Synthesis and Characterization of ROSA Dye - A Rhodamine B-type Fluorophore, Suitable for Bioconjugation and Fluorescence Studies in Live Cells

Background: Herein we report the multigram-scale synthesis, characterization and application of a rhodamine B-based fluorophore (ROSA) suitable for fluorescent studies in biological applications. This fluorophore is devoid of rhodamine spirolactone formation and furthermore characterized by a high molar extinction coefficient (ϵ=87250 ± 1630 M-1cm-1) and quantum yield (φ) of 0.589 ± 0.070 in water. Reported here is also the application of ROSA towards synthesis of a ROSA-PEG-GRGDS-NH2 fluorescent probe suitable for live cell imaging of αvβ3 integrins for in vitro assays. Objective: The main objective of this study is to efficiently prepare rhodamine B derivative, devoid of spirolactone formation that would be suitable for bioconjugation and subsequent bioimaging. Methods: Rhodamine B was transformed into rhodamine B succinimide ester (RhoB-OSu) using N-hydroxysuccinimide. RhoB-OSu was further coupled to sarcosine to obtain rhodamine Bsarcosine dye (ROSA) in good yield. The ROSA dye was then coupled to a αvβ3 integrin binding sequence using standard solid-phase conditions. Resulting ROSA-PEG-GRGDS-NH2 probe was used to image integrins on cancer cells. Results: The rhodamine B-sarcosine dye (ROSA) was obtained in multigram scale in good total yield of 47%. Unlike rhodamine B, the ROSA dye does not undergo pH-dependent spirolactone/spirolactam formation as compared with rhodamine B-glycine. It is also characterized by excellent quantum yield (φ) of 0.589 ± 0.070 in water and high molar extinction coefficient of 87250 ± 1630 M-1cm-1. ROSA coupling to the RGD-like peptide was proved to be efficient and straightforward. Imaging using standard filters on multimode plate reader and confocal microscope was performed. The αvβ3 integrins present on the surface of live WM-266-4 (melanoma) and MCF- 7 (breast cancer) cells were successfully imaged. Conclusion: We successfully derivatized rhodamine B to create an inexpensive, stable and convenient to use fluorescent probe. The obtained derivative has excellent photochemical properties and it is suitable for bioconjugation and many imaging applications.

OBTAINING SOME THERMOD AINING SOME THERMODYNAMIC PARAMETERS CONTAINING ME AINING METACRYLATE AND ALL TE AND ALLYL GROUPS CON YL GROUPS CONTAINING IN COMPOSITION

The article discusses the preparation of some thermodynamic parameters characterizing the final product of complex radical polymerization of amino alkyl acrylate derivatives containing both methacrylate and alkyl groups of low molecular weight compounds with electron acceptors - polymerization initiators at temperatures that exclude thermal decomposition of the initiators themselves. The equilibrium constant of the complication reaction and the molar extinction coefficient of the complex were determined by graphically solving the Bеneshe-Hildebrand equation, the heats of complex formation, entropy, Gibbs energy were found, and the elemental composition of low molecular weight and high molecular weight substances was determined. As can be seen from the studies, the molar extinction coefficient of the complex practically does not depend on temperature, and the equilibrium constant of the complexation reaction even in a narrow temperature range (283–293 K) increases with increasing temperature. The structure and composition of the synthesized monomeric and polymer salt were also considered in the work, and confirmed by IR-spectroscopy and analysis of elemental composition, the obtained spectroscopic data showed that the unit of the formed polymer corresponds in composition and structure to the monomeric quaternary salt, and the allyl bond in the polymers is preserved. It can be seen from the studies that, with an increase in the concentration of both the monomer and the initiator, the reaction rate naturally increases.

Extinction Coefficient of Plasmonic Nickel Sulfide Nanocrystals and Gold-Nickel Sulfide Core-Shell Nanoparticles

In the presented work, the molar extinction coefficient of plasmonic heazlewoodite (Ni3S2) nanoparticles and Au-Ni3S2 core-shell nanoparticles is determined for the first time. The results are compared to analogously determined extinction coefficients of pure Au nanocrystals (NCs), which themselves correlate very well with existing literature on the subject. The measured extinction coefficients at the localized surface plasmon resonance (LSPR) maximum wavelength of nickel sulfide particles are similar to the values of equally sized Au NCs. Therefore, considering the lower cost of the heazlewoodite material, it could be a reasonable alternative for optical applications of nanoparticles showing a LSPR in the visible regime of the electromagnetic spectrum. Furthermore, this study shows, that by growing a Ni3S2 shell onto a pure Au nanocrystal a highly tuneable optical material with variable LSPR frequency and molar extinction coefficient is obtained.

A subtle calculation method for nanoparticle’s molar extinction coefficient: The gift from discrete protein-nanoparticle system on agarose gel electrophoresis

Discrete biomolecule-nanoparticle (NP) conjugates play paramount roles in nanofabrication, in which the key is to get the precise molar extinction coefficient of NPs. By making best use of the gift from a specific separation phenomenon of agarose gel electrophoresis (GE), amphiphilic polymer coated NP with exact number of bovine serum albumin (BSA) proteins can be extracted and further experimentally employed to precisely calculate the molar extinction coefficient of the NPs. This method could further benefit the evaluation and extraction of any other dual-component NP-containing bio-conjugates.