Li-Jin Jiang

Studies of the sulfonation of hypocrellin A and the photodynamic actions of the product

Abstract Hypocrellin A (HA), a perylenoquinone derivative, is an efficient phototherapeutic agent. The reaction of HA with Na2SO3 in alkaline solution under reflux led to the formation of the sodium salt of 14-dehydroxy-15-deacetyl-hypocrellin A-13-sulfonate (13-SO3Na-DDHA). ESR studies in combination with chemical methods showed that 13-SO3Na-DDHA was an active photosensitizer. An ESR spectrum, assigned to the semiquinone radical anion of 13-SO3Na-DDHA was generated by illumination of a DMF buffer (1:1 by volume) solution of 13-SO3Na-DDHA in the presence of a series of reductive biomaterials. Using 13-SO3Na-DDHA as photosensitizer and 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as a spin trap, the ESR spectra of DMPO/SO3− radical adduct, ascorbyl radical and DMPO/O2·− radical adduct were obtained. Singlet oxygen formed from 13-SO3Na-DDHA photosensitization was detected by means of the 13-SO3Na-DDHA photosensitized oxidation of anthracene and ESR spectroscopy. It was therefore concluded that the photodynamic actions of 13-SO3Na-DDHA proceeded via both Type 1 and Type 2 reactions.

Pharmacokinetics, Tissue Distribution and Photodynamic Therapy Efficacy of Liposomal‐Delivered Hypocrellin A, a Potential Photosensitizer for Tumor Therapy

Hypocrellin A, from Hypocrella bambusae, is a novel photosensitizer of high singlet oxygen quantum yield for photodynamic therapy (PDT). Tissue distributions were studied in tumor‐bearing mice as a function of time following administration. The tumor model was S‐180 sarcoma transplanted into one hind leg of male Kunming mice; hypocrellin A (HA) was delivered to the mice by intravenous injection of 5 mg/kg of body weight as a suspension either as a unilamellar liposome or in dimethyl sulfoxide (DMSO)‐solubilized saline. The HA was isolated from several tissues and organs, as well as tumors and peritumoral muscles and skin. Quantitation was performed by a high‐performance liquid chromatographic technique with detection that utilizes the native fluorescence of HA. Independent of the delivery system, the dye was retained in tumors at higher concentrations than in normal tissues, except for kidney, liver, lung and spleen. The dye retention in tumors was high and was vehicle dependent. For the liposomal system, the maximal accumulation in tumor and maximal ratios of dye in tumor versus peritumoral muscle and skin occurred 12 h post‐injection; for the DMSO saline system, the maximal ratio occurred earlier, 6 h postadministration. Liposomal delivery improved the selective accumulation of the dye in tumor with higher maximal levels in tumor and higher ratios of tumor‐to‐muscle and tumor‐to‐skin. Levels of dye were very low or not detectable in the brain. The PDT efficacy of HA in the liposome and DMSO saline systems was determined by evaluating the tumor volume regression percent. The PDT efficacy of HA in liposomes was highest when light treatment was performed at 12 h postinjection, consistent with the highest retention of HA in tumors. Similarly, the maximal PDT efficacy in DMSO saline was attained at 6 h postinjection, the highest HA retention point in tumor. Moreover, the peak PDT efficacy of HA in liposomes was much higher than that of HA in DMSO saline and even hematoporphyrin mono‐methylether.

Generation of semiquinone radical anion and reactive oxygen (1O2, O2•− and •OH) during the photosensitization of a water-soluble perylenequinone derivative

Abstract 5(or 8)-sulfonated hypocrellin B (5-SO 3 H-HB) is a water-soluble perylenequinone derivative. Electron spin resonance studies in combination with chemical methods show that 5-SO 3 H-HB is an active photosensitizer. Visible-light illumination of 5-SO 3 H-HB solutions generates a semiquinone radical anion, singlet oxygen, a superoxide radical anion, a hydroxyl radical and hydrogen peroxide. In anaerobic solution, the semiquinone radical anion is predominately photoproduced, via self-electron transfer between the excited and ground states of 5-SO 3 H-HB. In aerobic solution, singlet oxygen is generated in the photosensitization of 5-SO 3 H-HB. In addition, a superoxide radical anion is also generated from 5-SO 3 H-HB on illumination in aerobic solution, via the reduction of oxygen by the semiquinone radical anion. In aqueous solution, the superoxide radical anion formed can rapidly transformed to a hydroxyl radical via the Fenton reaction and via the reaction of the semiquinone radical anion with H 2 O 2 . Based on these results, the pathways for the photodynamic action of 5-SO 3 H-HB have been proposed.

Studies on the chelation of hypocrellin a with aluminium ion and the photodynamic action of the resulting complex

Hypocrellin A (HA), a perylenoquinone derivative, is an efficient phototherapeutic agent. The chelation of HA with aluminium ion (Al3+) results in the formation of a 1:1 complex (Al(3+)-HA). The probable structure for this complex has been proposed. Compared with HA, this complex has a much higher solubility in water. In addition, Al(3+)-HA preserves the 1O2-generating function of HA as demonstrated by diphenylanthracene photo-bleaching experiments. In irradiated oxygen-saturated solutions of Al(3+)-HA superoxide radical anion and hydroxyl radical are formed as verified using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as the electron spin resonance (ESR) spin trapping agent. The production of superoxide radical anion is enhanced significantly in the presence of electron donors such as 1-benzyl-1,4-dihydronicotinamide (BNAH). These results suggest that the photodynamic action of Al(3+)-HA proceeds via both type 1 and type 2 reactions.

Photoreactions of hypocrellin B with thiol compounds

Abstract The photoreactions of hypocrellin B (HB) with thiol compounds (RSH) in ethanol buffer solutions have been investigated by electron spin resonance (ESR), α-phenyl-N-tert-butyl nitrone (PBN) spin trapping and quenching experiments. The mechanism of photothiylation of HB is proposed to involve a nucleophilic anion (RS−) addition of Michael type at the 5 or/and 8 positions followed by oxidation. The effects of oxygen and pH value have been studied. The photoreaction and thermal reaction have been compared.

Spectroscopic study on the photoreduction of hypocrellin A : generation of semiquinone radical anion and hydroquinone

Abstract Hypocrellin A (HA) is an efficient phototherapeutic agent. Illumination of HA in dimethylsulphoxide (DMSO) or DMSO-buffer (1:1 by volume, pH > 6.7) generated a strong electron paramagnetic resonance (EPR) signal. This EPR signal was intensified in the presence of reductants. The EPR spectrum obtained was assigned to the semiquinone radical anion of HA (HA .− ) based on a series of experimental results. Decay of HA .− , attributed to a radical-radical reaction, follows second-order kinetics. In acidic DMSO-buffer (1:1 by volume) solution, no EPR signal of HA .− was detected in the absence or presence of reductant. This was explained by a fast disproportionation of radicals, facilitated by protonation of radicals. The spectrophotometric measurements indicated that on illumination HA was directly reduced to its two-electron reduction product, i.e. hydroquinone, in acidic solution. The absorption maximum of the hydroquinone of HA which is at 496 nm at pH 5.8 shifts bathochromically with increase in pH of the medium. In DMSO or DMSO-buffer (1:1 by volume, pH > 6.7) solutions the semiquinone radical anion of HA was also observed spectrophotometrically. The absorption maximum of HA .− is at around 628 nm. Strong intramolecular hydrogen bonding was considered to exist in the chromophore of HA .− .

Electron spin resonance studies on photosensitized formation of hydroxyl radical by C-phycocyanin from Spirulina platensis.

Visible light (>470 nm) irradiation of an oxygen-saturated solution of C-phycocyanin (C-PC) in the presence of the spin trap 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO) gave an ESR spectrum characteristic of the DMPO-hydroxyl radical spin adduct DMPO-OH. The signal intensities of DMPO-OH adduct were enhanced by superoxide dismutase (SOD) and partly inhibited by catalase. It was partly responsible for the production of DMPO-OH that superoxide anion radical (O.-2) dismutated to generate hydrogen peroxide (H2O2) which decomposed ultimately to generate the highly reactive .OH. In addition, it can be concluded that singlet oxygen (1O2) was an important intermediate according to the strong inhibitory action of 1,4-diazabicyclo[2.2.2]octane (DABCO) and histidine on DMPO-OH formation. The experimental results suggest that photodynamic action of C-PC proceed via both type I and type II mechanisms. Furthermore, the decay kinetics of DMPO-OH adduct, the effects of DMPO and C-PC concentrations as well as irradiation time on DMPO-OH adduct formation were also discussed. Concentration of C-PC should be an important factor to influence the ESR signal intensities of DMPO-OH. Therefore, it may be concluded that reasonably lower concentration of C-PC might prolong the duration of photosensitized formation of .OH and might strengthen the photodynamic action.

Photodynamic action of hypocrellin dyes: structure–activity relationships

Abstract Hypocrellin and its derivatives were compared for their photodynamic effects on human oral cavity epithelial carcinoma KB cell line. The amphiphilicity as well as the singlet oxygen generating quantum yield of the hypocrellin dyes affected their photodynamic activity. The most hydrophilic dyes exhibited the lowest phototoxic activity, whereas the hydrophobic and amphiphilic dyes with higher singlet oxygen-generating quantum yield, exhibited high photodynamic activity. Cysteamine mono- and di-substituted hypocrellin B and cysteine mono-substituted hypocrellin B, demonstrating strong red absorption in the domain of phototherapeutic window (600–900 nm), proper hydrophobic and amphiphilic properties and high photocytotoxicity to KB cells, might prove to be potential phototherapeutic agents.

pH Effect on the spectroscopic behavior and photoinduced generation of semiquinone anion radical of hypocrellin B

Abstract The spectroscopic behavior and photoinduced generation of the semiquinone anion radical (HB ·− ) of hypocrellin B (HB), a naturally occurred pigment, have been investigated in ethanol-buffer (1:1 by volume) in a wide range of pH from −0.6 to 14. The absorption and emission properties of HB are strongly affected by pH. Protonation and deprotonation induce a hypsochromic and a bathochromic shift of the spectra, respectively, and a decrease in the quantum yield of fluorescence. The protonation and deprotonation pKa values of HB in the ground and excited singlet states have been determined by means of spectrophotometric and spectrofluorometric titrations and Forster Cycle calculation. The generation of HB ·− is influenced strongly by pH; HB ·− could not be detected either by electron paramagnetic resonance (EPR) or by spectrophotometric measurements in anaerobic acidic or strong alkaline media, and is stable in anaerobic neutral and weak alkaline environment. ©

STUDIES OF THE TRIPLET STATE PROPERTIES OF HYPOCRELLIN A BY NANOSECOND FLASH PHOTOLYSIS

Abstract Hypocrellin A (HA) is an efficient phototherapeutic agent. Its triplet properties have been investigated in cyclohexane using the nanosecond flash photolysis technique. The triplet—triplet (T—T) absorption spectrum of HA gives three bands with absorption maxima at around 520, 555 and 600 nm. The triplet energy level for HA has been estimated to be about 42.5 kcal mol −1 by the energy transfer method. The extinction coefficients for the T—T absorption of HA have been determined by the ground state depletion method. The triplet quantum yield of HA is 0.78±0.04 (by reference to benzophenone).