Fluorescence properties of the anti-tumour alkaloid luotonin A and new synthetic analogues: pH modulation as an approach to their fluorimetric quantitation in biological samples

Luotonin A is an alkaloid structurally related to the natural anti-tumour agent camptothecin. The fluorescence behaviour of luotonin A and a series of six analogues is described in the present work. The influence of solvent polarity and pH on the native fluorescence properties of these alkaloids was studied, finding that in organic solvents or in aqueous solutions (pH 5.5–7.2) the neutral form of the luotonin derivatives emit in the region of 410–450 nm but, in both media, acidification to pH values below 3.0 causes a new emission band to appear at about 500 nm. An ESPT reaction occurs due to the protonation of the basic nitrogen atoms of the pentacyclic ring. Acid-base titrations of luotonin A and its derivatives in aqueous and acetonitrile media were carried out in order to determine their pK_a^? values which were around 2, showing these compounds to be very weak bases. In aqueous media, the absence of an iso-emissive point in the emission spectra suggests the existence of more than two species in the proton transfer equilibria. The basicity of the luotonin A derivatives is increased in organic media, and a good correlation between the pK_a^? values and the chemical structure was found. The protonation of luotonin A was also studied by ¹H-NMR and ¹³C-NMR experiments, which proved the protonation of the nitrogen atoms at the positions 5 and 6 of the pentacyclic ring. The fluorescence quantum yields were determined in ethanol and in aqueous solutions under neutral and acidic conditions. The fluorescence quantum yields were higher in water for the case of the more polar compounds, and the opposite result was obtained for the more hydrophobic ones. The remarkable and interesting fluorescence properties of luotonin A prompted the development of its fluorimetric analytical quantitation, obtaining very good analytical features.     

13C, 15N CPMAS NMR and GIAO DFT calculations of stereoisomeric oxindole alkaloids from Cat's Claw (Uncaria tomentosa)

Oxindole alkaloids, isolated from the bark of Uncaria tomentosa [Willd. ex Schult.] Rubiaceae, are considered to be responsible for the biological activity of this herb. Five pentacyclic and two tetracyclic alkaloids were studied by solid-state NMR and theoretical GIAO DFT methods. The ¹³C and ¹⁵N CPMAS NMR spectra were recorded for mitraphylline, isomitraphylline, pteropodine (uncarine C), isopteropodine (uncarine E), speciophylline (uncarine D), rhynchophylline and isorhynchophylline. Theoretical GIAO DFT calculations of shielding constants provide arguments for identification of asymmetric centers and proper assignment of NMR spectra. These alkaloids are 7R/7S and 20R/20S stereoisomeric pairs. Based on the ¹³C CP MAS chemical shifts the 7S alkaloids (δ C3 70–71 ppm) can be easily and conveniently distinguished from 7R (δC3 74.5–74.9 ppm), also 20R (δC20 41.3–41.7 ppm) from the 20S (δC20 36.3–38.3 ppm). The epiallo-type isomer (3R, 20S) of speciophylline is characterized by a larger ¹⁵N MAS chemical shift of N4 (64.6 ppm) than the allo-type (3S, 20S) of isopteropodine (δN4 53.3 ppm). ¹⁵N MAS chemical shifts of N1–H in pentacyclic alkaloids are within 131.9–140.4 ppm.     

Study on the interaction between tabersonine and human serum albumin by optical spectroscopy and molecular modeling methods

The mechanism of interaction between tabersonine (TAB) and human serum albumin (HSA) was investigated by the methods of fluorescence spectroscopy, UV–vis absorption spectroscopy and molecular modeling under simulative physiological conditions. Results obtained from analysis of fluorescence spectrum and fluorescence intensity indicated that TAB has a strong ability to quench the intrinsic fluorescence of HSA through a static quenching procedure. The binding site number n and apparent binding constant K_a, corresponding thermodynamic parameters ΔG, ΔH and ΔS at different temperatures were calculated. The distance r between donor (human serum albumin) and acceptor (tabersonine) was obtained according to the Förster theory of non-radiation energy transfer. The effect of common ions on binding constant was also investigated. The synchronous fluorescence and three-dimensional fluorescence spectra were used to investigate the structural change of HSA molecules with addition of TAB. Furthermore, the study of molecular modeling indicated that TAB could bind to the site I of HSA and hydrophobic interaction was the major acting force, which was in agreement with the binding mode study.     

Remote detection of laser-induced autofluorescence on pure cultures of fungal and bacterial strains and their analysis with multivariate techniques

Remotely sensed laser-induced autofluorescence spectra of pure cultures of fungal strains (Aureobasidium pullulans, Verticillium sp.) and of bacterial strains (Bacillus sp., Pseudomonas sp.) are presented. The strains were isolated from samples collected in a Roman archaeological site (Tropaeum Traiani) near Constanta, Romania. The fluorescence spectra were detected in vivo from a distance of 25 m in the outdoor, using a high spectral resolution fluorescence LIDAR featuring a UV laser (XeCl@308 nm) as an excitation source. All the examined strains, except for the A. pullulans, showed fluorescence features such to allow their characterisation by processing data with multivariate techniques. Both Principal Component Analysis and Cluster Analysis were applied to the data set and compared to discriminate between the examined strains.Results demonstrate the feasibility of fluorescence-based detection and characterisation of fungi and bacteria in the outdoor with a high spectral resolution fluorescence LIDAR. In addition, they show that the proposed processing methods offer a means to discriminate between the fluorescence features due to the investigated samples and that of a fluorescence background of a known spectral shape, as that of the culture medium. This can be exploited for the remote fluorescence mapping of heterotrophic organisms on stone surfaces when the latter show a typical broad fluorescence band.     

Analysis of fluorescence quenching of pyronin B and pyronin Y by molecular oxygen in aqueous solution

The fluorescence quenching of pyronin B and pyronin Y molecules by molecular oxygen in aqueous solution was studied by using steady-state and time-resolved fluorescence and UV-Vis absorption spectroscopy techniques. In order to understand the quenching mechanism, fluorescence decays, absorption and fluorescence spectra of the probes were recorded as a function of the oxygen concentration and temperature. The quenching was found to be appreciable and shows positive deviation in the Stern-Volmer representation obtained from the fluorescence intensity ratio. Fluorescence quenching constants (k_q) were calculated from the τ_o/τ vs. [Q] plots having linear correlation and compared with calculated diffusion-controlled rate constants (k_diff) values. Experimental results were in good agreement with the simultaneous dynamic and static quenching model.     

Fluorometric investigation of the interaction between nitrophenols and 7-hydroxy-4-azidomethylcoumarin

The interaction between nitrophenols and 7-hydroxy-4-azidomethylcoumarin has been investigated by fluorescence and UV-vis absorbance spectroscopy. Quenching mechanisms have been evaluated by fluorescence measurements at different temperatures. Stern-Volmer quenching constant K_sv and corresponding thermodynamic parameters ΔH⁰, ΔG⁰ and ΔS⁰ were calculated. Binding studies concerning the number of binding sites ‘n’ and apparent binding constant ‘K’ were performed by fluorescence quenching method.     

Interactions between β-carboline alkaloids and bovine serum albumin: Investigation by spectroscopic approach

β-Carboline alkaloids are present in medicinal plants such as Peganum harmala L. that have been used as folk medicine in anticancer therapy. BSA¹ is the major soluble protein constituent of the circulatory system, and has many physiological functions including the transport of a variety of compounds. This study is the first attempt to investigate the binding of β-carboline alkaloids to BSA by using a constant protein concentration and varying drug concentrations at pH 7.2. FTIR² and UV–Vis³ spectroscopic methods were used to analyze the binding modes of β-carboline alkaloids, the binding constants and the effects of drug complexation on BSA stability and conformation. Spectroscopic evidence showed that β-carboline alkaloids bind BSA via hydrophobic interaction and van der Waals contacts along with H-bonding with the –NH groups, with overall binding constants of K_{harmine–BSA}=2.04×10⁴ M^?1, K_{tryptoline–BSA}=1.2×10⁴ M^?1, K_{harmaline–BSA}=5.04×10³ M^?1, K_{harmane–BSA}=1.41×10³ M^?1 and K_{harmalol–BSA}=1.01×10³ M^?1, assuming that there is one drug molecule per protein. The BSA secondary structure was altered with a major decrease of α-helix from 64% (free protein) to 59% (BSA–harmane), 56% (BSA–harmaline and BSA–harmine), 55% (BSA–tryptoline), 54% (BSA–harmalol) and β-sheet from 15% (free protein) to 6–8% upon β-carboline alkaloids complexation, inducing a partial protein destabilization.     

Photoinduced interaction between MPA capped CdTe QDs and certain anthraquinone dyes

Photoinduced interaction of mercapto propionic acid (MPA) capped CdTe quantum dots (QDs) with certain anthraquinone dyes namely alizarin, alizarin red S, acid blue 129 and uniblue has been studied by steady state and time resolved fluorescence measurements. Addition of anthraquinone dyes to CdTe QDs results in the reduction of electron hole recombination has been observed (i.e., fluorescence quenching). The Stern-Volmer constant (K_SV), quenching rate constant (k_q) and association constants (K) were obtained from fluorescence quenching data. The interaction of anthraquinone dyes with QDs occurs through static quenching was confirmed by unaltered fluorescence lifetime. The occurrence of electron transfer quenching mechanism has been proved by the negative free energy change (ΔG_et) obtained as per the Rehm-Weller equation.     

A1Σu+-X1Σg+ and B1Πu-X1Σg+ fluorescence of 6Li2 and 6Li7Li excited by a krypton ion laser

The A¹Σ_u⁺-X¹Σ_g⁺ and B¹Π_u-X¹Σ_g⁺ fluorescence of the ⁶Li₂ and ⁶Li⁷Li molecules has been studied for all krypton ion laser lines (468.0–799.3 nm) which might be expected to excite such fluorescence. Only two A-X fluorescence series of ⁶Li₂ were found (one excited by 647.1 nm, and one by 752.5 nm). No A-X fluorescence series of ⁶Li⁷Li was found. Five B-X fluorescence series of ⁶Li₂ were found (one each excited by 468.0, 476.2, and 530.9 nm, and two by 568.2 nm). Four B-X fluorescence series of ⁶Li⁷Li were found (one each excited by 468.0 and 482.5 nm and two by 520.8 nm). Calculated Einstein A coefficients and lifetimes for these transitions are also given.     

Design,synthesis, and anti-tobacco mosaic virus (TMV) activity of glycoconjugates of phenanthroindolizidines alkaloids

Glycoconjugates of phenanthroindolizidine alkaloids targeting tobacco mosaic virus (TMV) RNA were designed, synthesized, and evaluated for their antiviral activity against TMV for the first time. The glycoconjugation of $(S)$ -6-O-desmethylantofine (2) and 14-hydroxyltylophorines (3–6) was accomplished in three ways (O-glycosylation manner, using carbamoyloxy as linker arm, and using 1,2,3-triazole as linker arm) with three different sugar units (glucose, galactose, and mannose). The glycoconjugates showed improved water solubility and molecule polarity compared with phenanthroindolizidine alkaloids. The bioassay results showed that C6 was a suitable position for glycoconjugation and O-glycosylation can increase the antiviral activity of phenanthroindolizidine alkaloids indicating that the introduction of sugar units can improve the antiviral activity profile of glycoconjugates. Two O-glycosides of $(S)$ -6-O-desmethylantofine, (13aS)-6-O- $\upbeta $ -d-galactopyranosyl-2,3-dimethoxyphenanthro [9,10-b]-11-indolizidinone (10) and (13aS)-6-O- $\upbeta $ -d-mannopyranosyl-2,3-dimethoxyphenanthro [9,10-b]-11-indolizidinone (11) displayed significant higher activity than commercial ningnanmycin, and thus could be considered for novel therapy against plant virus infection.     

Effect of temperature and pH on interaction between bovine serum albumin and cetylpyridinium bromide: Fluorescence spectroscopic approach

The fluorescence spectroscopic technique has been efficiently employed to investigate the interaction between bovine serum albumin (BSA) and cetylpyridinium bromide (CPB) under different pH and temperature conditions. The binding constant, number of binding sites, thermodynamic parameters such as ΔG, ΔH, ΔS, and nature of binding forces between BSA and CPB were obtained by measuring the steady state fluorescence quenching of BSA by CPB. The experimental results showed that the fluorescence quenching of BSA by CPB was a result of the formation of CPB-BSA complex. The static quenching was confirmed from the Stern-Volmer quenching constant at different temperatures. The effect of CPB on the conformation of BSA was analyzed using synchronous and three-dimensional fluorescence spectroscopy. pH dependence complex formation between BSA-CPB is due to the interaction between cationic side chain of CPB and the net charge developed on BSA. The distance ‘r’ between BSA and CPB was obtained according to the fluorescence resonance energy transfer.     

Spectral and luminescent properties of jet-cooled dinaphthofuran

The fluorescence and fluorescence excitation spectra of jet-cooled dinaphtho[2,1-b:1′,2′-d]furan (dinaphthofuran) molecules, as well as their complexation with inert gases Ar, Kr, and Xe, are studied. The indicatrices of the degree of polarization of fluorescence of dinaphthofuran molecules upon excitation of the electronic transitions S ₀?S ₁ and S ₀?S ₂ are calculated as functions of the intramolecular orientation of the transition dipole moments. The fluorescence polarization spectrum is measured under excitation within the rotational contour of the line of the purely electronic transition v ₀ ⁰ = 29 294 cm^?1. In contrast to complex planar molecules, the S ₀?S ₂ fluorescence excitation spectrum of dinaphthofuran is found to be continuous, with the Q branch of the rotational contour being absent. The fluorescence excitation spectra of van der Waals complexes of dinaphthofuran with inert gases exhibit multiplet lines, which is associated with the helical structure of the molecules studied.     

Spectroscopic study on interaction between bisphenol A or its degraded solution under microwave irradiation in the presence of activated carbon and human serum albumin

In this study, the interaction between bisphenol A (BPA) or its degraded solution under microwave irradiation after their adsorption on activated carbon (AC/MW) and human serum albumin (HSA) was investigated by UV-vis and fluorescence spectroscopy techniques. The results showed that BPA could bind to HSA molecule, which could cause the stretch of peptide chains. Also, the degraded BPA solution with a few residues could still interact with HSA. Otherwise, the influences of pH and ionic strength on the interaction were estimated. The fluorescence quenching modes of HSA initiated by BPA at three temperatures (298, 310 and 315 K) were all obtained using Stern-Volmer and Lineweaver-Burk equations. The number of binding sites (n), binding constants (K_D) and energy transfer efficiency (E) were all calculated. The thermodynamic parameters (ΔH, ΔG and ΔS) and binding distances (r) were all measured at the three temperatures, respectively. Synchronous fluorescence spectroscopy was also carried out.     

Improvement of frequency sensitivity of series photodetector frequency circuit system and its application for Cy5 fluorescence detection

This study elucidated the frequency characteristics of series photodetector frequency circuit system for detection of DNA probe ET996 marked with fluorescence dye Cy5. We developed 48 MHz series photodetector frequency circuit system with good sensitivity for fluorescence detection. In accordance with the theory of series photodetector frequency circuit system, the frequency sensitivity can be improved by adjusting circuit parameters such as A (tan θ), C_q, C₀, and C_p. In this research of A adjustment, the capacitance parameter C_m of 48 MHz series photodetector frequency circuit system was adjusted to improve the frequency sensitivity for detection of fluorescence dye concentration; moreover, the bias of photodetector was also adjusted to improve the frequency sensitivity. In the optimal conditions of capacitance match and photodetector bias, the detection limit of ET996-Cy5 fluorescence dye concentration 2 pmol/L can be measured by 48 MHz sensor system. The results of fluorescence experiment also demonstrated that the frequency shift of 48 MHz sensor system was linearly related to the logarithm of fluorescence dye concentration from 200 nmol/L to 2 pmol/L. The frequency method can be applied simply and the detection limit of ET996-Cy5 fluorescence dye concentration was lower than the conventional fluorescence technique by 2 orders.     

The investigation of the interaction between edaravone and bovine serum albumin by spectroscopic approaches

The fluorescence and ultraviolet spectroscopies were explored to study the interaction between edaravone (EDA) and bovine serum albumin (BSA) under imitated physiological condition. The experimental results show that the fluorescence quenching mechanism between EDA and BSA is a combined quenching (dynamic and static quenching). The binding constants, binding sites, and the corresponding thermodynamic parameters (ΔG, ΔH, and ΔS) of the interaction system were calculated at different temperatures. According to Förster non-radiation energy transfer theory, the binding distance between EDA and BSA was calculated to be 3.10 nm. The effect of EDA on the conformation of BSA was analyzed using synchronous fluorescence spectroscopy. In addition, the effects of some common metal ions Mg²⁺, Ca²⁺, Cu²⁺, and Ni²⁺ on the binding constant between EDA and BSA were examined.     

Resonance fluorescence of gaseous chlorine excited with a single longitudinal mode argon ion laser

Resonance fluorescence spectra of gaseous chlorine, excited with a high power single longitudinal mode argon ion laser which was tuned through the lasing profiles of the 4880-, 4965-, 5017-, and 5145-Å lines, were studied. Five progressions of fluorescence lines were found and assigned as the fluorescence of ³⁵Cl³⁵Cl and ³⁷Cl³⁷Cl isotopic molecules resonantly excited from v = 0 and 1 levels of the X¹Σ state to the B³Π state. The fluorescence line positions agreed with those of the theoretical calculation within 0.2 cm^?1. The fluorescence intensities were very weak due to the extremely small transition probability between the B³Π and the X¹Σ states.     

Study on the interaction of tropisetron hydrochloride and l-tryptophan by spectrofluorimetry and its analytical application

A new method to determine tropisetron hydrochloride with l-tryptophan in the medium with pH=9.0 was studied, which is based on the fluorescence quenching effect of tropisetron hydrochloride on l-tryptophan. The fluorescence quenching mechanism and various factors influencing fluorescence quenching were discussed. Under the optimum conditions, the linear range and detection limit were 0.03-12.0 and 0.01 μg/mL (correlation coefficient r=0.9970), respectively. The calibration curve equation was ΔF=6.17+12.56 C (μg/mL). RSD was 3.4% (c=4.0 μg/mL, n=5); the detection limit estimated (S/N=3) was 0.01 μg/mL. The proposed method had been successfully applied to determine tropisetron hydrochloride in real samples and the obtained results were in good agreement with the results of the official method.     

UAE, MAE, SFE-CO2 and classical methods for the extraction of Mitragyna speciosa leaves

Mitragyna speciosa, a tropical plant indigenous to Southeast Asia, is well known for its psychoactive properties. Its leaves are traditionally chewed by Thai and Malaysian farmers and manual labourers as it causes a numbing, stimulating effect. The present study aims to evaluate alkaloid yield and composition in the leaf extracts. For this purpose we have compared several non-conventional extraction techniques with classic procedures (room temperature or under heating). Dried M. speciosa leaves belonging to three batches of different origin (from Thailand, Malaysia and Indonesia) were extracted using ultrasound-assisted extraction (UAE), microwave-assisted extraction (MAE) and supercritical carbon dioxide extraction SFE-CO₂, using methanol, ethanol, water and binary mixtures. The extracts were compared using an HPLC/ESI-MS analysis of mitragynine and four other related alkaloids which were present in the alkaloid fraction. The extraction technique influences both the raw product yield and the relative alkaloid content of M. speciosa leaves. Of the several methods tested, MAE in a closed vessel at 110 °C (60 W, methanol/water 1:1) gave the highest alkaloid fraction amount, while UAE with an immersion horn at 25 °C (21.4 kHz, 50 W, methanol) showed the best yield for mitragynine.This work may prove to be a useful contribution to forensic, toxicological and pharmacognosy studies. Although the potential applications of M. speciosa alkaloids clearly need further investigation, these results may facilitate the scaling-up of their extraction.     

Fluorescence and fluorescence excitation spectra of jet-cooled carbazole

The fluorescence excitation spectra of jet-cooled carbazole molecules at vibrational temperatures of 55 and 80 K and the fluorescence spectrum of these molecules excited by radiation at the frequency of a pure electronic transition are measured. As the vibrational temperature increases, the excitation spectra exhibit a series of lines of the same symmetry, which are caused by the interaction of the active vibration with a subensemble of optically inactive vibrations. The final symmetry of the totally and nontotally symmetric vibrations is determined from the shape of the rotational contours of the lines of vibronic transitions. The values of a decrease in the frequency of the nontotally symmetric vibrations in the first excited electronic state S ₁ due to their interaction with the electronic state S ₂ are calculated to be up to 100 cm^?1. The frequencies of the pure electronic transitions in the absorption and fluorescence spectra coincide with each other and are equal to 30809 cm^?1, the frequencies of vibrations in the ground state S ₀ exceeding the frequencies of the corresponding vibrations in the excited state S ₁. The degree of polarization of the integral fluorescence is determined for a series of vibronic transitions of the a ₁ and b ₂ final symmetry that are observed in the fluorescence excitation spectra, and the contribution of the intensity with the borrowed polarization θ_⊥ to the integral fluorescence is calculated. It is found that the intensity θ_⊥ is higher for the transitions of the b ₂ symmetry and can reach ≈50%.     

Analysis of the spectroscopic characteristics on the binding interaction between tosufloxacin and bovine lactoferrin

The interaction between tosufloxacin (TELX) and bovine lactoferrin (BLF) in aqueous solution was analyzed by fluorescence spectroscopy and absorbance spectra. The binding parameters and energy-transfer efficiency parameters were determined and the mechanism of interaction was discussed. The effect of tosufloxacin acting on the BLF’s conformation was detected and the unfolding procedure of BLF induced by tosufloxacin was explored by “fluorescence phase diagram”. Following experimental data of fluorescence polarization values P and r, the saturation characteristic of such kind of binding reaction was observed for the first time. The interaction between tosufloxacin and BLF influenced by Ni²⁺ and Co²⁺ were also preliminarily explored in this work.