THE ABSOLUTE VALUE OF THE REACTION RATE CONSTANT OF BILIRUBIN WITH SINGLET OXYGEN IN D2O

Abstract— The chemical reaction rate constant of bilirubin with singlet oxygen in basic aqueous solution has been redetermined to be 3.5 × 10⁸ M^-1 s^-1 by a competitive technique using a 1,3-diphenylisobenzofuran in sodium dodecyl sulfate micelles. Bilirubin also physically quenches a singlet oxygen with a rate constant of 9 × 10⁸ M ^-1 s^-1. The lifetime of singlet oxygen in D₂O solution has been determined to be 35 μ s . The absorption cross-section for the molecular oxygen ³∑_g-→¹δ _g + 1 v electronic transition at 1.06μn in aqueous solution is unexpectedly larger than the gas paase cross-section.     

REFLECTANCE SPECTROSCOPY FOR THE CHARACTERIZATION OF PHOTOCHROMISM IN THE CRYSTALLINE STATE

Abstract— The Kubelka-Munk theory for diffuse reflectance has been applied to a quantitative study of photochromism in the crystalline state. For three systems investigated it was found possible to assign first order rate constants to the thermal relaxation process and estimate the pre-exponential factor A and the activation energy E_a in Arrhenius equation. For the fading of the red photocolored form, Λ_max=490 mμ, of benzaldehyde phenylhydrazone A = 1.4×10⁸ min^-1 and E_a= 15.7 kcal mole^-1. For the fading of the blue photocolored form, Λ_max=590 mμ, of 2–(2,4-dinitrobenzyl)pyridine A= 5×10¹⁴ min^-1 E_a =23.3 kcal mole^-1, Cinnamaldehyde semicarbazone showing 'reversed phototropy' has a photoactivated state, Λ_max=400 mμ, which in dark is transformed into a strongly absorbing yellow species, Λ_max= 430 mμ with A = 14 × 10¹⁰ min^-1 and E_a= 18.7 kcal mole-¹.     

Near-infrared Light Emission from Nanomolar-level Singlet Molecular Oxygen Generated with an Ultra low Concentration Mixture of Hypochlorite and Hydrogen Peroxide

Abstract— We report the detection of a weak near-infrared light emission originating from 8 nM singlet molecular oxygen (¹O₂) produced in a mixture of 1 m M hypochlorite (OC1^-) and 8 n M hydrogen peroxide (H₂O₂). The measurements were made with a highly sensitive detection system for ultraweak light emission in the 1.0-1.5 μm wavelength region. The emission intensity exhibited linear dependence for H₂O₂ concentrations in the range of 8-670 n M . The mixture containing a lower concentration (33 μ M ) of OCl^- pseudocontinuously emitted near-infrared light for 5 s. The rate constant for ¹O₂ production obtained from the kinetic analysis agrees with that previously reported. Our results demonstrate the possibility of measuring very low concentrations of ¹O₂ in a OCi-/H₂O² mixture as well as ¹O₂ production in intact living systems.     

THE IN VITRO PHOTOCHEMISTRY OF BIOLOGICAL MOLECULES—II. THE TRIPLET STATES OF β-CAROTENE AND LYCOPENE EXCITED BY PULSE RADIOLYSIS

Abstract— –Pulse radiolysis has been used to excite the triplet states of β-carotene (τ# 9μ sec) and lycopene (τ= 8μsec) in hexane solution, both in the presence and absence of naphthalene as a triplet sensitiser. The absorption spectra of both triplets have been measured in the range 430–550 nm and have thus been extended into the region of the corresponding singlet absorptions. The overlap of the triplet and singlet spectra is discussed in relation to in vivo studies. Extinction coefficients of 1.3±0.1 × 10⁵ l/mole cm for β-carotene triplet 515 nm and 3.9±0.2 × 10⁵ l/mole cm for lycopene triplet at 525 nm were obtained. Isomerisation of the all- trans polyenes used was detected and preliminary measurements indicate that the yield of isomerisation was greater than the triplet yield. The rate of triplet energy transfer from naphthalene to β-carotene was estimated to be 1.5 × 10¹⁰ l/mole sec. The corresponding value for lycopene was 1.4× 10¹⁰ l/mole sec. The measured efficient quenching of triplet β-carotene by oxygen may occur by an energy transfer mechanism, leading to the formation of singlet oxygen (¹Δ_g. This would suggest that the triplet energy level of β-carotene lies between 121 and 94 kJ mole^-1.     

Quenching of Singlet Oxygen by a Carotenoid–Cyclodextrin Complex: The Importance of Aggregate Formation

The Girard's reagent P derivative of canthaxanthin ((GRP)₂-canthaxanthin), a dicationic carotenoid, forms a highly water-dispersible complex with (2-hydroxypropyl)-γ-cyclodextrin. The UV–visible light spectrum of the complex is consistent with some degree of aggregation, but the spectrum is independent of concentration from 7.5 to 750 μ m . Stern-Vomer plots for singlet-oxygen quenching by the complex are linear over a concentration range of 0–20 μ m . In the presence of 1 m m (2-hydroxypropyl)-γ-cyclodextrin, the singlet-oxygen quenching constant for the complex is 7.9 ± 0.9 × 10⁸  m ⁻¹s⁻¹. This is about an order of magnitude lower than the singlet-oxygen quenching constants for (GRP)₂-canthaxanthin in various organic solvents. The properties of the complex are also compared with the properties of (GRP)₂-canthaxanthin solubilized in neat water and in water containing various detergents. The singlet-oxygen quenching constant for (GRP)₂-canthaxanthin in micelles depends strongly on the specific detergent used, varying from 9.4 × 10⁸  m ⁻¹s⁻¹ for hexadecyltrimethylammonium bromide (CTAB) to 1.24 ± 0.4 × 10¹⁰  m ⁻¹s⁻¹ for sodium dodecyl sulfate. The small quenching constant in CTAB micelles correlates with spectroscopic evidence for aggregation of the (GRP)₂-canthaxanthin in this detergent.     

FLUORESCENCE CHARACTERISTICS OF INDOLES IN NON-POLAR SOLVENTS: LIFETIMES, QUANTUM YIELDS AND POLARIZATION SPECTRA

Abstract— Fluorescence lifetimes, quantum yields and polarization spectra were measured for indole, 3-methylindole and 2,3-dimethylindole in non-polar solvents. The results indicate simultaneous emission from thermally equilibrated ¹L_a and ¹L_b levels, with ¹L_a→¹ A dominating the 2,3-dimethylindole emission, and ¹L_b→¹ A dominating the indole emission. These results are consistent with previous assignments of the 0-0 transitions in absorption for these compounds. Radiative rates are: ¹L_a→¹ A , 2·0 × 10⁸ S^-1 and ¹L_b→¹ A . 0·62 → 10⁸ S^-1. In addition, the temperature dependence of the excitation and emission spectra are presented, which show that aggregation occurs with these indoles in hydrocarbons below approximately - 110°C. Possible applications to tryptophyl emission in the hydrophobic interiors of proteins are briefly discussed.     

FUNCTIONAL ROLE OF CALCIUM IN PHOTORECEPTOR CELLS

Abstract— Ca-uptake by disc membranes prepared from frog rod photoreceptor outer segments was examined. Ca-uptake study revealed two affinity sites which were saturated with 10–⁵ M and 10–³ M of ATP. When disc membranes in 20 m M Tris-HCl buffer (pH 7.5) were stored at -20°C for 6 h, more than 95% of Ca-uptake activity was lost. Ca-uptake activity was, however, preserved if the disc membrane suspension was mixed with 1–10m M ATP and stored at -20°C. Furthermore the reactivation of Ca-uptake was observed if disc membranes, which had lost Ca-uptake ability by storing at 4°C for 3 h, were mixed with 10 m M ATP and then frozen at -20°C for 5 h or 28 h (ATP-induced ATP-dependent Ca-uptake). When the contents of ATP bound to disc membranes were measured during a brief aging at 37°C, the decrement of bound ATP content was correlated well with the decreasing of Ca-uptake activity. Carbonylcyanide m -chlorophenylhydrazone (CCCP), a conductor of protons, inhibited Ca-uptake activity and half maximal inhibition was achieved at 2 × 10–⁸ M. When 10–⁶ M of CCCP was added to the ⁴⁵Ca-accumulated disc membranes, rapid release of ⁴⁵Ca from the disc membranes was observed. These results suggest that ATP may play a role in the Ca-pump regulation in disc membranes and a [H⁺] gradient across disc membrane may be linked to Ca-uptake mechanism.     

MECHANISM OF PHYSICAL QUENCHING OF SINGLET MOLECULAR OXYGEN BY CHLOROPHYLLS and RELATED COMPOUNDS OF BIOLOGICAL INTEREST

Abstract— The rate constant k₅/ > for physical quenching of singlet oxygen O₂(δ₁;) by the sensitizer in dye-sensitized photooxygenations is determined in the case of chlorophylls a and b (7.3 times 10⁸, 4.2 times 10⁸ M^-1 s^-1 respectively), pheophytins a and b (7.4 times 10⁷, 3.0 times 10⁷ M^-1 s^_1 respectively), tetraphenylporphyrin (4.4 times 10⁷ M^-1 s^_1), magnesium tetraphenylporphyrin (5.0 times 10⁸ M^-1 s^_1), zinc tetraphenylporphyrin (1.5 times 10⁸ M^-1 s^_l) and protoporphyrin IX-dimethylester (9.1 times 10⁷ M ^-1 s^_1) in benzene. These sensitizers show a linear correlation between log k_s^O , and their half-wave oxidation potentials and the value of the slope is similar to that observed for various compounds such as phenols. It is concluded that (i) the interaction between chlorophylls and related compounds with singlet oxygen may involve an exciplex as for phenols, and (ii) physical quenching may be envisaged as a spin-orbit-induced intersystem crossing within the exciplex.     

CURCUMIN-DERIVED TRANSIENTS: A PULSED LASER AND PULSE RADIOLYSIS STUDY

Abstract In this paper we report a time-resolved investigation of transients derived from curcumin, which may be intimately involved in the processes leading to its biological activity. Fluorescence and triplet quantum yields are respectively 0.06 and 0.11. The high percentage of internal conversion is proposed to proceed via H-transfer within the thermodynamically favored enol structure of what is formally a 1,3-diketone. The triplet energy (191 ± 2 kJ mol⁻¹), natural lifetime (1.5 μs) and self-quenching rate constant (5.0 × 10⁸ L mol⁻¹ s⁻¹) have been determined. Oxygen quenching of the triplet leads to the production of singlet oxygen with unit efficiency. Curcumin quenches the latter species very inefficiently (2.5 × 10⁵ L mob⁻¹ s⁻¹). The curcumin radical has been produced via three mechanistically distinct methods. This species is unreactive toward oxygen but is repaired by vitamins C and E and anthralin.     

A PULSED LASER SYSTEM FOR THE STUDY OF MICRO-SECOND DELAYED FLUORESCENCE FROM PLANTS

Abstract—A new laser system is designed for the investigation of S delayed fluorescence. This is intended to overcome some undesirable features present in other current designs, such as low actinic light fluxes and long illumination times, or slow responses owing to long turn-off times of the stimulation source and the presence of fluorescence artifacts.
The system uses a pulsed argon-ion laser and bears little resemblance to either the modified Becquerel phosphoroscope used by Lavorel (1971) or to those systems requiring shutters. Cheapness of design together with the ability to adjust such parameters as pulse duration easily, make this design attractive to most laboratories in which short (10 μs) high intensity (10⁵ W m^-2) pulses with rapid (1.5 μs) turn-off times and high on-to-off contrast ratios (10⁶:1) are required.
Certain design criteria are strictly imposed in order to produce kinetic data that can be meaningfully analysed.     

CHLORIDE ENHANCEMENT OF QUENCHING OF TRIPLET METHYLENE BLUE BY Fe(II)

Abstract— The influence of chloride ion on the rate of decay of triplet methylene blue in 0.01 M acid in the absence and presence of ferrous ions was investigated by means of laser flash-photolysis monitored by kinetic spectrophotometry. Chloride weakly accelerates decay of ³MBH^2± in aqueous solution in the absence of Fe(II). Quenching of ³MBH²⁺ by Fe(II) is more strongly catalyzed by Cl^- in both water and 50 v/v% aq. CH₃CN. The uncatalyzed quenching constant, k ₅, is of the order of 1 × 10⁶ M ^-1 s^-1 while in 4.8 M aqueous chloride ( μ – 7.2 M ) k ₅= (37.2 ± 1.8) × 10⁶ M ^-1 s^-1. A possible role of chloride is as a bridging species in quenching via electron transfer between ³MBH²⁺ and Fe(II).     

THE QUENCHING OF SINGLET OXYGEN BY AMINO ACIDS AND PROTEINS

Abstract— The physical quenching of singlet molecular oxygen (¹Δ_g) by amino acids and proteins in D₂O solution has been measured by their inhibition of the rate of singlet oxygen oxidation of the bilirubin anion. Steady-state singlet oxygen concentrations are produced by irradiating the oxygenated solution with the 1–06 μm output of a Nd-YAG laser, which absorbs directly in the electronic transition ¹Δ_g+ 1 v →³Σ_g^-. The rate of quenching by most of the proteins studied is approximated by the sum of the quenching rates of their amino acids histidine, tryptophan and methionine, which implies that these amino acids in the protein structure are all about equally accessible to the singlet oxygen. The quenching constants differ from those obtained by the ruby-laser methylene-blue-photosensitized method of generating singlet oxygen, or from the results of steady-state methylene-blue-photosensitized oxidation, where singlet oxygen is assumed to be the main reactive species. The singlet oxygen quenching rates in D₂O, pD 8, are (10⁷ℒ mol^-1 s^-1): alanine 0–2, methionine 3, tryptophan 9, histidine 17, carbonic anhydrase 85, lysozyme 150, superoxide dismutase 260, aposuperoxide dismutase 250.     

THE EFFECT OF MOLECULAR POLARIZATION ON THE ELECTROCHROMISM OF CAROTENOIDS—I. THE INFLUENCE OF A CARBOXYLIC GROUP

Abstract— Electrochromism of oriented all- trans -β-apo-8'-carotenoic acid is studied in thin capacitors. The linear electrochromism is very strong, in contrast to that of symmetrical carotenoids. It is proportional to the first derivative of the absorption spectrum. The quadratic electrochromism can be described as a superposition of fractions proportional to the first and second derivatives of the absorption spectrum. The permanent dipole moment difference between the ground state and the excited state of the carotenoic acid molecule is Δμ= 3.6 × 10^-29 C·m (±20%) (10.7 Debyes). The polarizability difference parallel to the long axis of the molecule is Δα|| = 1.17 × 10^-37 C·m²·V^-1 (±20%) (1050 ų). Furthermore, the relative permittivity of the solid carotenoic ethyl ester is _r= 3.5 ± 0.2.
Δμ is due to the polarizing force of the carboxylic group. This force is equivalent to a mean local electric field of F _t≅3 × 10⁶V/cm. Such a "local field" may also be exerted on a symmetrical carotenoid in the membrane of photosynthesis, e.g. by asymmetrical complex formation with a polarizing molecule. To obtain an effective permanent field of F _p≅ 2 × 10⁶V/cm across the membrane, as postulated in photosynthesis, a local field of F _l≅ 5.5 × 10⁵ V/cm would be sufficient. F _p is shown to be directed from inside to outside of the thylakoid. To realize this, e.g. a positive polar (i.e. electron-attracting) complex partner of the carotenoid, located more to the inside of the thylakoid, can be postulated.     

PHOTOBLEACHING OF A CYANINE DYE IN SOLUTION AND IN MEMBRANES

Abstract— N,N'-bis(2-ethyl-1,3-dioxolane)-kryptocyanine (EDKC), a lipophilic dye with a delocalized positive charge, photosensitizes cells to visible irradiation. In phosphate-buffered saline (PBS), EDKC absorbs maximally at 700 nm (ε= 1.2 × 10⁵ M⁻¹ cm⁻¹) and in methanol, the absorption maximum is at 706 nm (ε= 2.3 × 10⁵ M⁻¹ cm⁻¹). EDKC partitions from PBS into small unilamellar liposomes prepared from saturated phospholipids and into membranes prepared from red blood cells (RBC) and binds to human serum albumin (HSA). The EDKC fluorescence maximum red shifts from 713 nm in PBS to 720–725 nm in liposomes and RBC membranes and the fluorescence intensity is enhanced by factors of 14–35 compared to PBS (φ= 0.0046). EDKC is thermally unstable in PBS (T_1/2= 2 h at 1.3 × 10⁻⁵ M EDKC), but stable in methanol. In liposomes and RBC membranes, EDKC is 10 times more stable than in PBS, indicating that it is only partially exposed to the aqueous phase. Quenching of EDKC fluorescence in liposomes and RBC membranes by trinitrobenzene sulfonate also indicates that EDKC is not buried within the membranes. Photodecomposition of EDKC was oxygen-dependent and occurred with a low quantum yield (6.4 × 10⁻⁴ in PBS). Singlet oxygen was not detected upon irradiation of EDKC in membranes or with HSA since the self-sensitized oxidation of EDKC occurred at the same rate in D₂O as in H₂O and was not quenched by sodium azide or histidine.     

ON THE TRIPLET-TRIPLET ENERGY TRANSFER FROM CHLOROPHYLL TO CAROTENE IN TRITON X 100 MICELLES

Abstract— The kinetics of the triplet-triplet energy transfer of chlorophyll α (Cha) to β carotene (Car) has been investigated in Triton X100 micelles by 353 nm laser flash photolysis. This transfer consists of an intramicellar process involving pigment species located in the same micelle. A kinetic model using a bimolecular treatment leads to a rate constant of the energy transfer in the micellar phase ( k _tm≅ 6 × 10⁸ M ^-1 s^-1) lower than the previously determined values in homogeneous solvents ( k _t≅ 4.6 ≅ 10⁹ M ^-ls^-l); this result shows the high microviscosity of the micellar core. In addition, the apparent bimolecular rate constant ( k _t≅ 5.0 ≅ 10¹⁰ M ^-l s^-1) appears to be an order of magnitude higher than in homogeneous solvents. The lifetime of the carotene triplet state is the same in the hydrophobic core of Triton X100 micelles (τ a = 7.7 μs) as in organic solvents (hexane or carbon disulfide). The transfer yield is controlled by the distribution of chlorophyll and carotene molecules in the micelles.     

THERMODYNAMICS OF PORPHYRIN BINDING TO SERUM ALBUMIN: EFFECTS OF TEMPERATURE, OF PORPHYRIN SPECIES and OF ALBUMIN-CARRIED FATTY ACIDS

Abstract Porphyrin binding to serum albumin was studied at the molecular level probing the effects of: porphyrin self-aggregation, porphyrin species, temperature and protein-bound fatty acids. Human serum albumin was found to have a single high-affinity site for porphyrin monomers, with binding constants of 2 x 10⁶, 5 x 10⁷ and 3 x 10⁸ (37^o C, neutral pH, M ⁻¹), for hemato-, deutero- and protoporphyrins, respectively. Three equilibria models for the dimer binding were developed and tested. The data were found to fit best with a model proposing a single high-affinity binding site for the dimer, independent of and different than the monomer site. The binding constants of the hematoporphyrin and deuteroporphyrin dimers to human serum albumin (37^o C, neutral pH, M^−l) being 4 x 10* and 5 x 10⁸ respectively. The temperature dependence (Dp and HSA, 22-37^o C) of the monomer binding showed the process to be entropy-driven (δG^o= -45 kJ mol⁻¹; δS^o=+146 kJ mol⁻¹; δH^o= 0 kJ mol⁻¹). For the dimer binding, the enthalpy change was found to be highly temperature-dependent implying continuous changes in the heat capacity of the system over the entire temperature range, the trend at the 37^o C region fitting an entropy-driven process. The monomer vs dimer differences in temperature dependence strongly support separate and independent binding sites for these species. Similar thermodynamics were determined for fatty-acid carrying as well as for fatty-acid free HSA, with mild quantitative (but not qualitative) shifts.     

Fast Redox Perturbation of Aqueous Solution by Photoexcitation of Pyranine

Abstract— Intense illumination (60-120 MW/cm²) of an oxygen-free aqueous solution of pyranine (8-hydroxypyrene-l,3,6-tri-sulfonate) by the third harmonic frequency of an Nd-Yag laser (355 nm) drives a two successive-photon oxidative process of the dye. The first photon excites the dye to its first electronic singlet state. The second photon interacts with the excited molecule, ejects an electron to the solution and deactivates the molecule to a ground state of the oxidized dye (φ⁺). The oxidized product, φ⁺, is an intensely colored compound (Λ_max= 445 nm, ε= 43 000 ± 1000 M ⁻¹ cm⁻¹) that reacts with a variety of electron donors like quinols, ascorbate and ferrous compounds. In the absence of added reductant, φ⁺ is stable, having a lifetime of -10 min. In acidic solutions the solvated electrons generated by the photochemical reaction react preferentially with H⁺. In alkaline solution the favored electron acceptor is the ground-state pyranine anion and a radical, φ, of the reduced dye is formed. The reduced product is well distinguished from the oxidized one, having its maximal absorption at 510 nm with e = 25 000 ± 2000 M^-l cm⁻¹. The oxidized radical can be reduced either by φ^- or by other electron donors. The apparent second-order rate constants of these reactions, which vary from 10⁶ up to 10⁹M⁻¹ s⁻¹, are slower than the rates of diffusion-controlled reactions. Thus the redox reactions are limited by an energy barrier for electron transfer within the encounter complex between the reactants.     

STEADY-STATE NEAR-INFRARED DETECTION OF SINGLET MOLECULAR OXYGEN: A STERN-VOLMER QUENCHING EXPERIMENT WITH SODIUM AZIDE

Abstract— A sensitive near-infrared detection system incorporating improvements to existing methodologies has been used to characterize the sodium azide quenching of the steady-state luminescence of singlet molecular oxygen at 1270 nm. Stern-Volmer plots which were linear up to 80% quenching of the ¹O₂ generated by rose bengal and eosin Y yielded a rate constant of 5.8 ± 0.1 times 10⁸ M ⁻¹ s⁻¹ for the quenching of ¹O₂ in water, while the rate constants obtained in deuterium oxide with the same sensitizers were 6.28 times 10⁸ M ⁻¹ s⁻¹ and 6.91 times 10⁸ M ⁻¹ s⁻¹ respectively. A flow system minimized the effects of photobleaching of the rose bengal. With a mercury arc light source, the instrument can be used in photosensitization experiments to detect low levels of ¹O₂ production in aqueous media.     

TRANSIENT PHENOMENA IN THE PULSE RADIOLYSIS OF RETINYL POLYENES—7. RADICAL ANIONS OF VITAMIN A AND ITS DERIVATIVES

Abstract— Upon e^--pulse irradiation in nonprotic solvents, all- trans retinol (ROH) and retinylmethyl ether (ROMe) form transient species (τ= 0.5–7μs, λ_max=575–590 nm) identifiable as radical anions. Similar species are also formed upon laser pulse photoexcitation of these retinyl derivatives in the presence of N,N-dimethylaniline in acetonitrile. In contrast, electron transfer or attachment to all- trans retinyl acetate (ROAc) and palmitate (ROPa) results in 'instantaneous' loss of carboxylate anions from electron adducts giving the retinylmethyl radical (R-, λ_max= 395 nm, τ_k > 100 μ,s); the radical anions in these cases are too short-lived to be detected by nanosecond pulse radiolysis. The lifetimes of radical anions of ROH and ROMe are very sensitive to water and alcohols (e.g. k_q = 10⁷ M ^-1 s^-1 with methanol as quencher for ROH^- in tetrahydrofuran). Based on these findings, the spectral dissimilarity of the one-electron reduction products from ROH and ROAc in alcohols and aqueous micelles becomes explainable in terms of fast formation of protonated radical anions (RH(OH), τ_1/2, > 100 μs, λ_max=370–375 nm) in the case of ROH and of retinylmethyl radical via loss of AcO^- from radical anion in the case of ROAc. In tetrahydrofuran, the complexation of ROH^- with cations such as Na⁺ and Bu₄N⁺ affects the relative importance of its major decay modes, namely, protonation and dehydroxylation, the latter process being significantly enhanced by the presence of Na⁺.     

ANALYSIS OF FLUORESCENCE KINETICS AND ENERGY TRANSFER IN ISOLATED α SUBUNITS OF PHYCOERYTHRIN FROM Nostoc sp.

Abstract— The fluorescence decay profiles, relative quantum yield, and transmission of the phycoerythrin a subunit, isolated from the photosynthetic antenna system of Nostoc sp., were measured using single picosecond laser excitation. The fluorescence decay profiles were found to be intensity independent for the intensity range investigated (4 × 10¹³ and 4 × 10¹⁵ photons-cm-² per pulse). The decay profiles were fitted to a model assuming both chromophores absorb and fluoresce. The inferred total deactivation rates for the two chromophores, in the absence of energy transfer and when the effects of the response time of the streak camera and the finite pulse width are properly included, are 1.0 × 10¹⁰s' and 1.0 × 10⁹ s ¹ for the s and f chromophores. respectively, whereas the transfer rate between the two fluorophorcs is estimated to be 1.0 × 10¹⁰ s⁻¹ giving a s→ f transfer rate on the order of (100 ps)⁻¹. Steady-atate polarization measurements were found to be equal to those calculated using the rate parameters inferred from the kinetic model fit to the fluorescence decays. The apparent decrease in the relative fluorescence quantum yield and increase of the relative transmission with increasing excitation intensity is suggestive of ground state depletion and upper excited state absorption. Evidence suggests that exciton annihilation is absent within isolated α subunits for the intensity range investigated (4 × 10¹³ to 4 × 10¹⁵ photons-cm ² per pulse).