Numerical Modeling Studies on Photophysical Transitions of Autofluorescent Flavin Coenzymes with A Microfluidic-based Fluorescence Microscope System

Yıl 2025, Cilt: 37 Sayı: 1, 73 - 91, 25.03.2025

Selim Can Dirican , Bahar Tezcan , Süleyman Yiğit Dölek , Barış Demirbay

https://doi.org/10.7240/jeps.1558538

Öz

In this research, numerical modeling studies are presented for a microfluidic-based fluorescence microscope system capable of resolving photophysical transitions of flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) coenzymes that are very sensitive to light and solvent environment, have very small excitation cross-section and weak fluorescence. Different photophysical models were used for both molecules considering the molecular structure, photophysical properties and chemical reactions of FMN and FAD. The electronic states in these models were considered as a first order linear differential equation system and each electronic state population was solved in time, and image and signal data were obtained using the geometric dimensions of the laser excitation area and microscope parameters with the microfluidic chip. The laser excitation intensity at two different flow rates and the effects of redox agents such as ethanol, ascorbate and tryptophan added to the solvent on the normalized fluorescence signal and electronic state populations were simulated. In addition to signal and electronic state analyses, sCMOS image data used in generating the signals were simulated for different experimental conditions and compared with the laser excitation field. The method proposed in the study showed that it has the capacity to distinguish different dark state populations from each other at different flow rates and to resolve the effect of different dark states on normalized fluorescence signals and camera images under different experimental conditions. When compared with existing methods, the numerical findings demonstrate the potential of the method presented in the study to prevent flavin photodegradation to a large extent and enable optimization studies to be conducted regarding the conditions under which photophysical properties of different molecules can be observed.

Anahtar Kelimeler

Flavin mononucleotide, flavin adenine dinucleotide, photophysics, fluorescence spectroscopy and microscopy, electronic state model, numerical simulation

Proje Numarası

124F110

Mikroakışkan Temelli Floresans Mikroskop Sistemi ile Otofloresan Flavin Koenzimlerinin Fotofiziksel Geçişleri Üzerine Nümerik Modelleme Çalışmaları

Öz

Bu araştırmada ışığa ve çözücü ortamına oldukça hassas, çok küçük uyarım kesit alanına sahip ve zayıf floresan ışıma yapabilen flavin mononükleotit (FMN) ve flavin adenin dinükleotit (FAD) koenzimlerinin fotofiziksel geçişlerini çözümleme kapasitesine sahip mikroakışkan temelli bir floresans mikroskop sistemi için nümerik modelleme çalışmaları sunulmuştur. FMN ve FAD’nin moleküler yapısı, fotofiziksel özellikleri ve girdikleri kimyasal reaksiyonlar dikkate alınarak her iki molekül için farklı fotofiziksel modeller kullanılmıştır. Bu modellerde yer alan elektronik durumlar 1. mertebeden lineer diferansiyel denklem sistemi olarak ele alınmış olup her bir elektronik durum popülasyonu zamana bağlı olarak çözülmüş, mikroakışkan çip ile lazer uyarım alanının geometrik boyutları ve mikroskop parametreleri kullanılarak görüntü ve sinyal verisi olarak elde edilmiştir. İki farklı akış hızında lazer uyarım şiddeti, çözücüye eklenen etanol, askorbat ve triptofan gibi redoks ajanlarının normalize floresan sinyaline ve elektronik durum popülasyonlarına olan etkisi simüle edilmiştir. Sinyal ve elektronik durum analizlerine ek olarak sinyallerin oluşturulmasında kullanılan sCMOS görüntü verileri farklı deneysel koşullar için simüle edilmiş ve lazer uyarım alanıyla kıyaslanmıştır. Araştırmada önerilen yöntem farklı akış hızlarında farklı karanlık durum popülasyonlarının birbirinden ayırt edilebilirliğini ve farklı deneysel koşullarda değişen karanlık durumların normalize floresan sinyaline ve kamera görüntülerine olan etkisini çözümleme kapasitesine sahip olduğunu göstermiştir. Mevcut yöntemlerle kıyaslandığında, elde edilen sayısal bulgular, çalışmada sunulan yöntemin flavin foto-bozunumunu büyük ölçüde önleyebilme potansiyelini ispatlamıştır ve farklı moleküllerin fotofiziksel özelliklerinin hangi koşullarda gözlemlenebileceği ile ilgili optimizasyon çalışmalarının yapılmasına olanak sağlamaktadır.

Anahtar Kelimeler

Flavin mononükleotit, flavin adenin dinükleotit, fotofizik, floresans spektroskopisi ve mikroskobu, elektronik durum modeli, nümerik simülasyon

Destekleyen Kurum

TÜBİTAK

Proje Numarası

124F110

Teşekkür

Bu makale Türkiye Bilimsel ve Teknolojik Araştırma Kurumu (TUBİTAK) tarafından fon desteğine hak kazanan 124F110 numaralı 3501 projesinin bilimsel bir ürünüdür. Yazarlar ve proje ekibi olarak bu araştırmaya katkılarından dolayı TÜBİTAK’a teşekkür ederiz.

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