Bunyamin Karagoz

Polymerization-induced self-assembly (PISA) - Control over the morphology of nanoparticles for drug delivery applications

In this paper, we describe the synthesis of asymmetric functional POEGMA-b-P(ST-co-VBA) copolymers in methanol, yielding in one-pot polymerization a range of nanoparticle morphologies, including spherical micelles, worm-like, rod-like micelles and vesicles. The presence of the aldehyde group was then exploited to form crosslinks or to conjugate chemotherapy compounds, such as doxorubicin, via pH-breakable bonds (Schiff base or imine) directly to the preformed nanoparticles. The influence of the nanoparticle morphologies on the MCF-7 breast cancer cell line uptake was investigated using flow cytometry and confocal microscopy. Finally, the IC50 of DOX, following nanoparticle delivery, was studied showing significant influence of the nanoparticle carrier morphology on therapeutic efficacy for breast cancer.

Pair correlation microscopy reveals the role of nanoparticle shape in intracellular transport and site of drug release

Nanoparticle size, surface charge and material composition are known to affect the uptake of nanoparticles by cells. However, whether nanoparticle shape affects transport across various barriers inside the cell remains unclear. Here we used pair correlation microscopy to show that polymeric nanoparticles with different shapes but identical surface chemistries moved across the various cellular barriers at different rates, ultimately defining the site of drug release. We measured how micelles, vesicles, rods and worms entered the cell and whether they escaped from the endosomal system and had access to the nucleus via the nuclear pore complex. Rods and worms, but not micelles and vesicles, entered the nucleus by passive diffusion. Improving nuclear access, for example with a nuclear localization signal, resulted in more doxorubicin release inside the nucleus and correlated with greater cytotoxicity. Our results therefore demonstrate that drug delivery across the major cellular barrier, the nuclear envelope, is important for doxorubicin efficiency and can be achieved with appropriately shaped nanoparticles.

Immobilization of catalase via adsorption on poly(styrene-d-glycidylmethacrylate) grafted and tetraethyldiethylenetriamine ligand attached microbeads

Fibrous poly(styrene-d-glycidylmethacrylate) (P(S-GMA)) brushes were grafted on poly(styrene-divinylbenzene) (P(S-DVB)) beads using surface initiated-atom transfer radical polymerization (SI-ATRP). Tetraethyldiethylenetriamine (TEDETA) ligand was incorporated on P(GMA) block. The multi-modal ligand attached beads were used for reversible immobilization of catalase. The influences of pH, ionic strength and initial catalase concentration on the immobilization capacities of the P(S-DVB)-g-P(S-GMA)-TEDETA beads have been investigated. Catalase adsorption capacity of P(S-DVB-g-P(S-GMA)-TEDETA beads was found to be 40.8 ± 1.7 mg/g beads at pH 6.5 (with an initial catalase concentration 1.0mg/mL). The K(m) value for immobilized catalase on the P(S-DVB-g-P(S-GMA)-TEDETA beads (0.43 ± 0.02 mM) was found about 1.7-fold higher than that of free enzyme (0.25 ± 0.03 mM). Optimum operational temperature and pH was increased upon immobilization. The same support was repeatedly used five times for immobilization of catalase after regeneration without significant loss in adsorption capacity or enzyme activity.

Amine functional monodisperse microbeads via precipitation polymerization of N-vinyl formamide: immobilized laccase for benzidine based dyes degradation.

Densely cross-linked poly(vinylamine) microbeads (∼ 2 μm) were prepared by precipitation copolymerization of N-vinyl formamide and ethylene glycoldimethacrylate in acetonitrile. The formamido groups of the microbeads were hydrolyzed into amino groups. Then, amino-functionalized microbeads were used for covalent immobilization of laccase via glutaraldehyde coupling. The average amount of immobilized enzyme was 18.7 mg/g microbeads. Kinetic parameters, V(max) and K(m) values were determined as 20.7 U/mg protein and 2.76 × 10(-2)mmol/L for free enzyme and 15.8 U/mg protein and 4.65 mmol/L for the immobilized laccase, respectively. The immobilized laccase was operated in a batch reactor for the degradation of two different benzidine based dyes (i.e., Direct Blue 1 and Direct Red 128). The laccase immobilized on the microbeads was very effective for removal of these dyes which interfere with the hormonal system.

Functionalization of Poly(divinylbenzene) Microspheres by Combination of Hydrobromination and Click Chemistry Processes: A Model Study

Fluorescent pyrene-functional poly(divinylbenzene) (PDVB) cross-linked core microspheres were synthesized using the click chemistry strategy as a model study. First, the core monodisperse microspheres were prepared by precipitation polymerization using 2,2′-azobisisobutronitrile (AIBN) as initiator in acetonitrile in the absence of any stabilizer. Residual double bonds were converted to azide-functions by hydrobrominated with hydrogen bromide followed by azidation with NaN3 in DMF solution. Propargyl pyrene was prepared independently as a fluorescent click component. Finally, azide-functionalized microspeheres were coupled with propargyl pyrene with high efficiency by click chemistry. The modified PDVB microspheres are characterized by using FT-IR spectroscopy, UV-Vis absorption spectroscopy, fluorescence spectroscopy and fluorescence microscopy.

Gadolinium-functionalized nanoparticles for application as magnetic resonance imaging contrast agents via polymerization-induced self-assembly

Polymerization-induced self-assembly (PISA) is an easily applied synthetic technique for the preparation of polymer nanoparticles with various shapes and at high concentrations. Although PISA has been applied to prepare nanoparticles for a variety of different applications, to date there have been no attempts to employ nanoparticles prepared via PISA as a novel platform from which to prepare positive contrast agents for magnetic resonance imaging (MRI). To this end, here we report an efficient synthesis of surface-functional polymer-based nanoparticles with tunable size and morphology (micelles, filomicelles and vesicles) via PISA, their post-synthetic functionalisation and an initial investigation into their use as a positive MRI contrast agent. A short functional block of poly(glycidyl methacrylate) was prepared using reversible addition-fragmentation chain transfer (RAFT) polymerization, and this was chain extended with oligo(ethylene glycol) methyl ether methacrylate to create a novel macromolecular chain transfer agent for PISA. A library of surface-functional self-assembled nanoparticles with different shapes was then synthesized, and the versatility of the glycidyl methacrylate units for post-synthetic surface functionalization was demonstrated by ring opening the epoxide with a primary thiol, a primary amine and sodium azide. The resulting azide functional materials could be further modified via copper-catalysed azide–alkyne cycloaddition. The epoxide groups were also exploited to conjugate Gd-DOTA to the polymeric nanoparticles and the application of the diverse polymeric nanoparticles for T1-weighted MRI was investigated, with the filomicelle emerging as a promising candidate due to both a good gadolinium-labelling efficiency and a high T1 relaxivity. Given that filomicelles typically exhibit enhanced blood circulation times, the gadolinium-labelled filomicelles could have potential applications as a blood pool agent for magnetic resonance angiography, and in cancer diagnostics/theranostics.

Synthesis of Methacrylate-Based Functional Monomers via Boron Ester Acidolysis and Their Polymerization

A new method is presented for the synthesis of functional methacrylate ester monomers. In this method, boron esters of appropriate alcohols were reacted with methacrylic acid to give the corresponding methacrylic ester monomers. The monomers N,N-dimethylaminoethyl methacrylate (DAMA), 2-chloroethyl methacrylate (CEMA), 2-bromoethyl methacrylate (BEMA), 2-methoxyethyl methacrylate (MEMA) and butyl methacrylate (BMA) were prepared in a laboratory scale by a two-step one-pot process. In the first step, the corresponding boron esters were formed by azeotropic removal of water. Subsequently, addition of methacrylic acid and heating at 140–160°C in the presence of pyridine resulted in acidolysis of the boron esters, yielding the functional methacrylate ester monomers in acceptable yields (41–71%). The monomers obtained were characterized by 1H-NMR and FT-IR spectra after isolation and purification. Their polymerizability was also demonstrated.

Modification of polydivinylbenzene microspheres by a hydrobromination/click-chemistry protocol and their protein-adsorption properties.

Hydrophobic- and/or hydrophilic-polymer-grafted PDVB microspheres are synthesized by the combination of hydrobromination and click-chemistry processes. The modified-PDVB microspheres and the intermediates at various stages of synthesis are characterized using GPC, ¹H NMR and FTIR spectroscopy and TGA analysis. Use of the microspheres as a support matrix for reversible protein immobilization via adsorption is investigated. The system parameters such as the adsorption conditions (i.e., enzyme concentration, medium pH) and desorption are studied and evaluated with regards to the biocatalytic activity and adsorption capacity.

Highly fluorescent sensing of nitroaromatic explosives in aqueous media using pyrene-linked PBEMA microspheres

Crosslinked 2-bromoethyl methacrylate polymer (PBEMA) was prepared in micro-spherical form (2-5µm) by precipitation polymerization methodology. The bromide substituent was substituted with an azide group, which was then coupled with 1-[(2-Propynyloxy)methyl]pyrene] via alkyne-azide click chemistry. The pyrene-linked microspheres showed an intense green-blue excimer emission with a maximum at 480nm, implying π-π stacking between the pyrene moieties on the microsphere surfaces. This fluorescence emission is extremely sensitive to the aromatic nitro compounds. So that the green-blue light fades immediately upon addition of trace amounts of 2,4,6-trinitrotoluene (TNT), 2,4-dinitrotoluene (DNT) and 2,4,6-trinitro phenol (TNP) in 100% aqueous media. Stern-Volmer plots were employed for comparison of their fluorescence quenching effects. The plots revealed Stern-Volmer constants of 1.33 × 105, 2.451 × 105 and 1.076 × 105M-1 for TNT, TNP and DNT, respectively. Furthermore, it has been observed that, the microspheres can be reused several times, without losing excimer emission properties.

New water-soluble polymer with allyl pendant groups

Polymerization of N-allyl maleamic acid (AMA) is described. The monomer, AMA, was prepared by reaction of maleic anhydride with allyl amine. It was demonstrated for the first time that only maleate double bonds are involved in the radical polymerization and the allyl groups remain unreacted. Water-soluble oligomers (1800–2100 Da) with allyl pendant groups were obtained in reasonably high yields (up to 78% within 45 h) by using 2,2′-azobis(2-methylpropionamidine) dihydrochloride as a radical source. Notably no cyclo-polymers are formed in this polymerization. Experiments also showed that polymerization of this process is rather slow and proceeds with first-order kinetics (k = 1.03 × 10−6 s−1) in ethanol, at 75°C. Interestingly, in the co-polymerization with some other vinyl monomers, such as acrylamide, acrylic acid and vinyl acetate, AMA behaves as a bi-functional monomer and gives cross-linked polymers.