buffered solutions of different pH values. Changes in absorption spectra, depending on the polarity of the solvent and the pH value of the medium, indicate the potential applicability of compounds 9, 10 and 12 as optical pH sensors
Stability of nitrilase during synthesis of (R)-4-chloro-3-hydroxybutyric acid
No full textEnzimi, odnosno biokatalizatori, imaju brojne prednosti nad kemijskim katalizatorima te se zbog toga njihova upotreba i potražnja u farmaceutskoj industriji neprestano povećava. Enzimi koji kataliziraju reakcije hidrolize nitrila u karboksilne kiseline i amonijak bez stvaranja amidnih međuprodukata nazivaju se nitrilazama. Nitrilaza se koristi kao biokatalizator u reakcijama pretvorbe (R)-4-klor-3-hidroksibutironitrila u (R)-4-klor-hidroksibutansku kiselinu. Zatim dolazi do spontane ciklizacije (R)-4-klor-3-hidroksibutanske kiseline u
(R)-3-hidroksi-γ-butirolakton koji je svestrani kiralni gradivni blok, a često se koristi u sintezama mnogih farmaceutskih polimera i otapala. U ovome se radu pratila stabilnost nitrilaze pri sintezi (R)-4-klor-hidroksibutanske kiseline. Sinteza (R)-4-klor-hidroksibutanske kiseline provedena je u kotlastom reaktoru. Spektrofotometrijskim mjerenjima određivana je aktivnost nitrilaze pri različitim koncentracijama supstrata, (R)-4-klor-3-hidroksibutironitrila, kroz vremensko razdoblje od 24 h te su uspoređeni eksperimentalno dobiveni podaci s onima predviđenim matematičkim modelom. Uočeno je da aktivnost nitrilaze pada s vremenom, odnosno dolazi do deaktivacije nitrilaze. Iz dobivenih eksperimentalnih podataka procijenjena je konstanta deaktivacije pri različitim koncentracijama supstrata i enzima te je zaključeno kako početni uvjeti u reaktoru utječu na stabilnost nitrilaze.Enzymes, also known as biocatalysts, have numerous advantages over chemical catalysts and because of that their use and demand in pharmaceutical industry is constantly growing. The enzymes that catalyze reactions of nitrile hydrolisis into carboxylic acids and ammonia without the formation of amide intermediates are called nitrilases. Nitrilases are used in conversion reactions (R)-4-chlorine-3-hydroxybutyronitrile into
(R)-4-chlorine-3-hydroxybutyric acid. Afretwords, a spontaneous reaction of cyclisation (R)-4-chlorine-3-hydroxybyric acid into (R)-3-hydroxy-γ-butyrolactone occurs, a versatile chiral building block, often used in synthesis of numerous pharmaceutical polymers and solutions. In this paper, the stability of nitrilase during the synthesis of
(R)-4-chlorine-3-hydroxybutyric acid was monitored. The synthesis of
(R)-4-chlorine-3-hydroxybutyric acid takes place in a batch reactor. The activity of nitrilase at different contrentrations of the substrate, (R)-4-chlorine-3-hydroxybutyronitrile, over a period of 24 hours was measured with the spectrophotometric assay. The experimentally obtained data was compared with those predicted by the mathematical model. It was observed that the activity of nitrilase decreases with time and it was concluded that deactivation of nitrilase occurs. The deactivation constant at different substrate and enzyme concentration was estimated from the obtained experimental data and it was concluded that the initial conditions in the reactor affect the stability of nitrilase
Improving bioremediation of waste streams through exogenous microbial cultures
No full textU modernom svijetu tehnološkog napretka, antropogene aktivnosti dovode do sve većeg onečišćenja voda. Procjedne vode koje nastaju iz organske frakcije miješanog komunalnog otpada sadrže visoko opterećenje te zahtijevaju fizikalnu, kemijsku ili biološku obradu prije ispuštanja u okoliš. Tehnologija bioremedijacije postaje sve važnija u suočavanju s navedenim izazovima jer pruža održivo i visokoučinkovito rješenje za sanaciju različitih vrsta onečišćenja. Onečišćujuće tvari putem procesa biorazgradnje ili biotransformacije prevode se u netoksične produkte i nusprodukte djelovanjem enzimskog kataboličkog potencijala mikroorganizama. Kako bi se povećala brzina i djelotvornost tehnologije bioremedijacije uvode se dodatni alati kao što su optimiranje procesnih čimbenika putem biostimulacije te korištenje autohtonih ili egzogenih kultura kao inokuluma u procesu bioaugmentacije.
U ovom radu istraživana je bioremedijacija procjedne vode iz organske frakcije iz miješanog komunalnog otpada. Uspoređena je djelotvornost uklanjanja organskog onečišćenja uz bioaugmentaciju egzogenih mikrobnih kultura Pseudomonas putida te aktivnog mulja.
Pokusi su provedeni u šaržnom reaktoru pri submerznim uvjetima u periodu od 10 dana. Ukupna prosječna vrijednost učinkovitosti procesa uklanjanja organske tvari za istraživane sustave iznosila je 89 %. Provedena istraživanja pružaju uvide u praktičnu primjenu bioremedijacije i doprinose daljnjem razvoju strategija za obradu otpadnih voda.In the modern society of technological progress, anthropogenic activities are leading to increasing water pollution. Leachates originating from the organic fraction of mixed municipal waste are highly contaminated and require physical, chemical or biological treatment before being discharged into the environment. Bioremediation technology is becoming increasingly important in overcoming these challenges as it offers a sustainable and highly effective solution for the remediation of various types of pollution. Pollutants, through processes of biodegradation or biotransformation, are converted into non-toxic products and byproducts through the enzymatic catabolic potential of microorganisms. To enhance the speed and effectiveness of bioremediation technology, additional tools are introduced, such as the optimisation of process factors through biostimulation and the use of indigenous or exogenous cultures as inoculants in bioaugmentation. In this work, the bioremediation of leachate from the organic fraction of mixed municipal waste was investigated.
The effectiveness of the removal of organic pollution removal with bioaugmentation with exogenous microbial cultures Pseudomonas putida and activated sludge was compared.
The experiments were carried out in a batch reactor under submerged conditions for 10 days. The average overall efficiency of the organic matter removal in the investigated systems was 89 %. The conducted research provides insights into the practical application of bioremediation and contributes to the further development of wastewater treatment strategies
Novel imidazo[4,5-b]pyridine conjugates: synthesis, structural characterization and biological activity
No full textU okviru doktorskog rada opisana je sinteza, strukturna karakterizacija i biološka aktivnost novih derivata imidazo[4,5-b]piridina. Za sintezu novih spojeva korištene su klasične metode organske sinteze, sinteza potpomognuta mikrovalovima, paralelna sinteza i organometalna sinteza. Akrilonitrilni derivati 21–40 i 58–89 priređeni su aldolnom kondenzacijom 2-cijanometilimidazo[4,5-b]piridina s različitim benzaldehidima. Ispitan je utjecaj supstitucije u položaju N3 imidazo[4,5-b]piridinske jezgre, broj i položaj metoksi i hidroksi skupina, te prisutnost N,N-dialkilnog supstituenta u para-položaju fenilne jezgre na biološku aktivnost i spektroskopske karakteristike. Nadalje, reakcijom ciklokondenzacije
2-cijanometilimidazo[4,5-b]piridina i salicilbenzaldehida priređeni su iminokumarinski derivati 97–119 supstituirani u položajima 6 i 7 iminokumarinske jezgre, te na N3 dušikovom atomu imidazo[4,5-b]piridinske jezgre. 2-fenilamidino-supstituirani derivati imidazo[4,5-b]piridina 126–133 priređeni su iz cijano-supstituiranih prekursora 122–125, dok su reakcijom metiliranja priređeni derivati 134–136. Pinnerovom reakcijom u dva stupnja, priređeni su svi derivati supstituirani cikličkim amidinima, a one-pot reakcijom s reagensom LiHMDS derivati supstituirani nesupstituiranim amidinom. Suzukijevom reakcijom priređeni su derivati 142–146 i 149–154 supstituirani u položaju 6 imidazo[4,5-b]piridinske jezgre, a ovisno o korištenoj bornoj kiselini, u para-položaju fenilnog prstena supstituirani su različitim skupinama. N3-metil-supstituirani amidini imidazo[4,5-b]piridina 163–171 priređeni su iz cijano-supstituiranih prekursora 152, 161 i 162 Pinnerovom reakcijom. Serija konjugata imidazo[4,5-b]piridina i amidino-supstituiranih benzazola 182–190 priređena je kondenzacijom amidino-supstituiranih prekursora 173–181, odnosno
4-amidino-1,2-fenilendiamin-hidroklorida ili 5-amidino-2-aminobenzentiola s
imidazo[4,5-b]piridin-6-karbaldehidom 172. Schiffove baze imidazo[4,5-b]piridina 194–197 priređene su iz imidazo[4,5-b]piridina-karbaldehida 172 s anilinima, a Schiffove baze 202–214 iz 2-aminofenil-supstituiranih imidazo[4,5-b]piridina s benzaldehidima. Priređene su i dvije klase amidnih derivata imidazo[4,5-b]piridina koje se međusobno razlikuju prema položaju amidne veze. Derivati 217, 218, 230, 231 i 234 priređeni su iz benzoilnog klorida i amino-supstituiranih derivata, dok je derivat 222 priređen katalitički. Amidino-supstituirani derivati 219, 220, 232 i 233 priređeni su Pinnerovom reakcijom u dva stupnja.
Amino-supstituirani imidazo[4,5-b]piridini 102, 111, 147, 155, 233 i 235 priređeni su redukcijom iz odgovarajućih nitro-supstituiranih derivata. Radi bolje topljivosti derivati 103, 112, 119, 148, 156, 224 i 236 su pripravljeni kao amonijeve hidrokloridne soli. Strukture svih priređenih ciljanih spojeva i prekursora potvrđene su 1H i 13C NMR spektroskopijom, a za pojedine spojeve i dodatno masenom spektrometrijom. Detaljna spektroskopska karakterizacija u svrhu pronalaženja novih potencijalnih optičkih pH-senzora, provedena je za odabrane spojeve iz klasa akrilonitrila, iminokumarina i Schiffovih baza, a pKa vrijednosti određene su eksperimentalno i računalnim metodama. Svim ciljanim novopriređenim spojevima ispitana je antiproliferativna aktivnost in vitro na nekoliko staničnih linija humanih karcinoma. Za najaktivnije derivate istražen je mehanizam biološkog djelovanja koji je dodatno potvrđen, za neke derivate, i računalnom analizom. Antioksidativna aktivnost in vitro ispitana je spektroskopskim metodama DPPH, FRAP i ABTS, te elektrokemijski, dok je mehanizam antioksidativnog djelovanja potvrđen i računalnom analizom. Najaktivnijim amidinskim derivatima ispitana je interakcija s ct-DNK. Odabranim derivatima ispitana je antibakterijska aktivnost in vitro na nekoliko sojeva Gram-pozitivnih i Gram-negativnih bakterija, te antivirusna aktivnost in vitro prema odabranim vrstama virusa.Within this work, the synthesis, structural characterization and biological activity of novel imidazo[4,5-b]pyridine derivatives was described. For the synthesis of targeted compounds, classical methods of organic chemistry, parallel synthesis, and microwave assisted synthesis as well as organometal chemistry were used. Acrylonitrile derivatives 21–40 and 58–89 were prepared in the aldol condensation from 2-cyanomethylimidazo[4,5-b]pyridines and various benzaldehydes. Influence of N3 substitution on imidazo[4,5-b]pyridine core, the number and the position of hydroxy and methoxy groups as well as presence of N,N-dialkyl substituent placed at para-position of the phenyl ring on biological activity and spectroscopic characteristics was investigated. In the cyclocondensation reaction from 2-cyanomethylimidazo[4,5-b]pyridines and salicyl benzaldehydes, iminocoumarin derivatives 97–119 substituted at positions 6 and 7 of iminocoumarin core as well as at the position N3 of imidazo[4,5-b]pyridine nucleus were prepared. 2-phenylamidino-substituted
imidazo[4,5-b]pyridine derivatives 126–133 were prepared from cyano-substituted precursors 122–125, while derivatives 134–136 were prepared in the reaction of methylation. Two step Pinner reaction was used to prepare compounds substituted with cyclic amidines, while one pot reaction with LiHMDS was used to prepare derivatives with unsubstituted amidines. Suzuki coupling was used to prepare derivatives 142–146 and 149–154 substituted at the postition 6 of imidazo[4,5-b]pyridine core, depending on used boronic acid, substituted at the p–position of the phenyl ring with different substituents.
N3-methyl-substituted amidine derivatives 163–171 were prepared from cyano-substituted precursors 152, 161 and 162 in Pinner reaction and one pot reaction with LiHMDS. Benzazole conjugates of imidazo[4,5-b]pyridine with amidino-substituted benzazoles 182–190 were synthesized by condensation amidino-substituted precursors 173–181, respectively from 4-amidino-1,2-phenylenediamines hydrochlorides or 5-amidino-2-amino-substituted benzenthiolates with imidazo[4,5-b]pyridine-6-carbaldehyde 172. Imidazo[4,5-b]pyridine derived Schiff bases 194–197 were prepared from imidazo[4,5-b]pyridine-6-carbaldehyde 172 with various anilines, while Schiff bases 202–214 obtained from
2-aminophenyl-substituted imidazo[4,5-b]pyridines with benzaldehydes. Two series of amide derivatives were prepared which differ from position of amide bond. Derivatives 217, 218, 230, 231 and 234 were prepared by reactions between benzoil chlorides and
amino-substituted derivatives, while derivative 222 was prepared catalytically.
Amidino-substituted derivatives 219, 220, 232 and 233 were prepared by Pinner reaction.
All amino-substituted imidazo[4,5-b]pyridines 102, 111, 147, 155, 233 i 235 were prepared by reduction of corresponding nitro-substituted compounds. Some derivatives 103, 112, 119, 148, 156, 224 and 236 were prepared as ammonium hydrochloride salts in order to achieve better solubility. Structures of targeted compounds and precursors were analysed by means of 1H and 13C NMR spectroscopy, and were further confirmed by mass spectrometry for selected compounds. Thorough spectroscopic characterization was conducted for selected acrylonitrile, iminocoumarin and Schiff base derivatives, while pKa values were determined experimentally and computationally. All targeted compounds were evaluated for their antiproliferative activity in vitro on several human cancer cell lines. Mechanism of action was further investigated for the most active compounds and for some chosen derivatives confirmed also by computational analysis. Antioxidative activity in vitro was determined by using spectroscopic methods DPPH, FRAP and ABTS as well as electrochemically, while their mechanism of antioxidative action was explained by computational analysis. The most active amidine compounds were chosen for studying their interaction with ct-DNA. Selected compounds were tested also for their antibacterial activity in vitro against some Gram positive and Gram negative bacterial strains, as well as antiviral activity in vitro against selected viruses