Ultrafast Dynamics of Metal Complexes of Tetrasulfonated Phthalocyanines at Biological Interfaces: Comparison between Photochemistry in Solutions, Films, and Noncancerous and Cancerous Human Breast Tissues

International audienceA promising material in medicine, electronics, opto-electronics, electrochemistry, catalysis, and photophysics, Al(III) phthalocyanine chloride tetrasulfonic acid (AlPcS4) is investigated at biological interfaces of human breast tissue by means of steady-state and time-resolved pump?probe spectroscopies: IR, Raman, UV?vis, fluorescence, and electronic transient absorption by pump?probe spectroscopy. Spectrally resolved pump?probe data were recorded on time scales ranging from femtoseconds to nanoseconds and give insight into molecular interactions and primary events in the interfacial region. The nature of these fast processes and pathways of the competing relaxation processes from the initially excited electronic states in AlPcS4 films and at biological interfaces of human breast cancerous and noncancerous tissues is studied. Comparison between photochemical dynamics in the biological environment of the human breast tissues and that occurring in aqueous solutions is presented. The excited-state absorption (ESA) decays and bleaching recovery of the ground state have been fitted in the time window extending to nanoseconds (0?1 ns). We found that the excited-state dynamics of AlPcS4 at biological interfaces of human breast tissue is extremely sensitive to the biological environment and differs drastically from that observed in solutions and films. We demonstrated that the ultrafast dynamics at biological interfaces is described by three time constants in the ranges of 110?170 fs, 1?7 ps, and 20?60 ps. We were able to ascribe these three time constants to the primary events occurring in phthalocyanine at biological interfaces. The shortest time constants have been assigned to vibrational wavepacket dynamics in the Franck?Condon region down to the local minimum of the excited-state S1. The 1?7 ps components have been assigned to vibrational relaxation in the excited and ground electronic states. In contrast to the dynamics observed in aqueous solutions with the components in the range of 150?500 ps assigned to decay from S1 to the ground electronic state, these slow components have not been recorded in human breast tissue. We have shown that the lifetimes characterizing the first excited-state S1 in the interfacial regions of the breast tissue are markedly shorter than those in solution. It suggests that molecular structures responsible for harvesting of the light energy in biological tissue find their own ways for recovery through some special features of the potential energy surfaces such as conical intersections, which facilitate the rate of radiationless transitions. We found that the dynamics of photosensitizers in normal (noncancerous) breast tissue is markedly faster than that in cancerous tissue

Biochemical fingerprint of colorectal cancer cell lines using label-free live single-cell Raman spectroscopy

Label-free live single-cell Raman spectroscopy was used to obtain a chemical fingerprint of colorectal cancer cells including the classification of the SW480 and SW620 cell line model system, derived from primary and secondary tumour cells from the same patient. High-quality Raman spectra were acquired from hundreds of live cells, showing high reproducibility between experiments. Principal component analysis with linear discriminant analysis yielded the best cell classification, with an accuracy of 98.7±0.3% (standard error) when compared to discrimination trees or support vector machines. SW480 showed higher content of the disordered secondary protein structure amide III band, whereas SW620 showed larger α-helix and β-sheet band content. The SW620 cell line also displayed higher nucleic acid, phosphates, saccharide, and CH2 content. HL60, HT29, HCT116, SW620 and SW480 live single-cell spectra were classified using PCA/LDA with an accuracy of 92.4±0.4% (standard error), showing differences mainly in the β-sheet content, the cytochrome C bands, the CH-stretching regions, the lactate contributions and the DNA content. The lipids contributions above 2900 cm-1 and the lactate contributions at 1785 cm-1 appeared to be dependent on the colorectal adenocarcinoma stage, the advanced stage cell lines showing lower lipid and higher lactate content. The results demonstrate that these cell lines can be distinguished with high confidence, suggesting that Raman spectroscopy on live cells can distinguish between different disease stages, and could play an important role clinically as a diagnostic tool for cell phenotyping

Analysis of Human Colon by Raman Spectroscopy and Imaging-Elucidation of Biochemical Changes in Carcinogenesis

Noninvasive Raman imaging of non-fixed and unstained human colon tissues based on vibrational properties of noncancerous and cancerous samples can effectively enable the differentiation between noncancerous and tumor tissues. This work aimed to evaluate the biochemical characteristics of colon cancer and the clinical merits of multivariate Raman image and spectroscopy analysis. Tissue samples were collected during routine surgery. The non-fixed, fresh samples were used to prepare micrometer sections from the tumor mass and the tissue from the safety margins outside of the tumor mass. Adjacent sections were used for typical histological analysis. We have found that the chemical composition identified by Raman spectroscopy of the cancerous and the noncancerous colon samples is sufficiently different to distinguish pathologically changed tissue from noncancerous tissue. We present a detailed analysis of Raman spectra for the human noncancerous and cancerous colon tissue. The multivariate analysis of the intensities of lipids/proteins/carotenoids Raman peaks shows that these classes of compounds can statistically divide analyzed samples into noncancerous and pathological groups, reaffirming that Raman imaging is a powerful technique for the histochemical analysis of human tissues. Raman biomarkers based on ratios for lipids/proteins/carotenoids content were found to be the most useful biomarkers in spectroscopic diagnostics

Effect of PHRs and PCPs on Microalgal Growth, Metabolism and Microalgae-Based Bioremediation Processes: A Review

In this review, the effect of pharmaceuticals (PHRs) and personal care products (PCPs) on microalgal growth and metabolism is reported. Concentrations of various PHRs and PCPs that cause inhibition and toxicity to growths of different microalgal strains are summarized and compared. The effect of PHRs and PCPs on microalgal metabolism (oxidative stress, enzyme activity, pigments, proteins, lipids, carbohydrates, toxins), as well as on the cellular morphology, is discussed. Literature data concerning the removal of PHRs and PCPs from wastewaters by living microalgal cultures, with the emphasis on microalgal growth, are gathered and discussed. The potential of simultaneously bioremediating PHRs/PCPs-containing wastewaters and cultivating microalgae for biomass production in a single process is considered. In the light of reviewed data, the feasibility of post-bioremediation microalgal biomass is discussed in terms of its contamination, biosafety and further usage for production of value-added biomolecules (pigments, lipids, proteins) and biomass as a whole

Vitamin C—Protective Role in Oxidative Stress Conditions Induced in Human Normal Colon Cells by Label-Free Raman Spectroscopy and Imaging

Colorectal cancer is the second most frequently diagnosed cancer worldwide. Conventional diagnostics methods of colorectal cancer can detect it at an advanced stage. Spectroscopic methods, including Raman spectroscopy and imaging, are becoming more and more popular in medical applications, and allow fast, precise, and unambiguous differentiation of healthy and cancerous samples. The most important advantage of Raman spectroscopy is the ability to identify biomarkers that help in the differentiation of healthy and cancerous cells based on biochemistry of sample and spectra typical for lipids, proteins, and DNA. The aim of the study was to evaluate the biochemical and structural features of human colon cell lines based on Raman spectroscopy and imaging: normal cells CCD-18 Co, normal cells CCD-18 Co under oxidative stress conditions, and normal cells CCD-18 Co at first treated by using tert-Butyl hydroperoxide and then supplemented by vitamin C in high concentration to show the protective role of vitamin C in micromolar concentrations against ROS (Reactive Oxygen Species). Raman data obtained for normal cells injured by ROS were compared with spectra typical for cancerous cells. Statistically assisted analysis has shown that normal ROS-injured and cancerous human colon cells can be distinguished based on their unique vibrational properties. The research carried out proves that label-free Raman spectroscopy may play an important role in clinical diagnostics differentiation of normal and cancerous colon cells and may be a source of intraoperative information supporting histopathological analysis

Surface-Enhanced Raman Spectroscopy Analysis of Human Breast Cancer via Silver Nanoparticles: An Examination of Fabrication Methods

Breast cancer in a traditional way is diagnosed using mammography, computer tomography, ultrasounds, biopsy, and finally, histopathological analysis. Histopathological analysis is a gold standard in breast cancer diagnostics; however, it is time consuming and prone to the human interpretations. That is why new methods based on optical properties of analyzed human tissue samples are needed to be introduced to the clinical practice objective, costless and fast diagnostic protocols. Nowadays, Raman spectroscopy-based methods are gaining more and more importance. Raman spectroscopy and imaging allow to characterize human tissue samples using an electromagnetic radiation from a safe range, and simultaneously, a minimal sample preparation is required. During measurements, a natural differentiation in tissues components’ scattering cross sections is used to build 2D and 3D maps of the chemical component distribution. The paper presents the application of SERS (surface-enhanced Raman spectroscopy) measurements for analysis of human breast cancer (adenocarcinoma). The advantages of SERS application in cancer diagnostics are also discussed. Moreover, the detailed chemical composition of human breast cancer tissue based on Raman bands of DNA/RNA, amino acids, lipids, and proteins which are significantly enhanced is presented. Three different methods of NP preparation are presented, and the effectiveness of Raman signal enhancement of Ag nanoparticles synthetized by these methods is compared. The enhancement effect of NPs synthetized by reduction of silver nitrate with sodium borohydride (method no. 1) and silver nitrate-hydroxylamine hydrochloride reduction (method no. 2) was stronger when compared with the polyol method (method no. 3). Presented SERS results confirmed that the clearly resolved and high-intensity Raman spectra of cancer human breast tissue can be recorded using integration times of the order of fractional seconds and one milliwatt of the excitation laser power

Raman Imaging—A Valuable Tool for Tracking Fatty Acid Metabolism—Normal and Cancer Human Colon Single-Cell Study

Altered metabolism of lipids is a key factor in many diseases including cancer. Therefore, investigations into the impact of unsaturated and saturated fatty acids (FAs) on human body homeostasis are crucial for understanding the development of lifestyle diseases. In this paper, we focus on the impact of palmitic (PA), linoleic (LA), and eicosapentaenoic (EPA) acids on human colon normal (CCD-18 Co) and cancer (Caco-2) single cells using Raman imaging and spectroscopy. The label-free nature of Raman imaging allowed us to evaluate FAs dynamics without modifying endogenous cellular metabolism. Thanks to the ability of Raman imaging to visualize single-cell substructures, we have analyzed the changes in chemical composition of endoplasmic reticulum (ER), mitochondria, lipid droplets (LDs), and nucleus upon FA supplementation. Analysis of Raman band intensity ratios typical for lipids, proteins, and nucleic acids (I1656/I1444, I1444/I1256, I1444/I750, I1304/I1256) proved that, using Raman mapping, we can observe the metabolic pathways of FAs in ER, which is responsible for the uptake of exogenous FAs, de novo synthesis, elongation, and desaturation of FAs, in mitochondria responsible for energy production via FA oxidation, in LDs specialized in cellular fat storage, and in the nucleus, where FAs are transported via fatty-acid-binding proteins, biomarkers of human colon cancerogenesis. Analysis for membranes showed that the uptake of FAs effectively changed the chemical composition of this organelle, and the strongest effect was noticed for LA. The spectroscopy studies have been completed using XTT tests, which showed that the addition of LA or EPA for Caco-2 cells decreases their viability with a stronger effect observed for LA and the opposite effect observed for PA. For normal cells, CCD-18 Co supplementation using LA or EPA stimulated cells for growing, while PA had the opposite impact