Alternative Biosynthetic Starter Units Enhance the Structural Diversity of Cyanobacterial Lipopeptides.

Puwainaphycins (PUWs) and minutissamides (MINs) are structurally analogous cyclic lipopeptides possessing cytotoxic activity. Both types of compound exhibit high structural variability, particularly in the fatty acid (FA) moiety. Although a biosynthetic gene cluster responsible for synthesis of several PUW variants has been proposed in a cyanobacterial strain, the genetic background for MINs remains unexplored. Herein, we report PUW/MIN biosynthetic gene clusters and structural variants from six cyanobacterial strains. Comparison of biosynthetic gene clusters indicates a common origin of the PUW/MIN hybrid nonribosomal peptide synthetase and polyketide synthase. Surprisingly, the biosynthetic gene clusters encode two alternative biosynthetic starter modules, and analysis of structural variants suggests that initiation by each of the starter modules results in lipopeptides of differing lengths and FA substitutions. Among additional modifications of the FA chain, chlorination of minutissamide D was explained by the presence of a putative halogenase gene in the PUW/MIN gene cluster of Anabaena minutissima strain UTEX B 1613. We detected PUW variants bearing an acetyl substitution in Symplocastrum muelleri strain NIVA-CYA 644, consistent with an O-acetyltransferase gene in its biosynthetic gene cluster. The major lipopeptide variants did not exhibit any significant antibacterial activity, and only the PUW F variant was moderately active against yeast, consistent with previously published data suggesting that PUWs/MINs interact preferentially with eukaryotic plasma membranes.IMPORTANCE Herein, we deciphered the most important biosynthetic traits of a prominent group of bioactive lipopeptides. We reveal evidence for initiation of biosynthesis by two alternative starter units hardwired directly in the same gene cluster, eventually resulting in the production of a remarkable range of lipopeptide variants. We identified several unusual tailoring genes potentially involved in modifying the fatty acid chain. Careful characterization of these biosynthetic gene clusters and their diverse products could provide important insight into lipopeptide biosynthesis in prokaryotes. Some of the variants identified exhibit cytotoxic and antifungal properties, and some are associated with a toxigenic biofilm-forming strain. The findings may prove valuable to researchers in the fields of natural product discovery and toxicology.

Viscosity reduction of isotonic solutions of the photosensitizer TPCS2a by cyclodextrin complexation.

Meso-tetraphenyl chlorin disulphonate (TPCS2a) is a photosensitizer (PS) particularly developed and patented for use in the technology of photochemical internalization (PCI) against cancer. TPCS2a is known to aggregate in aqueous media even at low concentrations (≥0.1 µM) and to form a high-viscosity network at clinically relevant concentrations (mM). The aim of this work was to evaluate the effect of two hydroxypropylated cyclodextrin derivatives of beta and gamma type, respectively i.e. HPßCD and HPγCD, on the aggregation and solubilization of TPCS2a in isotonic solutions. Samples containing micromolar concentrations of TPCS2a were studied spectrophotometrically, while samples containing a clinical relevant concentration (10 mM = 9 mg/ml) of TPCS2a were evaluated by dynamic viscosity measurements. HPßCD was determined to be a more suitable solubilizer of TPCS2a than HPγCD in aqueous media both in the absence and presence of salt. The complexation stoichiometry between TPCS2a/HPßCD at micromolar to millimolar concentrations of TPCS2a was determined to be 1:3 and 1:2 in the absence and presence of isotonic NaCl, respectively. The network of TPCS2a (10 mM) was broken down in the presence of 3% w/v (= 20 mM) HPßCD, i.e. a 1:2 molar ratio between TPCS2a and the cyclodextrin. Formation of the inclusion complex resulted in low viscosity samples both in water and in the presence of isotonic NaCl or phosphate buffered saline (PBS) at 25 °C and 37 °C.

Evaluation of porphyrin loaded dry alginate foams containing poloxamer 407 and ß-cyclodextrin-derivatives intended for wound treatment.

Dry alginate foams intended for antibacterial photodynamic therapy (aPDT) of infected wounds were prepared. aPDT is a treatment modality involving light of appropriate wavelength, oxygen and a photosensitizer (PS) to produce radicals that attack biological targets (e.g. bacteria). The hydrophobic 5,10,15,20-tetrakis(4-hydroxyphenyl)porphyrin (THPP) was chosen as a model PS and loaded into the foams. The foams contained a poloxamer (Pluronic F127) and/or ß-cyclodextrin (ßCD)-derivatives, i.e. methylßCD (MßCD), heptakis(2,6-di-O-methyl)-ßCD (HkßCD) or hydroxypropylßCD (HPßCD).The physical characteristics of the various foams prepared and the THPP load, release and photostability were evaluated. All the foams were thin and flexible and could easily be handled and cut to fit a potential wound. Fast disintegration were observed for dry foams in water or phosphate buffered saline, while in Hanks' balanced salt solution the foams absorbed liquid without losing their integrity. All the foams had a similar THPP load of 0.12-0.13% w/w. The presence of ßCD-derivatives improved THPP release, however, no differences were observed between the different types of ßCD-derivatives evaluated. THPP loaded into the foams was photolabile, which induced a color change combined with fading. Alginate foams formulated with PF127, ßCD-derivatives and THPP demonstrated applicability as delivery vehicles for PSs in aPDT of wounds.

Interaction between curcumin and human serum albumin in the presence of excipients and the effect of binding on curcumin photostability.

Curcumin (Cur) is known to bind to human serum albumin (HSA) which may lead to a reduced phototoxic effect of the compound in the presence of serum or saliva. The influence of excipients on the Cur-HSA binding was studied by HSA florescence quenching and Cur absorption and emission spectroscopy in the presence and absence of the selected excipients. Photostabilty of Cur in the presence of HSA was evaluated, as well as the effect of excipients on HSA bound Cur photodegradation. Cyclodextrins (CDs) (2-hydroxypropyl-ß-cyclodextrin and 2-hydroxypropyl-γ-cyclodextrin) and polymers (polyethylene glycol 400, PEG 400 and Pluronic F-127, PF-127) were selected for the study. CDs and PF-127 seem to decrease Cur binding to HSA, probably through competitive binding. Cur was still bound to HSA in polyethylene glycol (PEG) solutions at the highest investigated concentration (5% w/v). However, high PEG concentration appears to have effect on the protein conformation, as shown by the fluorescence quenching study. Low Cur photostability in the presence of HSA could be improved by the addition of hydroxylpropyl-γ-cyclodextrin (HPγCD) to the samples, whereas PEG and PF-127 showed no effect.

Rheological characterization of an injectable alginate gel system.

BACKGROUND: This work investigates a general method for producing alginate gel matrices using an internal mode of gelation that depends solely on soluble alginate and alginate/gelling ion particles. The method involves the formulation of two-component kits comprised of soluble alginate and insoluble alginate/gelling ion particles. Gelling kinetics, elastic and Young's moduli were investigated for selected parameters with regard to soluble alginate guluronate content, molecular weight, calcium or strontium gelling ions and alginate gelling ion particle sizes in the range between 25 and 125 micrometers. RESULTS: By mixing the two components and varying the parameters mentioned above, alginate gel matrices with tailor-made viscoelastic properties and gelling kinetics were obtained. Final gel elasticity depended on alginate type, concentration and gelling ion. The gelling rate could be manipulated, e.g. through selection of the alginate type and molecular weight, particle sizes and the concentration of non-gelling ions. CONCLUSIONS: Formulations of the injectable and moldable alginate system presented have recently been used within specific medical applications and may have potential within regenerative medicine or other fields.

Formulation and characterization of lyophilized curcumin solid dispersions for antimicrobial photodynamic therapy (aPDT): studies on curcumin and curcuminoids LII.

CONTEXT: Bacterial resistance to antibiotics is increasing and alternative antibacterial treatments like antimicrobial photodynamic therapy (aPDT) are needed. Curcumin is under investigation as a potential photosensitizer in aPDT. OBJECTIVE: The purpose of this study was to develop rapidly dissolving formulations of curcumin that could photoinactivate both Gram-positive and Gram-negative bacteria. MATERIALS AND METHODS: Curcumin solid dispersions with methyl-ß-cyclodextrin and hyaluronic acid (HA), hydroxypropyl methylcellulose (HPMC) or both HA and HPMC were prepared through lyophilization. The lyophilizates were characterized by curcumin drug load [% (w/w)], differential scanning calorimetry, photostability, thermal stability, their ability to form supersaturated solutions and by in vitro photoinactivation of Enterococcus faecalis and Escherichia coli. RESULTS AND DISCUSSION: The lyophilizates were amorphous solid dispersions with a curcumin drug load in the range of 1.4-5.5% (w/w) depending on the included polymer and the ratio between curcumin and the cyclodextrin. The lyophilizates were photolabile, but thermally stable and dissolved rapidly in contact with water to form supersaturated solutions. Selected lyophilizates demonstrated >log 6 reduction of colony forming units/ml of both E. faecalis and E. coli after exposure to low curcumin concentrations (0.5-10 µM) and blue light dose (11-16 J/cm(2)). The high drug load of the lyophilizates, rapid dissolution, ability to form relatively stable supersaturated solutions and the very high phototoxicity towards both E. faecalis and E. coli make these lyophilizates suitable for in vivo aPDT. CONCLUSIONS: This treatment with optimized curcumin formulations should be explored as an alternative to topical antibiotics in the treatment of wound infections.

Poloxamer-based curcumin solid dispersions for ex tempore preparation of supersaturated solutions intended for antimicrobial photodynamic therapy.

Phototoxic effect of curcumin supersaturated solutions toward pathogenic bacteria has already been demonstrated. However, to be useful in the clinical practice, a supersaturated solution needs to be physically and chemically stabile over the relevant time period. Poloxamer-based solid dispersions (SD) intended for ex tempore preparation of a supersaturated solution were designed to simultaneously facilitate dissolution and inhibit precipitation of curcumin in vitro. Due to the transformation of the crystalline compound to an amorphous form, as shown by X-ray powder diffraction, the desired concentration of curcumin could easily be achieved upon hydration of SDs. The efficiency of selected poloxamers (Pluronic® F-127, F-68 and P-123) as the precipitation inhibitors (PIs) and influence of an additional PI (polyethylene glycol 400, hydroxypropyl methylcellulose, and hyaluronic acid) on the physical stability of the solutions were examined by UV-Vis spectrophotometry. HPLC-PDA was employed to evaluate hydrolytic and photolytic stability of curcumin in the SD solutions. At sub-micellar concentrations, Pluronics® F-127 and P-123, but not F-68, delayed curcumin precipitation in aqueous media and protected the compound from hydrolytic degradation up to 24 h. Complete inactivation of Enterococcus faecalis was achieved after exposure to solutions of selected SDs at curcumin concentration ≥1 µM and the light dose of 9.4 J/cm2.

The influence of Pluronics® on dark cytotoxicity, photocytotoxicity, localization and uptake of curcumin in cancer cells: studies of curcumin and curcuminoids XLIX.

In order to apply curcumin as a photosensitizer in photodynamic therapy (PDT) one needs a formulation that can solubilize and stabilize the compound. Pluronics® (Pluronic) are reported to both solubilize and stabilize curcumin against hydrolytic degradation. The aim of the present work was therefore to investigate the influence of Pluronic formulation on the photocytotoxicity of curcumin. Interactions between curcumin and Pluronics were investigated by fluorescence emission and absorption spectroscopy. Cell survival was measured with the MTT assay. The location of curcumin in the cells was investigated with fluorescence microscopy, and the cellular uptake was measured with fluorescence emission spectroscopy. Pluronics P123 and F127 in contrast to Pluronic P85 and PEG 400 may solubilize curcumin under non-cytotoxic conditions. An inverse relationship between the concentration of Pluronic and the photocytotoxicity of curcumin was observed. Curcumin could rapidly translocate across the cell membrane by passive diffusion. The fluorescence from curcumin in the cells (in the cytoplasm) after 1 hour of incubation was lowered by the presence of Pluronics in the formulation. However, the absolute amount of cell-bound curcumin after 1 hour of incubation was independent of the presence of Pluronics. Curcumin was bound more strongly to cells when incubated with formulations without Pluronics compared to cells incubated with curcumin formulations with Pluronics. Incubation of WiDr cells with curcumin for 6 hours resulted in lysosomal accumulation of curcumin independent of the presence of Pluronics. Lysosomally located curcumin could not be observed in HT1080 cells after 6 hours of incubation. The Pluronics P123 and F127 were found to be suitable for solubilizing and stabilizing curcumin, but inhibited photocytotoxic effects of curcumin unless the Pluronic concentration during treatment of the cells was less than 5-10× above the critical micellar concentration.

Neutral natural deep eutectic solvents as anti-biofilm agents.

Natural deep eutectic solvents (NADES) are a class of liquids with promising properties as components in pharmaceutical formulations, such as a low toxicity profile, biodegradability and versatility. Recently, their potential use as anti-biofilm agents has been proposed, due to their ability to solubilize and stabilize biological macromolecules. In the current work, the ability to break down biofilm matrix and the biofilm killing activity of three NADES of neutral pH were investigated against Staphylococcus aureus ATCC 6538 and Pseudomonas aeruginosa ATCC 9027 biofilms. The tested NADES were choline chloride:xylitol (ChX), choline chloride:glycerol (ChG) and betaine:sucrose (BS). Two of the NADES (ChX and ChG) significantly reduced the number of remaining viable cells of both bacterial species in pre-formed biofilm by 4-6 orders of magnitude, while the average biofilm biomass removal for all NADES was 27-67% (S. aureus) and 34-49% (P. aeruginosa). The tested NADES also inhibited biofilm formation of both bacterial species at concentrations at or below 0.5 x the minimal inhibitory concentration (MIC), possibly in part due to observed restrictions imposed by NADES on planktonic growth. These results demonstrate the potential value of neutral NADES as anti-biofilm agents in future antimicrobial preparations.

Studies on curcumin and curcuminoids. XXXIX. Photophysical properties of bisdemethoxycurcumin.

The steady-state absorption and fluorescence, as well as the time-resolved fluorescence properties of bisdemethoxycurcumin dissolved in several solvents differing in polarity and H-bonding capability were measured. The photodegradation quantum yield of the compound in acetonitrile and methanol was determined. The bisdemethoxycurcumin decay mechanisms from the S(1) state were discussed and compared with those of curcumin. The differences in S(1) dynamics observed between bisdemethoxy-curcumin and curcumin could be ascribed to a difference in H-bond acceptor/donor properties of the phenolic OH and a difference in strength of the intramolecular H-bond in the keto-enol moiety within the two molecules.

Protein-Based Systems for Topical Antibacterial Therapy.

Recently, proteins are gaining attention as potential materials for antibacterial therapy. Proteins possess beneficial properties such as biocompatibility, biodegradability, low immunogenic response, ability to control drug release, and can act as protein-mimics in wound healing. Different plant- and animal-derived proteins can be developed into formulations (films, hydrogels, scaffolds, mats) for topical antibacterial therapy. The application areas for topical antibacterial therapy can be wide including bacterial infections in the skin (e.g., acne, wounds), eyelids, mouth, lips, etc. One of the major challenges of the healthcare system is chronic wound infections. Conventional treatment strategies for topical antibacterial therapy of infected wounds are inadequate, and the development of newer and optimized formulations is warranted. Therefore, this review focuses on recent advances in protein-based systems for topical antibacterial therapy in infected wounds. The opportunities and challenges of such protein-based systems along with their future prospects are discussed.

Photostability studies of GarKS peptides for topical formulation development.

There is a growing interest in the use of antimicrobial peptides (AMPs) as potent alternatives for conventional antibiotics, especially in chronic infected wounds. The development of a suitable topical formulation requires a thorough assessment of the photostability profiles of AMPs. In this study, we sought to investigate the photostability of novel Garvicin KS (GarKS; composed of three peptides GakA, GakB, and GakC) peptides either as an individual peptide or in combinations. The photostability of the aqueous peptide solution was determined using Suntest (indoor and outdoor conditions). Furthermore, the antimicrobial efficacy of the peptides was evaluated following UVA irradiations. Photodegradation of the peptides under indoor and outdoor conditions followed first-order kinetics. Individual peptides (GakA, GakB, and GakC) were more prone to photodegradation as compared to combination peptides (GakA+GakB, GakB+GakC, and GakA+GakC) both under indoor and outdoor conditions where the GakA+GakB combination was the most photostable. A combination of GakA+GakB+GakC enhanced photostability under indoor conditions but was reduced under outdoor conditions. A combination of three peptides with an antioxidant (glutathione) or superoxide/hydrogen peroxide scavenger (trehalose) enhanced the photostability of peptides with the highest stability achieved at a peptide:photostabilizer molar ratio of 1:0.8 for glutathione. A nominal increase in the MIC value for the peptide combinations as opposed to a larger increase for individual peptides further supports the photostability effects of combination peptides following UVA irradiations. These results suggest that the GakA+GakB or GakA+GakB+GakC combinations exhibited the highest photostability with excellent antimicrobial efficacy deemed suitable for the development of a potent AMP formulation for topical applications.

Hybrid hydrogels for bacteriocin delivery to infected wounds.

Superficial infections in chronic wounds can prevent the wound healing process by the development of persistent infections and drug-resistant biofilms. Topically applied antimicrobial formulations with stabilized and controlled release offer significant benefits for the effective treatment of wound infections. Bacteriocins are the antimicrobial peptides (AMPs) produced by bacteria that are viable alternatives to antibiotics owing to their natural origin and low propensity for resistance development. Herein, we developed a hybrid hydrogel composed of Pluronic F127 (PF127), ethylenediaminetetraacetic acid (EDTA) loaded liposomes, glutathione (GSH), and the bacteriocin Garvicin KS (GarKS) referred to as "GarKS gel". The GarKS gel exhibited suitable viscosity and rheological properties along with controlled release behavior (up to 9 days) for effective peptide delivery following topical application. Potent in vitro antibacterial and anti-biofilm effects of GarKS gel were evident against the Gram-positive bacterium Staphylococcus aureus. The in vivo treatment of methicillin resistant S. aureus (MRSA) infected mouse wounds suggested potent antibacterial effects of the GarKS gel following multiple applications of once-a-day application for three consecutive days. Altogether, these results provide proof-of-concept for the successful development of AMP loaded topical formulation for effective treatment of wound infections.

A natural deep eutectic solvent (NADES) as potential excipient in collagen-based products.

Natural deep eutectic solvents (NADES) have previously shown antibacterial properties alone or in combination with photosensitizers and light. In this study, we investigated the behavior of the structural protein collagen in a NADES solution. A combination of collagen and NADES adds the unique wound healing properties of collagen to the potential antibacterial effect of the NADES. The behavior of collagen in a NADES composed of citric acid and xylitol and aqueous dilutions thereof was assessed by spectroscopic, calorimetric and viscosity methods. Collagen exhibited variable unfolding properties dependent on the type of material (telo- or atelocollagen) and degree of aqueous dilution of the NADES. The results indicated that both collagen types were susceptible to unfolding in undiluted NADES. Collagen dissolved in highly diluted NADES showed similar results to collagen dissolved in acetic acid (i.e., NADES network possibly maintained). Based on the ability to dissolve collagen while maintaining its structural properties, NADES is regarded as a potential excipient in collagen-based products. This is the first study describing the solubility and structural changes of an extracellular matrix protein in NADES.

Topical antimicrobial peptide formulations for wound healing: Current developments and future prospects.

Antimicrobial peptides (AMPs) are the natural antibiotics recognized for their potent antibacterial and wound healing properties. Bare AMPs have limited activity following topical application attributable to their susceptibility to environment (hydrolysis, oxidation, photolysis), and wound (alkaline pH, proteolysis) related factors as well as minimal residence time. Therefore, the formulation of AMPs is essential to enhance stability, prolong delivery, and optimize effectiveness at the wound site. Different topical formulations of AMPs have been developed so far including nanoparticles, hydrogels, creams, ointments, and wafers to aid in controlling bacterial infection and enhance wound healing process in vivo. Herein, an overview is provided of the AMPs and current understanding of their formulations for topical wound healing applications along with suitable examples. Furthermore, future prospects for the development of effective combination AMP formulations are discussed. STATEMENT OF SIGNIFICANCE: Chronic wound infection and subsequent development of antibiotic resistance are serious clinical problems affecting millions of people worldwide. Antimicrobial peptides (AMPs) possess great potential in effectively killing the bacteria with minimal risk of resistance development. However, AMPs susceptibility to degradation following topical application limits their antimicrobial and wound healing effects. Therefore, development of an optimized topical formulation with high peptide stability and sustained AMP delivery is necessary to maximize the antimicrobial and wound healing effects. The present review provides an overview of the state-of-art in the field of topical AMP formulations for wound healing. Current developments in the field of topical AMP formulations are reviewed and future prospects for the development of effective combination AMP formulations are discussed.

Effect of curcumin and cosolvents on the micellization of Pluronic F127 in aqueous solution.

The drug solubilization capacity of poloxamers like Pluronic F127 (PF127, poloxamer 407) is dependent on the physical form of the polymer; i.e. the distribution between unimers, aggregates, and micelles. Further, the formation of micelles can alter the stability and pharmacological activity of a drug molecule. It is therefore important to understand how the micellization process is influenced by the addition of excipients and drug molecules. Curcumin is considered a photosensitizer in antimicrobial photodynamic therapy (aPDT). The aPDT effect is optimized at a poloxamer concentration just below the critical micellar concentration (CMC). We aimed to evaluate the effect of curcumin in the presence of 1% ethanol (EtOH) or dimethyl sulfoxide (DMSO) on PF127 micellization. These organic solvents are commonly used in topical preparations as a cosolvent or penetration enhancer (in the case of DMSO). The micellization process was investigated by UV-vis spectroscopy, dynamic light scattering (DLS), and differential scanning calorimetry (DSC). The micellization process of PF127 was slightly influenced by the addition of 1% EtOH or DMSO; however, the presence of 20 µM curcumin enhanced the effect. Micellization was favored in PBS compared to MilliQ water. Structures were formed between PF127 and curcumin at poloxamer concentrations ≥0.3 µM which facilitated solubilization of the photosensitizer. The optimal PF127 concentration required to solubilize 20 µM curcumin but avoid micellization was in the range 0.3 µM-0.04 mM in PBS in the presence of 1 % EtOH or DMSO. A careful consideration of the curcumin, cosolvents, and PF127 concentrations is required to enhance the curcumin solubility and prevent the PF127 micellization.

Preformulation studies on novel garvicin KS peptides for topical applications.

Antimicrobial peptides (AMPs) are emerging as a viable alternative to antibiotics attributable to their potent antimicrobial effects and low propensity for resistance development, especially in chronic infected wounds. The development of an optimized topical formulation of AMPs is thus warranted. Preformulation studies for determination of the suitability and optimization requirements of AMPs in topical formulation development are important. Therefore, we sought to investigate the preformulation studies for a novel bacteriocin garvicin KS (GarKS), which is composed of three peptides (GakA, GakB, and GakC). The effects of physiological fluids and varying temperatures on GarKS peptide stability were determined. The antimicrobial effects of the peptides and their combinations were evaluated in Staphylococcus aureus (methicillin sensitive and resistant strains). Furthermore, their effects on fibroblast viability and proliferation were determined. The GarKS peptides were stable in water and PBS at room and physiological temperatures, however, the peptides were significantly degraded in simulated wound fluid. The antimicrobial and fibroblast cell viability/proliferation effects of either individual GarKS peptides or their combinations varied. A careful consideration of the peptide stability, antimicrobial efficacy, and fibroblast viability/proliferation effects suggests GakA+GakB as a potent combination for the development of an optimized topical formulation of GarKS peptides.

Investigation of the antimicrobial effect of natural deep eutectic solvents (NADES) as solvents in antimicrobial photodynamic therapy.

Natural deep eutectic solvents (NADES) are a third class of liquids, separate from water and lipids. Some NADES, especially those containing organic acids, have been suggested to possess antimicrobial properties. Such properties may be advantageous when NADES are used as solvents in e.g. antimicrobial photodynamic therapy. However, to control the toxicity of acid-containing NADES, they must retain their specific qualities upon dilution. Hence, the aims of this study were to investigate the effect of dilution on the acid-containing NADES network, their antimicrobial activity on different planktonic microorganisms, and their influence on phototoxicity when used as solvents for a photosensitiser. Four bacteria and one fungus were exposed to the NADES, CS (citric acid:sucrose) and MFG (malic acid:fructose:glucose) (molar ratios 1:1 and 1:1:1, respectively), at ≤1:200 dilution. Additionally, the antimicrobial properties of the NADES were studied in Escherichia coli in terms of pH and chelating effects. In investigations of phototoxicity, the microorganisms were exposed to the photosensitiser meso-tetra(p-hydroxyphenyl)porphine (THPP; 1nM) dissolved in diluted NADES combined with blue light (27J/cm2). The eutectic network appeared to remain upon dilution ≤1:200. CS (1:200) was less toxic than an equal concentration of citric acid in the Gram-negative bacteria Klebsiella pneumoniae and E. coli (p<0.05). A higher degree of phototoxicity was induced in E. coli (~1% survival) when THPP was dissolved in CS or MFG than in phosphate buffer (~61% survival; p<0.05). No conclusion could be drawn as to whether the observed toxicity in E. coli exposed to NADES was due to the pH of the solutions or chelation of outer membrane-bound cations.

Localized internal radiotherapy with 90Y particles embedded in a new thermosetting alginate gel: a feasibility study in pigs.

BACKGROUND: Internal radiotherapy requires the localization of the radionuclide to the site of action. A new injectable alginate gel formulation intended to undergo immediate gelation in tissues and capable of encapsulating radioactive particles containing 90Y was investigated. METHODS: The formulation was injected intramuscularly, into the bone marrow compartment of the femur and intravenously, respectively, in pigs. The distribution of radioactivity in various tissues was determined. RESULTS: Following intramuscular injection, more than 90% of the radioactivity was found at the site of injection. Following injection into bone marrow, 30-40% of the radioactivity was retained at the site of injection, but a considerable amount of radioactivity was also detected in the lungs (35-45%) and the liver (5-18%). Following intravenous injection, 80-90% of the radioactivity was found in the lungs. CONCLUSION: The present formulation appears suitable for localized radiotherapy in organs and tissues having low perfusion.

Physicochemical characterisation and antimicrobial phototoxicity of an anionic porphyrin in natural deep eutectic solvents.

Natural deep eutectic solvents (NADES) are a newly discovered group of eutectics which has shown promise as a solvent in antimicrobial photodynamic therapy (aPDT). The purpose of this study was to investigate preparations of an anionic porphyrin, meso-tetra-(4-carboxyphenyl)-porphine (TCPP), solubilised in NADES, with regard to their physicochemical and antibacterial properties. The NADES CS (pH∼0), ChX (pH∼4) and MFG (pH∼1) solubilised TCPP with absorption maximum ∼443nm and emission maximum ∼678nm, indicating formation of the TCPP dication. Dilution of TCPP-NADES>1:1 (water) reduced the physical stability of the preparations. The photostability half-lives of TCPP in methanol, MFG, and CS were ∼9h, 6.9h and 3.2h, respectively. Nanomolar concentrations of TCPP solubilised in diluted MFG combined with ⩽27J/cm(2) blue light increased Gram-positive and Gram-negative bacterial phototoxicity, >99.98% and 96% bacterial reduction, respectively, compared to TCPP in PBS/ethanol under equivalent treatment conditions. TCPP solubilised in diluted CS was toxic to bacteria both in the absence (36-72% reduction) and presence of light. TCPP in CS, and in the CS component citric acid, induced a TCPP-concentration dependent increase in Gram-negative phototoxicity relative to controls, which was most pronounced for TCPP-CS. The mechanism behind the increased toxicity is unknown.