Hypericin nanoparticles for self-illuminated photodynamic cytotoxicity based on bioluminescence resonance energy transfer.

The purpose of this study was to investigate the self-sensitization of photosensitizer without an external light source to produce a photodynamic therapy (PDT) effect based on the principle of bioluminescence resonance energy transfer (PDT-BRET). First, we demonstrated that HeLa cells could efficiently express firefly luciferase (FLase) after the firefly luciferase gene was transfected with the FLase-gene plasmid (FLase-GP), and proved that FLase could act on the substrate firefly D-luciferin (FLuc) to produce photons. The generated photons activate the photosensitizer hypericin (Hyp) and induce cytotoxicity. Then, we successfully prepared carboxymethyl chitosan-modified poly(lactic-co-glycolic acid) nanoparticles (CPNPs) loaded with FLuc (FLuc-CPNPs) and with loaded Hyp (Hyp-CPNPs). Their physicochemical and pharmaceutical characteristics indicated that they were an excellent drug delivery system. Characterization of the biological effects showed that they could be located in the mitochondrial, had higher ROS generation and stronger cytotoxicity. In vivo results also showed that PDT-BRET was as effective as classic PDT. PDT-BRET and the related drug delivery system are expected to become a new platform for anticancer therapy.

Breaking the reduced glutathione-activated antioxidant defence for enhanced photodynamic therapy.

Photodynamic therapy (PDT) has been applied in cancer treatment by utilizing reactive oxygen species (ROSs) to kill cancer cells. However, a high concentration of reduced glutathione (GSH) is present in cancer cells and can consume ROSs and sharply reduce the PDT activity. To address this problem, herein, we synthesized a thymine modified Zn phthalocyanine (ZnPc, a monomer and an active form for PDT) and prepared its nanoparticle form (an aggregator and an inactive form) with Hg2+ providing the driving force for the "thymine-Hg2+-thymine" interaction. The nanoparticles could remain in the inactive form during the delivery process in blood. Once endocytosed by cancer cells, the nanoparticles are disintegrated, and deprived of Hg2+ by intracellular GSH, which decreases the level of GSH. Simultaneously, the activity of the released monomer ZnPc is recovered and high PDT activity is observed.

Study on the conformation changes of Lysozyme induced by Hypocrellin A: the mechanism investigation.

The interactions between Lysozyme and Hypocrellin A are investigated in details using time-resolved fluorescence, fourier transform infrared spectroscopy (FTIR), circular dichroism spectroscopy (CD), three-dimensional fluorescence spectra, and thermal gravimetric analysis (TGA) techniques. The results of time-resolved fluorescence suggest that the quenching mechanism is static quenching. FTIR and CD spectroscopy provide evidences of the reducing of α-helix after interaction. Hypocrellin A could change the micro-environmental of Lysozyme according to hydrophobic interaction between the aromatic ring and the hydrophobic amino acid residues, and the altered polypeptide backbone structures induce the reduction of α-helical structures. Moreover, TGA study further demonstrates the structure changes of Lysozyme on the effect of Hypocrellin A. This study could provide some important information for the derivatives of HA in pharmacy, pharmacology and biochemistry.

Characterization and photodynamic activity of a new phthalocyanine nanoparticles.

Nanoparticles of a hydrophobic photosensitizer, tetrakis (3-trifluoromethylphenoxy) zinc phthalocyanine (FPcZn) have been synthesized by using a simple reprecipitation technique. The resulting drug nanoparticles (FPcZn-NP) were spherical, highly monodispersed and stable in aqueous system, without an additional stabilizer. Comparative studies with FPcZn-NP and FPcZn indicated that after the formation of nanoparticles, FPcZn-NP maintained the efficiency of (1)O(2) generation. Further more, the in vitro studies demonstrated that such nanoparticles can be efficiently taken up by Hela cells, which might be resulted to cell death by light irradiation. These properties could make the FPcZn-NP to be a promising candidate in clinical photodynamic therapy.

Graphene oxide noncovalent photosensitizer and its anticancer activity in vitro.

Graphene oxide (GO) was investigated as a potential drug-delivery system due to its special properties and biocompatibility. Thus far, little has been done to use GO as a photosensitive drug-delivery system and to explore its anticancer activity in vitro in photodynamic therapy applications. Here, a novel GO-hypocrellin A (GO-HA) hybrid was prepared by a simple noncovalent method and its photodynamic activity was studied for the first time. The results showed that an efficient loading amount of HA on GO was as high as 1.0 mg mg(-1) and the stability of the hybrid was superior to that of the free hypocrellin A in aqueous solution. Furthermore, GO-HA can be excited by irradiation with light of appropriate wavelength to generate singlet oxygen, and in vitro experiments illustrated that GO-HA was efficiently taken up by tumor cells, and that light irradiation of such impregnated cells resulted in significant cell death. Thus, these properties of GO-HA could possibly make it especially promising for use in clinical photodynamic therapy.

[Spectroscopic studies on the interaction of Elsinochrome A with myoglobin].

The interaction between Elsinochrome A (EA) and myoglobin (Mb) was investigated using UV-Vis and fluorescence spectroscopy. The results suggested that there was a strong interaction between EA and Mb. In the dark, the interaction occurred on the surfaces amidic acid of Mb, but when illuminated, the interactions happened both on amidic acid and the interior structure of hemachrome of Mb. According to the values of the quenching constant, the thermodynamics parameters, the binding constants and the binding sites, it was showed that the binding interaction of EA and Mb was mainly hydrophobic in nature and the quenching mechanism was static quenching procedure. The change in the micro-circumstance of aminos of myoglobin was studied by synchronous fluorescence spectroscopy.

Isolation and anti-inflammatory activity evaluation of triterpenoids and a monoterpenoid glycoside from Harpagophytum procumbens.

A new triterpenoid glycoside, designated harproside (1), and a new iridoid glycoside, named pagide (2), along with six known triterpenoids (3-8), were obtained from the tubers of Harpagophytum procumbens D. C. (devil's claw), and their structures were established through chemical methods and spectroscopic analyses. In an in vitro assay, the six triterpenoids showed anti-inflammatory activity. Compounds 3, 7, and 8 showed significant inhibitory activity against neutrophil respiratory burst stimulated by PMA, while compounds 4, 5, and 6 showed marginal inhibitory activity.

A nanoencapsulated hypocrellin A prepared by an improved microemulsion method for photodynamic treatment.

A new hypocrellin A (HA) encapsulated silica nanoparticles was prepared by an improved microemulsion method based on the unique character of cetyl trimethyl ammonium bromide (CTAB). Stable aqueous dispersions of the HA-loaded nanoparticles, with the diameter about 50 nm, owned superior photo-stability and singlet oxygen generation ability to free HA. In vitro studies demonstrated the active uptake of HA-doped nanoparticles into the cytosol of HeLa (human cervix epithelioid carcinoma) cells. Significant morphology change and phototoxicity to such impregnated tumor cells was observed upon irradiation with light. Thus, the potential of using this method to prepare silica nanoparticles as drug carriers for photodynamic therapy has been demonstrated.

Mitochondria-targeting photosensitizer-encapsulated amorphous nanocage as a bimodal reagent for drug delivery and biodiagnose in vitro.

The use of ceramic nano-carriers containing anti-cancer drugs for targeted delivery that span both fundamental and applied research has attracted the interest of the scientific community. In this paper, a hydrophobic photodynamic therapy drug, hypocrellin A (HA), was successfully encapsulated in water-soluble amorphous silica nanocage (HANC) by an improved sol-gel method. These nanocages are of ultrasmall size, highly monodispersed, stable in aqueous suspension, and retain the optical properties of HA. Moreover, these nanocages can be effectively delivered, subsequently taken up by cancer cells and finally targeted to mitochondria. In addition, incubation time dependent photodynamic efficacy difference between HANC and HA was investigated for the first time. Especially, the nanocages, owning extremely high stable fluorescence comparing with free HA, also have potentials as efficient probes for optical biodiagnose in vitro. All these properties of HANC could possibly make it especially promising to be used as a bimodal reagent for photodynamic therapy and biodiagnose.

A new sol-gel silica nanovehicle preparation for photodynamic therapy in vitro.

We report a facile silica nanovehicle preparation procedure for hydrophobic drug delivery, which is carried out in water without adding any surfactant and additional catalyst. This strategy includes hydrophobic drug nanopaticle preparation by reprecipitation method and in situ hydrolyzation and polymerization to encapsulate this naoparticle using only N-(beta-amimoethyl)-gamma-aminopropyltriethoxysilane (AETPS). To demonstrate this technique hypocrellin A (HA), a hydrophobic photosensitizing anticancer drug, is embedded into silica nanovehicle using this simple method. The resulting HA encapsulated nanovehicles (HANV) are monodisperse and stable in aqueous solution. Comparative studies with free HA and entrapped HA have demonstrated that the encapsulation effect on the embedded photosensitizer nanoparticle significantly enhances the efficacy of singlet oxygen generation and, thereby, the in vitro photodynamic efficacy.

[Study on interaction of caffeine with myoglobin by fluorescence spectroscopy].

The interaction of caffein and myoglobin was investigated by fluorescence spectroscopy and synchronous fluorescence spectroscopy. The intrinsic fluorescence of myoglobin was significantly quenched by caffein under the physiological condition (pH 7.4). The results indicated that caffeine was capable of binding with myoglobin to form a 1:1 complex and the quenching mechanism of myoglobin affected by caffeine was shown to be a static quenching procedure by calculating quenching constant, binding sites and binding constant. According to the thermodynamic parameters, the main binding force of the interaction is electrostatic force and hydrophobic force. The change in the micro-circumstance of aminos of myoglobin was analyzed by synchronous fluorescence spectrometry. The result indicated that caffeine can change the conformation of the protein, leading to the change in the micro-environment of tryptophane and tyrosine residues from hydrophobic environment to hydrophilic environment to different extent.

[Study on the interaction between p-HPcZn and myoglobin].

The interaction of p-HPcZn and myoglobin was studied by fluorescence spectra and synchronous fluorescence spectra methods under the physiological condition. The p-HPcZn can quench the fluorescence of myoglobin effectively, and it is indicated that there is a strong interaction occurring between p-HPcZn and myoglobin. The results of the fluorescence spectra with changing temperature proved that the interaction can lead to the formation of complex of p-HPcZn with myoglobin, and quench the fluorescence of myoglobin through the static quenching mechanism. Dealing with the values of fluorescence spectra, the binding parameter and the binding site of the interaction can be obtained, whose values are 2.481 X 10(5) and 0.444 respectively. In addition, the interaction can change the conformation of myoglobin markedly also.

Iridoid glycosides from Harpagophytum procumbens D.C. (devil's claw).

Iridoid glycosides, harprocumbide A (6''-O-alpha-D-galactopyranosylharpagoside, 1) and harprocumbide B (6''-O-(cis-p-coumaroyl)-procumbide, 2) were isolated from the tubers of Harpagophytum prucumbens D.C., along with nine known iridoid glycosides 6-O-alpha-D-galactopyranosylharpagoside (3), and harpagoside (4), harpagide (5), 8-cinnamoylmyoporoside (6), 8-O-feruloylhapagide (7), procumbide (8), 6''-O-(p-coumaroyl)-procumbide (9), 8-O-(p-coumaroyl)-harpagide (10) and 8-O-(cis-p-coumaroyl)-harpagide (11). Compound 10 showed marginal inhibition activity against macrophages respiratory burst.

[Study on interaction between hypocrellin A and hemoglobin or myoglobin using synchronous fluorescence spectra].

The effects of hypocrellin A (HA) on the conformational changes of hemoglobin and myoglobin were studied using synchronous fluorescence spectroscopy. The results indicated that HA can change the conformation of these two proteins, leading to the change in the micro-environment of tryptophane and tyrosine residues from hydrophobic environment to hydrophilic environment to different extent.

[Study on interaction between CT-DNA and cytochrome C using UV-vis spectroscopy and electrochemistry].

The interaction between CT-DNA immobilized on the positively charged nylon membrane and cytochrome C was studied using UV-Vis reflectance absorption spectroscopy and electrochemistry. It was found that cytchrome C can be adsorbed on the negative charged CT-DNA surface with the active positive charge area. Therefore, CT-DNA can promote the direct electrochemical reaction of cytochrome C.

A tyrosine-modified hypocrellin B with affinity for and photodamaging ability towards calf thymus DNA.

An enhanced photodamaging ability towards CT-DNA was achieved in a tyrosine-modified hypocrellin B by improving the affinity of the sensitizer to DNA.

The photodynamic property improvement of hypocrellin A by chelation with lanthanum ions.

A 1:1 complex of lanthanum ion with hypocrellin A (La3+-HA) possessing high singlet oxygen generation efficiency, large absorbance in the phototherapeutic window, and great water solubility exhibits promising photodynamic properties.