PICOSECOND KINETICS and A MODEL FOR THE PRIMARY EVENTS OF BACTERIORHODOPSIN

The picosecond absorption kinetics of light adapted bacteriorhodops in (BR) are reported for water and glycerol-water suspensions of purple membrane from Halobacieriwn hulobium at different temperatures (77-300 K) and pH (5-10). It is shown that a photoproduct of BR, called P-BR (pseudo-bacteriorhodopsin). is responsible for the 16 ± 2 ps relaxation lifetime observed under steady state irradiation (i.e. with a train of ps pulses) at room temperature. At 77 K the absorption recovery lifetime is 62 ± 5 ps in good agreement with previous fluorescence lifetime studies. The observed signal is very sensitive to the sample flow rate and at the highest flow rates a fast absorbance increase (≤ 2 ps) is observed at λ > 620 nm. while at 576 nm a similarly fast absorption recovery alter bleaching is observed. These results imply that the reaction BR → K-BR occurs within 2 ps. In order to explain the simultaneous formation of P-BR and K-BR at 77 K. a model for the primary events including a traus-cis isomerization and a protein relaxation about the chromophore is suggested. The same model can also be used in explaining the simultaneous formation of batho- and hypsorhodopsin in some rhodopsins. Finally, a scheme for the last steps in the photocycle of bacteriorhodopsin including protonation, protein relaxation and cis-trans isomerization is proposed     

FLUORESCENCE KINETICS OF SPINACH CHLOROPLASTS MEASURED WITH A PICOSECOND OPTICAL KERR GATE

Abstract. An overview of the reported chlorophyll a fluorescence lifetimes from green plant photosystems is presented and the problems encountered in the measurement of fluorescence lifetime using two currently available picosecond techniques are discussed.
The fluorescence intensity of spinach chloroplasts exposed to 10 ps flashes was measured as a function of time after the flash and wavelength of observation by the ultrafast Kerr shutter technique. Using a train of 100 pulses separated by 6ns and with an average photon flux per pulse of ˜2 times 10¹⁴ photons/cm², the fluorescence intensity at 685 nm (room temperature) was found to decay with two components, a fast one with a 56 ps lifetime, and a slow one with a 220 ps lifetime. The 730 nm fluorescence intensity at room temperature decays as a single exponential with a 100 ps lifetime. The 730 nm fluorescence lifetime was found to increase by a factor of 6 when the temperature was lowered from room temperature to 90 K while the lifetime of 685 and 695 nm fluorescence were unchanged. At room temperature, the fast and slow components at 685 nm are attributed to the emission from pigment system I (PS I) and PS II, respectively. It is likely that the absolute values of lifetimes, reported here, may increase if single ps low intensity flashes are used for these measurements.     

PICOSECOND FLUORESCENCE KINETICS IN SPINACH CHLOROPLASTS AT LOW TEMPERATURE

Abstract— At 77 K the fluorescence from spinach chloroplasts excited using picosecond mode-locked laser pulses at 620 nm is made up of 5 separate kinetic components. Three of these are predominant at short wavelengths. between 650 and 690 nm, and they appear to correspond to the 3 decay phases seen at room temperature. The 2 new components. a 100 ps rise and a 3000 ps decay, characterize the longer (730–770 nm) wavelength fluorescence. The temperature dependence of the kinetic components of the long wavelength fluorescence shows that the 3000 ps decay accounts for essentially all of the large increase in fluorescence yield observed at low temperature. Furthermore, it appears that this increase does not result entirely from an increase in the fluorescence lifetime, as has been proposed. The dependences of these 2 new components (the 100 ps rise and 3000 ps decay) on emission wavelength and temperature are similar enough to suggest they have a common origin, presumably the chlorophyll pigment component C705. The amplitudes (yield/lifetime) of these 2 phases are approximately equal, and they are opposite in sign. Thus. we see evidence of time-resolved excitation transfer from those pigment molecules that absorb the 620 nm radiation to those that give rise to the long wavelength fluorescence at low temperature.     

PICOSECOND FLUORESCENCE STUDY OF PHOTOSYNTHETIC MUTANTS OF Chlamydomonas reinhardii: ORIGIN OF THE FLUORESCENCE DECAY KINETICS OF CHLOROPLASTS

Abstract— The fluorescence decay kinetics of photosynthetic mutants of Chlamydomonas reinhardii which lack photosystem II (PS II), photosystem I (PS I), and both PS II and PS I have been measured. The PS II mutant strain8–36C exhibits fluorescence decay lifetime components of 53, 424 and 2197 ps. The fluorescence decay of a PS I mutant strain12–7 contains two major fluorescence decay components with lifetimes of 152 and 424 ps. The fluorescence decay of mutant strain C2, which lacks both PS II and PS I, is nearly single exponential with a lifetime of 2561 ± 222 ps. In simulations in which it is assumed that wild-type decays are a simple sum of the major decay components of the isolated parts of the photosynthetic unit as measured in the mutants, curves are obtained that fit the wild-type C. reinhardii fluorescence decay data when the absorption cross-sections of PS II and PS I are weighted approximately equally. The 89 ps lifetime component in the wild-type is an average of 53 and 152 ps components arising from excitation transfer to and trapping in PS I and PS II. The single step transfer time in PS I is estimated to be between 100 and 700 fs depending on assumptions about array size. We find that between two and four visits to the PS I reaction center are required before final trapping.     

(Sub)-picosecond spectral evolution of fluorescence in photoactive proteins studied with a synchroscan streak camera system

The spectral evolution of three photoactive proteins has been investigated by measuring the fluorescence with good temporal and wavelength resolution and a high signal-to-noise ratio. Upon excitation at 400 nm wild-type (wt) PYP both at neutral pH and in the low-pH blueshifted pBdark state exhibited a strong quenching of the fluorescence, the major part of which could be described by lifetimes of about 1.7 and 7.7 ps. The remaining fluorescence decay occurred multiexponentially with lifetimes between 30 and 125 ps. Additionally, in wtPYP at neutral pH, a dynamic Stokes shift was found to occur with a time constant of about 0.25 ps. In a PYP preparation that was reconstituted with the chromophore 7-hydroxy-coumarin-3- carboxylic acid rather than the native coumaric acid, and which is therefore not capable of performing the cis-trans-isomerization that initiates the photocycle in wtPYP, the fluorescence was found to decay multiexponentially with lifetimes of 51 ps, 0.33 and 3.77 ns. Additionally, dynamic Stokes shifts were observed with time constants of about 0.1 and 3.5 ps. Upon comparison of the dynamics of this preparation with that of wtPYP the multiexponential decay with lifetimes of 1.7 and 7.7 ps found in wtPYP was attributed to photochemistry of the p-coumaric-acid chromophore. The emission from bacteriorhodopsin mutant D85S upon excitation at 635 nm decays biexponentially with estimated lifetimes of 5.2 and 19.1 ps. No dynamic Stokes shift was observed here. Four lifetimes were needed to describe the decay of the emission from the A* state in the green fluorescent protein. From a target analysis it was concluded that the longer lifetimes are accompanied by a decreasing probability of forming I*, which approaches zero with the longest A* lifetime of 1.5 ns. These observations may be explained by heterogeneity of A and by relaxation of A*. In all three systems studied, multiexponential decay of emission was present, suggesting that heterogeneity is a common feature of these chromophore protein complexes.     

USE OF A PULSED LASER DIODE TO MEASURE PICOSECOND FLUORESCENCE LIFETIMES

Abstract— The use of an inexpensive pulsed laser diode (Hamamatsu picosecond light pulser PLP-01) as the excitation source for a single photon timing spectrolluorimeter with microchannel plate photomultiplier detection was dem-onstrated. The performance of the instrument was tested with two very short-lived fluorescent dyes and two pho-tosynthetic systems with wcll-defined decay characteristics. Individual fluorescence decays were analyzed by modeling with a convolution of the instrument response function to a sum of exponential decay components. Accurate fluorcscence lifetimcs of the dyes cryptocyanine (55 ps in acetone and 83 ps in ethanol) and 1,1'-diethyl-2,2'-dicarbocyanine iodide (13 ps in acetone and 26 ps in ethanol) were obtained by analysis of the decay kinetics with a single exponential component. Fits to the fluorescence decay kinetics of isolated photosystem I particles and intact cyanobacterial cells required three and four decay components. respectively. The decay kinetics of the isolated photosystem I preparation were dominated (99%) by a very fast 9 ps lifetime, reflecting the preparation's small antenna size of approximately 30 chlorophyll a . The cyanobackria showed decay components of 35 ps, 160 ps, 400 ps and 1.95 ns similar to those described previously by Mullincaux and Holzwarth ( Rinchim. Biophys. Acfa 1098 , 68–78, 1991). The performance of the pulsed laser diode as an excitation source for single photon timing is discussed in comparison with conventional sources of picosecond light pulses.     

Time-resolved fluorescence spectra of arterial fluorescent compounds: reconstruction with the Laguerre expansion technique

The time-resolved fluorescence spectra of the main arterial fluorescent compounds were retrieved using a new algorithm based on the Laguerre expansion of kernels technique. Samples of elastin, collagen and cholesterol were excited with a pulsed nitrogen laser and the emission was measured at 29 discrete wavelengths between 370 and 510 nm. The expansion of the fluorescence impulse response function on the Laguerre basis of functions was optimized to reproduce the observed fluorescence emission. Collagen lifetime (5.3 ns at 390 nm) was substantially larger than that of elastin (2.3 ns) and cholesterol (1.3 ns). Two decay components were identified in the emission decay of the compounds. For collagen, the decay components were markedly wavelength dependent and hydration dependent such that the emission decay became shorter at higher emission wavelengths and with hydration. The decay characteristics of elastin and cholesterol were relatively unchanged with wavelength and with hydration. The observed variations in the time-resolved spectra of elastin, collagen and cholesterol were consistent with the existence of several fluorophores with different emission characteristics. Because the compounds are present in different proportions in healthy and atherosclerotic arterial walls, characteristic differences in their time-resolved emission spectra could be exploited to assess optically the severity of atherosclerotic lesions.     

PICOSECOND FLUORESCENCE KINETICS and ENERGY TRANSFER IN THE ANTENNA CHLOROPHYLLS OF GREEN ALGAE*

Single-photon timing measurements on flowing samples of Chlorella vulgaris and Chlamydomonas reinhardtii at low excitation intensities at room temperature indicate two main kinetic components of the fluorescence at open reaction centers (F₀) of photosystem II with lifetimes of approx. 130 and 500 ps and relative yields of about 30 and 70%. Closing the reaction centers progressively by preincubation of the algae with increasing concentrations of 3-(3′,4′-dichlorophenyl)-l,l-dimethylurea (DCMU) and hydroxylamine gave rise to a slow component with a lifetime increasing from 1.4 to 2.2 ns (F_max) The yield of the slow component increased to 65-68% of the total fluorescence yield in parallel to a decrease in the yield of the fast component to a value close to zero at the f_max-level. The 130 ps lifetime of the fast component remained unchanged. The middle component showed an increase of its lifetime from 500 to 1100 ps and of its yield by a factor of 1.5. Spacing of the ps laser pulses by 12 μs allowed us to resolve a new long-lived fluorescence component of very small amplitude which is ascribed to a small amount of chlorophyll not connected to functional antennae. The opposite dependence of the yield of the fast and the slow component on the state of the reaction centers at almost constant lifetimes is consistent with a mechanism of energy conversion in largely separately functioning photosystem II units. Yields and lifetimes of these two components are in agreement with the high quantum yield of photosynthesis. The lower lifetime limit of 1.4 ns of the slow component is assigned to the average transfer time of an excited state from a closed to a neighboring open reaction center and the increase in the lifetime to 2.2 ns is evidence for a limited energy transfer between photosystems II. Relative effects of changing the excitation wavelength from 630 to 652 nm on the relative fluorescence yields of the kinetic components were studied at the fluorescence wavelengths 682, 703 and 730 nm. Our data indicate that (i) the middle component has its fluorescence maximum at shorter wavelength than the fast component and (ii) that the antennae chlorophylls giving rise to the middle component are preferentially excited by 652 nm light. It is concluded that the middle component originates from the light-harvesting chlorophyll alb protein complexes and the major portion of the fast component from the chlorophyll a antennae of open photosystem II reaction centers.     

FLUORESCENCE DECAY KINETICS and CHARACTERISTICS OF BIMOLECULAR EXCITON ANNIHILATION IN CHLOROPLASTS

Abstract— The decay profiles of the fluorescence of dark-adapted spinach chloroplasts (0^A°C) excited with single 30 ps 532 nm laser pulses of varying intensities were measured with a low-jitter streak camera system. By comparing the decay profiles of the fluorescence at low and high laser fluences, i.e. in the absence and presence, respectively, of dynamic bimolecular exciton-exciton annihilation effects, the duration of such dynamic annihilation events can be estimated. A simple model suggests that the influence of bimolecular annihilation events on the fluorescence decay kinetics should disappear within a time interval corresponding to the low intensity, unimolecular lifetime of the exciton population which is subject to exciton-exciton annihilation. The low intensity fluorescence decay profiles are characterized by three to four lifetimes (Reviewed by A. R. Holzwarth, Photochem. Photobiol. 43,707–725, 1986); it is shown here that only the shortest fluorescence components are subject to exciton annihilation, since the kinetics of the fluorescence decay are influenced by annihilations only within the initial 150–200 ps time interval after the excitation pulse. The amplitudes (but not the decay kinetics) of the longer-lived fluorescence components are decreased at high levels of laser pulse excitations, suggesting that these components are derived from the shorter-lived fluorescence decay components. The implications of these results are*discussed within the contexts of current models of the fluorescence in chloroplasts.     

PICOSECOND LASER PHOTOLYSIS OF SQUID RHODOPSIN AT ROOM AND LOW TEMPERATURES

Abstract. Squid rhodopsin extracted with 2% digitonin (pH 10.5 or 7.0) was excited with a 347 nm light pulse from a mode-locked ruby laser at room temperature. Within 19 ps after the excitation, absorbance at 430 nm due to hypsorhodopsin increased and subsequently decreased with a decay time of 45 ± 10 ps. Absorbance at 550 nm due to bathorhodopsin increased with a rise time of 50 ± 10 ps. These results are the first observations of hypsorhodopsin at room temperature and clearly show that hypsorhodopsin is a precursor of bathorhodopsin which has been considered to be the earliest photoproduct in the photobleaching process of rhodopsin.
Hypsorhodopsin appeared with a rise time of 70 ± 10 ps at 421 nm at liquid nitrogen temperature without any bathorhodopsin being observed during the formation of hypsorhodopsin. An experiment using an N₂ laser showed that squid bathorhodopsin converted to lumirhodopsin with a decay time of about 300 ns at room temperature.     

Analysis of Fluorescence Lifetime of Protochlorophyllide and Chlorophyllide in Isolated Etioplast Membranes Measured from Multifrequency Cross-correlation Phase Fluorometry

The fluorescence decays of protochlorophyllide (Pchlide) and of chlorophyllide (Chlide) in wheat etioplast membranes were analyzed using a multiexponential fluorescence decay model. Using different excitation wavelengths from 430 to 470 nm, we found that a triple-exponential model at 14°C and a double-exponential model at — 170°C were adequate to describe the Pchlide fluorescence decay. We discuss the origin of the three fluorescence lifetime components at 14°C on the basis of the dependence of their fractional intensities on the excitation wavelength and by correlating the fractional intensities with integrated fluorescence intensities of different Pchlide forms in steady-state fluorescence spectra. The fluorescence decay of the main Pchlide form, photoactive Pchlide-F657, is shown to have a complex character with a fast component of 0.25 ns and a slower component of about 2 ns. Two lifetime components of 2 ns and 5.5–6.0 ns are ascribed to the second photoactive form, Pchlide-F645, and to nonphotoactive Pchlide forms, respectively. In etioplast membranes preilluminated by a short saturating light pulse, we found a single 5.0 ns component for Chlide-F688 (the Chlide-NADPH: protochlorophyllide oxidoreductase [PORJ-NADP⁺complex) and an additional 1.6 ns component when the formation of Chlide-F696 (the Chlide-POR-NADPH complex) was promoted by exogenous NADPH. From the fluorescence lifetime results we evaluated the quantum yield of the primary photoreaction by Chlide-F696 as being 70%.     

Fluorescence Lifetime Spectroscopy of Glioblastoma Multiforme¶

Fluorescence spectroscopy of the endogenous emission of brain tumors has been researched as a potentially important method for the intraoperative localization of brain tumor margins. We investigated the use of time‐resolved, laser‐induced fluorescence spectroscopy for demarcation of primary brain tumors by studying the time‐resolved spectra of gliomas. The fluorescence of human brain samples (glioblastoma multiforme, cortex and white matter: six patients, 23 sites) was induced ex vivo with a pulsed nitrogen laser (337 nm, 3 ns). The time‐resolved spectra were detected in a 360–550 nm wavelength range using a fast digitizer and gated detection. Parameters derived from both the spectral‐ (intensities from narrow spectral bands) and the time domain (average lifetime) measured at 390 and 460 nm were used for tissue characterization. We determined that high‐grade gliomas are characterized by fluorescence lifetimes that varied with the emission wavelength (>3 ns at 390 nm, <1 ns at 460 nm) and their emission is overall longer than that of normal brain tissue. Our study demonstrates that the use of fluorescence lifetime not only improves the specificity of fluorescence measurements but also allows a more robust evaluation of data collected from brain tissue. Combined information from both the spectraland the time domain can enhance the ability of fluorescencebased techniques to diagnose and detect brain tumor margins intraoperatively.     

EXCITED STATE pKa BEHAVIOUR OF DAPI. A RATIONALIZATION OF THE FLUORESCENCE ENHANCEMENT OF DAPI IN DAPI-NUCLEIC ACID COMPLEXES*

Abstract— The steady state and time resolved fluorescence of the drug and chromosomal staining agent, 4′,6-diamidino-2-phenylindole dihydrochloride, DAPI, was examined under different solvent conditions. In solutions between pH 3 and pH 9 the fluorescence spectral maximum of DAPI was found at 460 nm. The fluorescence decayed with double exponential kinetics, with decay times of 2.86 and 0.144 ns, at all wavelengths below 550 nm. At 550 nm single exponential decay kinetics with a lifetime of 0.153 ns was observed. The fluorescence spectrum could be resolved into two components, the 2.86 ns component having a spectral maximum near 450 nm and the 0.144 ns component having a spectral maximum near 490 nm. The results are rationalized in terms of there being two different configurations of DAPI, one of which undergoes a rapid protonation of the indole ring by proton transfer from the 6-amidinium group in the excited singlet state. The 0.144 ns component is assigned as the fluorescence from the excited state of the protonated indole ring. The results provide an explanation of the fluorescence enhancement in DAPI-nucleic acid complexes.     

Optical and electric signals from dried oriented purple membrane of bacteriorhodopsins

All the intermediates of the bacteriorhodopsin photocycle are excitable with light of suitable wavelength. This property might regulate the activity in the cells when they are exposed in the nature to high light intensity. On the other hand this property is involved in many applications. In this study the ground state and M intermediate of dried oriented samples of wild-type bacteriorhodopsin and its mutant D96N were excited with 406 nm laser flashes. Substantial M populations were generated with quasi-continuous illumination. The decay of the absorption of M intermediate had three components: their lifetimes were very different for laser flash and quasi-continuous illuminations in cases of both bacteriorhodopsin species. The optical answer for the excitation of M intermediate had a lifetime of 2.2 ms. Electric signals for M excitation had large fast negative components and small positive components in the 100 μs time domain. The results are expected to have important implications for bioelectronic applications of bacteriorhodopsin.     

FLUORESCENCE OF BACTERIORHODOPSIN UNDER SUBPICOSECOND LIGHT EXCITATION

Abstract— A streak camera detection technique has been used to record the fluorescence of bacteriorhodopsin at room temperature induced by single subpicosecond light pulses. The fluorescence lifetime of bacteriorhodopsin has been found to be less than 2 ps.     

EXCITED STATE PROPERTIES OF THE N-(DEOXYGUANOSIN-8-YL)-2-AMINOFLUORENE ADDUCTS

The spectroscopic characteristics of adducts derived from the covalent binding of the carcinogen 2-aminofluorene to the C8 position of deoxyguanosine [N-(deoxyguanosin-8-yl)-2-amino-fluorene, dGuo-C8-AF], and from an adduct of similar structure formed with the synthetic polynucleotide poly(dG-dC).poly(dG-dC), were investigated. At 77 K both adducts are characterized by well-defined and rather narrow fluorescence emission spectra with maxima at 370 and 390 nm characteristic of the aromatic, monomolecular 2-aminofluorene (AF) residue. In contrast, at room temperature, the fluorescence is characterized by a broad, structureless emission band with a maximum at 460 nm in aqueous mixtures, shifting to 415 nm in solvents of lower polarity (100% propanol); the maxima are located at intermediate wavelengths in solutions of different propanol/water compositions, and this emission is attributed to an excited state complex (exciplex). The fluorescence quantum yield decreases when either the solvent polarity or the temperature are increased, varying from 5.4% (100% propanol) to 0.04-0.05% (100% H2O). The fluorescence decay profiles of dGuo-C8-AF adducts (measured at the National Synchrotron Light Source facility at the Brookhaven National Laboratory) can be roughly, but not exactly, modeled in terms of two exponential decay components in the range of about 0.3-1.0 ns with the propanol concentration greater than 60%; at lower propanol concentrations, a single short lifetime is observed and in 100% water solutions its value is 0.08 ns. The shorter lifetime, favored in solvent mixtures of higher polarities, is attributed to an exciplex with significant charge-transfer character.(ABSTRACT TRUNCATED AT 250 WORDS)     

A femtosecond time-resolved investigation of dual fluorescence from N6,N6-dimethyladenine

The excited electronic state dynamics of N(6),N(6)-dimethyladenine (DMAde), a molecule known to emit dual fluorescence, has been studied in aqueous solution using femtosecond fluorescence up-conversion spectroscopy. Time profiles of the fluorescence of DMAde excited at lambda= 258 nm were measured at a series of wavelengths in the range 320 nm or= 500 nm), which appeared slightly delayed compared to the UV fluorescence, the long-lived fluorescence component (tau(3)) dominated, the second component (tau(2)) disappeared. The results are consistent with the assumption that DMAde is primarily excited to a short-lived local excited (LE) electronic state that fluoresces mostly in the UV and decays rapidly, on a approximately 0.5 ps timescale, to an intramolecular charge transfer (ICT) state that emits only at longer wavelengths in the visible spectrum. The fluorescence-time profiles and transient fluorescence spectra reconstructed from the time profiles provided further information on secondary relaxation processes within and between the excited states and their non-radiative relaxation to the electronic ground state.     

PICOSECOND DYNAMICS OF THE EXCITED STATE RELAXATIONS IN PHYTOCHROME

Abstract— A comparative study of the picosecond kinetics of rye ( Secale cereale L.) phytochrome, its 39 and 23 kDa chromopeptides and deuterated rye phytochrome has been carried out. Evidence is presented that the fluorescence decay of Pr contains a very short lifetime component (14 ps) which has escaped detection in the fluorescence studies reported so far. Thus, the overall decay is well described by four exponential components, two rapid (14 and 44 ps) and two slower ones (157 and 690 ps). The fluorescence decays of deuterated Pr and of a 39 and 23 kDa chromopeptide of Pr also require the analysis in terms of four exponentials for a good fit. Some of the lifetime and amplitude values obtained differ significantly from the values estimated for Pr. In the chromopeptides, the two longer components have distinctly slower decays. For the two faster components the lifetimes remain approximately the same, but their relative amplitudes vary greatly. In deuterated Pr, the lifetimes are affected only slightly by deuteration. In contrast, the decay amplitudes are strikingly altered. Moreover, from a rate equation simulation modelling the observed fluorescence kinetics, it turns out that the yields for the various deactivation steps in the chromopeptides and in deuterated Pr reveal differences from the corresponding values in Pr. The implications of the results presented with respect to the influence of the protein moiety of Pr on the picosecond relaxation process are discussed.     

Biochemical Imaging of Human Atherosclerotic Plaques with Fluorescence Lifetime Angioscopy

A prototype angioscopy system with fluorescence lifetime imaging microscopy (FLIM) capabilities was built and applied for biochemical imaging of human coronary atherosclerotic plaques. The FLIM angioscopy prototype consisted of a thin flexible angioscope suitable for UV-excited autofluorescence imaging, and a FLIM detection system based on a pulse sampling approach. The angioscope was composed of an imaging bundle attached to a gradient index objective lens and surrounded by a ring of illumination fibers (2 mm outer diameter, 50 μm spatial resolution). For FLIM detection based on the pulse sampling approach, a gated-intensified charge-couple device camera (200 ps temporal resolution) was used. Autofluorescence was excited with a pulsed UV laser (337 nm) and FLIM images were acquired at three emission bands (390/40 nm, 450/40 nm, 550/88 nm). The system was characterized on standard fluorophores and then used to image postmortem human coronary arteries. The FLIM angioscope allowed us to distinguish elastin-dominant plaques (peak emission at 450 nm, ∼1.5 ns lifetimes) from collagen-dominant plaques (peak emission at 390 n, ∼2–3 ns lifetimes) based on their intrinsic fluorescence spectral and lifetime differences. This study demonstrates the potential of FLIM angioscopy for biochemical imaging of human coronary atherosclerotic plaques.     

胞嘧啶水溶液体系辐解的瞬态产物研究

Using the ns pulse radolysis, we studied the characteristic absorption spectrum and kinetic decay of cytosine anion radical (Cyt-). Results showed that the characteristic absorption of Cyt- was located at λ=355±5 nm, and decayed following the first order kinetics with τ1/2=265 ns at pH=7.0. The decay became slower and τ1/2 rapidly rised with the increment of pH value, Cyt- protonated at C6 in acidic solution, and the characteristic absorption was located at λ=310±5 nm, and decayed following the second order kinetics: Cyt- protonated at N3 in aqueous solution of pH≥7, and the characteristic absorption was located at λ=295±5 nm, and decayed following the second order kinetics.