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Hemicellulose hydrolysates (HH), which could be an interesting carbon source to feed mixed microbial cultures (MMC) able to accumulate high value-added compounds. This research focused on the evaluation of a culture strategy to achieve the simultaneous biological production of Levulinic Acid (LA) and Polyhydroxyalcanoates (PHA) by MMC fed with a synthetic HH (SHH). The culture strategy involves the use of sequential batch reactors (SBR) to select microorganisms capable of producing LA and PHA. This work proved that the cultivation strategy used allowed the biological production of LA, reaching 37%w/w when the SHH was composed of 85% pentoses. In addition, the simultaneous biological production of LA and PHB was possible when the SHH was enriched with acetate (45% pentoses - 50% acetate). Finally, this study showed that the composition of the SHH impacts directly on the selected microorganism genus and the type and quantity of the value-added compounds obtained.

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The objective of this study was to investigate whether the conjugation of gold nanoparticles (GNPs) to 5-aminolevulinic acid (5-ALA) could enhance the anti-tumor efficiency of photodynamic therapy (PDT) in epidermoid carcinoma cells. The mRNA and protein expression levels were determined by quantitative real-time PCR and western blot, respectively. Cell viability, apoptosis, invasion, and migration were determined by MTT assay, flow cytometry, transwell invasion assay, and migration assay, respectively. Singlet oxygen generation was detected by the singlet oxygen sensor green reagent assay. Our results showed that PDT with 5-ALA and GNPs-conjugated 5-ALA (5-ALA-GNPs) significantly suppressed cell viability, increased cell apoptosis and singlet oxygen generation in both HaCat and A431 cells, and PDT with 5-ALA and 5-ALA-GNPs had more profound effects in A431 cells than that in HaCat cells. More importantly, 5-ALA-GNPs treatment potentiated the effects of PDT on cell viability, cell apoptosis, and singlet oxygen generation in A431 cells compared to 5-ALA treatment. Further in vitro assays showed that PDT with 5-ALA-GNPs significantly decreased expression of STAT3 and Bcl-2 and increased expression of Bax in A431 cells compared with PDT with 5-ALA. In addition, 5-ALA-GNPs treatment enhanced the inhibitory effects of PDT on cell invasion and migration and Wnt/ß-catenin signaling activities in A431 cells compared to 5-ALA treatment. In conclusion, our results suggested that GNPs conjugated to 5-ALA significantly enhanced the anti-tumor efficacy of PDT in A431 cells, which may represent a better strategy to improve the outcomes of patients with cutaneous squamous cell carcinoma.

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The objective of this study was to investigate whether the conjugation of gold nanoparticles (GNPs) to 5-aminolevulinic acid (5-ALA) could enhance the anti-tumor efficiency of photodynamic therapy (PDT) in epidermoid carcinoma cells. The mRNA and protein expression levels were determined by quantitative real-time PCR and western blot, respectively. Cell viability, apoptosis, invasion, and migration were determined by MTT assay, flow cytometry, transwell invasion assay, and migration assay, respectively. Singlet oxygen generation was detected by the singlet oxygen sensor green reagent assay. Our results showed that PDT with 5-ALA and GNPs-conjugated 5-ALA (5-ALA-GNPs) significantly suppressed cell viability, increased cell apoptosis and singlet oxygen generation in both HaCat and A431 cells, and PDT with 5-ALA and 5-ALA-GNPs had more profound effects in A431 cells than that in HaCat cells. More importantly, 5-ALA-GNPs treatment potentiated the effects of PDT on cell viability, cell apoptosis, and singlet oxygen generation in A431 cells compared to 5-ALA treatment. Further in vitro assays showed that PDT with 5-ALA-GNPs significantly decreased expression of STAT3 and Bcl-2 and increased expression of Bax in A431 cells compared with PDT with 5-ALA. In addition, 5-ALA-GNPs treatment enhanced the inhibitory effects of PDT on cell invasion and migration and Wnt/β-catenin signaling activities in A431 cells compared to 5-ALA treatment. In conclusion, our results suggested that GNPs conjugated to 5-ALA significantly enhanced the anti-tumor efficacy of PDT in A431 cells, which may represent a better strategy to improve the outcomes of patients with cutaneous squamous cell carcinoma.

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BACKGROUND: To study the effective therapeutic concentration, drug application duration, irradiation duration and irradiation dosage of 5-aminolevulinic acid photodynamic therapy (ALA-PDT) for treating vitiligo and observe its clinical efficacy. OBJECTIVE: To assess the clinical efficacy of ALA-PDT for treating vitiligo. METHODS: ALA with different drug concentrations, drug application durations, irradiation durations and irradiation dosages were adopted to treat skin lesions caused by vitiligo to determine the effective drug concentration, drug application duration, irradiation duration and irradiation dosage for treating three vitiligo volunteers and observe the therapeutic results. RESULTS: The clinical trial suggested that ALA at a drug concentration of 1.5%, drug application duration for 3 hours, irradiation dosage of 80mw/cm2 and irradiation duration for 20 min was effective in treating vitiligo. Under these parameters, ALA-PDT was effective to the three vitiligo volunteers, with mild pain and feeling of burning but no other adverse reaction during treatment. STUDY LIMITATIONS: Due to the small sample size in this study for the effectiveness of PDT in treating vitiligo and the potential variations in the efficacy for treating the disease at different areas, further studies shall be conducted for confirmation. CONCLUSIONS: ALA with a drug concentration at 1.5%, drug application duration for 3 hours, irradiation dosage of 80 mw/cm2 and irradiation duration for 20 min is effective in treating vitiligo. Therefore, ALA-PDT is safe and effective in treating the disease, with minor adverse events, providing a new method for treating vitiligo in the future.

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Abstract: Background: To study the effective therapeutic concentration, drug application duration, irradiation duration and irradiation dosage of 5-aminolevulinic acid photodynamic therapy (ALA-PDT) for treating vitiligo and observe its clinical efficacy. Objective: To assess the clinical efficacy of ALA-PDT for treating vitiligo. Methods: ALA with different drug concentrations, drug application durations, irradiation durations and irradiation dosages were adopted to treat skin lesions caused by vitiligo to determine the effective drug concentration, drug application duration, irradiation duration and irradiation dosage for treating three vitiligo volunteers and observe the therapeutic results. Results: The clinical trial suggested that ALA at a drug concentration of 1.5%, drug application duration for 3 hours, irradiation dosage of 80mw/cm2 and irradiation duration for 20 min was effective in treating vitiligo. Under these parameters, ALA-PDT was effective to the three vitiligo volunteers, with mild pain and feeling of burning but no other adverse reaction during treatment. Study limitations: Due to the small sample size in this study for the effectiveness of PDT in treating vitiligo and the potential variations in the efficacy for treating the disease at different areas, further studies shall be conducted for confirmation. Conclusions: ALA with a drug concentration at 1.5%, drug application duration for 3 hours, irradiation dosage of 80 mw/cm2 and irradiation duration for 20 min is effective in treating vitiligo. Therefore, ALA-PDT is safe and effective in treating the disease, with minor adverse events, providing a new method for treating vitiligo in the future.

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Sonodynamic therapy (SDT) is emerging as new atherosclerosis treatment. The use of gold nanoparticles (AuNPs) as the vehicle for a sensitizer delivery improves reactive oxygen species formation. In this study, methyl ester of aminolevulinic acid (MALA) gold nanoparticles (MALA:AuNPs) functionalized with polyethylene glycol (PEG) were synthesized by photoreduction and characterized by ultraviolet/visible optical absorption, zeta potential and electron microscopy. The reactive oxygen species generation induced by ultrasound irradiation of MALA:AuNPs solutions was studied by observing the decrease in the 1,3-diphenylisobenzofuran emission band. The potential use of MALA:AuNPs as sensitizer for sonodynamic therapy was investigated on THP-1 macrophages. The cytotoxicity test was also described. The findings suggested that ultrasound combined with MALA:AuNPs provides impressive results in in vitro studies. Sonodynamic therapy with MALA:AuNPs through 2 minutes of ultrasound exposure (1 MHz and 1 W/cm2) culminated with total macrophage reduction. Thus, sonodynamic therapy combined with MALA:AuNPs has potential as a treatment for atherosclerosis.

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Toxic concentrations of monocarboxylic weak acids present in lignocellulosic hydrolyzates affect cell integrity and fermentative performance of Saccharomyces cerevisiae. In this work, we report the deletion of the general catabolite repressor Mig1p as a strategy to improve the tolerance of S. cerevisiae towards inhibitory concentrations of acetic, formic or levulinic acid. In contrast with the wt yeast, where the growth and ethanol production were ceased in presence of acetic acid 5 g/L or formic acid 1.75 g/L (initial pH not adjusted), the m9 strain (Δmig1::kan) produced 4.06 ± 0.14 and 3.87 ± 0.06 g/L of ethanol, respectively. Also, m9 strain tolerated a higher concentration of 12.5 g/L acetic acid (initial pH adjusted to 4.5) without affecting its fermentative performance. Moreover, m9 strain produced 33% less acetic acid and 50-70% less glycerol in presence of weak acids, and consumed acetate and formate as carbon sources under aerobic conditions. Our results show that the deletion of Mig1p provides a single gene deletion target for improving the acid tolerance of yeast strains significantly.

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The present study aims to identify the renewable resources available in Brazil such as açai seed, coconut husks, coffee husks, rice husks, eucalyptus sawdust, grass, soy peel, bamboo, banana stems and banana stalks. To identify such renewable energy sources, samples were examined for their physical and chemical characteristics using X-ray diffraction (XRD), proximate and ultimate analyses, thermogravimetric analysis (TGA), calorific value determination, near-infrared (NIR) spectroscopy, UV spectroscopy, high-pH anion-exchange chromatography (HPAEC-PAD) and accelerated solvent extraction (ASE). Among the biomasses, açai and coffee exhibited higher total sugar content, 67.70% and 62.55%, respectively. Sawdust exhibited low ash, along with the highest calorific value and lignin content. The highest glucose contents were observed in bamboo (44.65%) and sawdust (38.80%). The maximum yield for the bioproducts levulinic acid (LA), formic acid (FA) and furfural were estimated; açai exhibited the highest yield of LA and FA, while coffee exhibited the best furfural yield. All of these properties indicate that the residues are potential candidates for bioenergy production.

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The conversion of biomass-derived levulinic acid (LA) into gamma-valerolactone (GVL) using formic acid (FA) and Fe3(CO)12 as the catalyst precursor was achieved in 92% yield. To mimic a biorefinery setting, crude liquor (containing 20% LA) from the acid hydrolysis of sugarcane biomass in a pilot plant facility was directly converted into GVL in good yield (50%), without the need for isolating LA.

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BACKGROUND: Estimation of the time period that precedes an injury is critical in forensic medicine. However, there is no reliable method that can be used to evaluate the oldness of a lesion. The aim of this work is to develop a fluorimetric method that can be used to follow the aging process of lesions by applying methyl-ALA (MAL) on wounds and by quantifying protoporphyrin IX (PPIX) fluorescence during the healing process. We also aim to understand the changes in PPIX fluorescence by establishing a correlation with histological evaluations during the healing process. METHODS: Standardized linear wounds were made on the dorsum of 72 mice, which were divided in control (MAL -) and experimental (MAL +) groups. In vivo fluorescence spectra (FS) were collected from normal and wound skin sites of control and experimental groups, corresponding to four groups of FS spectra: (a) FS of skin wound after MAL (+/+); (b) FS of normal skin after MAL (-/+); (c) FS of skin wound without MAL (+/-) and (d) FS of normal skin without MAL (-/-). Animals were monitored periodically for 3 months and euthanized. Tissue specimens were processed for histological analysis using design-based stereological methods. Serial cross-sections were analyzed to evaluate the organization of the dermis and epidermis, collagen deposition and cellular proliferation. RESULTS: FS of skin wound with MAL (+/+) showed an expressive intensity increase from the beginning of the experiment to the 34th day, with maximum fluorescence being observed on the ≈ 11 th day after wounding. There was preferential PPIX accumulation in healing sites as compared to adjacent normal skin (+/-) in the early stage of healing. Histological findings allowed correlation of the fluorescence increase mainly with cell proliferation. The drastic decrease in the FS intensity observed in the end of the healing process was correlated with the decrease in the proliferation rate as well as with the presence of new extracellular fibrous materials. CONCLUSIONS: In the mice wound-healing model tested here, it was possible to distinguish whether the injury was in early or advanced stages by using PPIX fluorescence induced by MAL. We conclude that this method is a promising approach to evaluate the age of skin wounding and we hope this work will stimulate human studies to allow this technique to become standardized in forensic medicine.

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The Photodynamic Therapy (PDT) is a cancer treatment based on specific accumulation of a photosensitizer in the malignant tissue and its subsequent irradiation at appropriate wave lengths induce the production of oxygen singlets responsible for peroxidation of cell organelles and cell death. A prototype of a 630 nm non coherent light source designed and constructed at the Centre of Electron Microscopy of The Cordoba National University allowed a successful application of PDT, in nonmelanoma skin tumors, for the first time in this Country. A topical treatment with 20% delta aminolevulinic acid (ALA) in aqueous solution was applied in 100 lesions of actinic keratosis of 27 patients. The results obtained in this study were the following: Complete Remission (RC) 84%, Partial Remission (RP) 10%, No Response (SR) 0% and No data (SD) 6%. In the latter group are included those patients who did not return for reevaluation. The high efficiency plus the excellent cosmetic response and low aggressiveness, make the PDT the method of choice treatment of this skin pathology. In addition to be no invasive and well tolerated, the prototype of light source used in this study, was remarkably effective.

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La Terapia Fotodinámica (PDT) es un tratamiento del cáncer basado en la acumulación específica de una droga fotosensible en el tejido maligno. Su posterior radiación con una longitud de onda apropiada, induce la producción de singuletes de oxígeno responsable de la peroxidación de las organelas y la muerte de las células neoplásicas. En el Centro de Microscopía Electrónica de la Universidad Nacional de Córdoba se diseñó y construyó un prototipo de fuente de irradiación no coherente de 630nm el que posibilitó la aplicación de PDT por primera vez en nuestro país. Este prototipo ha sido aplicado satisfactoriamente en el tratamiento de la queratosis actínica. Fueron tratadas 100 lesiones en 27 pacientes utilizando como fotosensibilizador al ácido d amino levulínico (ALA) al 20 por ciento La activación lumínica duró de 5 a 20 minutos dependiendo de la extensión y profundidad de la lesión. Los resultados obtenidos fueron los siguientes: Remisión Completa de las lesiones (RC) 84por ciento, Remisión parcial (RP) 10 por ciento, Sin respuesta (SR) 0 por ciento y Sin datos (SD) 6 por ciento. En el último grupo están incluidos aquellos pacientes que no retornaron para su evaluación. La alta efectividad, sumada a la inmejorable respuesta cosmética y la reducida agresividad, hacen de PDT el método de elección en el tratamiento de esta patología. El prototipo utilizado en este estudio demostró ser además de no invasivo y bien tolerado, altamente efectivo.

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La Terapia Fotodinámica (PDT) es un tratamiento del cáncer basado en la acumulación específica de una droga fotosensible en el tejido maligno. Su posterior radiación con una longitud de onda apropiada, induce la producción de singuletes de oxígeno responsable de la peroxidación de las organelas y la muerte de las células neoplásicas. En el Centro de Microscopía Electrónica de la Universidad Nacional de Córdoba se diseñó y construyó un prototipo de fuente de irradiación no coherente de 630nm el que posibilitó la aplicación de PDT por primera vez en nuestro país. Este prototipo ha sido aplicado satisfactoriamente en el tratamiento de la queratosis actínica. Fueron tratadas 100 lesiones en 27 pacientes utilizando como fotosensibilizador al ácido d amino levulínico (ALA) al 20 por ciento La activación lumínica duró de 5 a 20 minutos dependiendo de la extensión y profundidad de la lesión. Los resultados obtenidos fueron los siguientes: Remisión Completa de las lesiones (RC) 84por ciento, Remisión parcial (RP) 10 por ciento, Sin respuesta (SR) 0 por ciento y Sin datos (SD) 6 por ciento. En el último grupo están incluidos aquellos pacientes que no retornaron para su evaluación. La alta efectividad, sumada a la inmejorable respuesta cosmética y la reducida agresividad, hacen de PDT el método de elección en el tratamiento de esta patología. El prototipo utilizado en este estudio demostró ser además de no invasivo y bien tolerado, altamente efectivo. (AU)

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delta-Aminolevulinic acid (ALA), a heme precursor accumulated in acute intermittent porphyria and saturnism, undergoes autoxidation leading to ammonium ion and probably the corresponding alpha-ketoaldehyde. This reaction is accelerated by addition of oxyhemoglobin (oxyHb) and other iron complexes. OxyHb is concomitantly oxidized to metHb; the apparent second-order rate constant of oxyHb/ALA coupled oxidation is ca. 10 M-1 min-1.1H NMR and uv spectral studies suggest that ALA undergoes enolization before consuming the dissolved oxygen. Spin-trapping experiments demonstrate formation of both the hydroxyl radical and a substrate-derived carbon-centered radical during ALA oxidation. Generation of active oxygen species by ALA might be related to the neuropathy associated to some acquired and inherited porphyrinpathies.

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In order to study the porphyrinogenic ability of lindane in mammals, rats were treated with the pesticide suspended with the aid of Tween or dissolved in oil during about 3 months. The urinary excretion of porphyrins and its precursors: delta-aminolaevulinate (ALA) and porphobilinogen (PBG), as well as the faecal excretion of coproporphyrin (COPRO) and protoporphyrin (PROTO) was determined weekly. At the end of the treatment the hepatic activities of ALA Synthase (ALA-S), the first and rate limiting enzyme of haem pathway, and porphyrinogen carboxy-lyase (PCL), enzyme which sequentially decarboxylates uroporphyrinogen (8 COOH) to coproporphyrinogen (4 COOH), were assayed. Lindane moderately increased the urinary excretion of porphyrins and its precursors, being the former the mainly affected parameter. The faecal excretion of COPRO and PROTO was also increased. However, the hepatic activity of ALA-S was not altered. This would suggest that the regulatory haem pool was not affected. Nor was PCL activity altered in spite of being the key enzyme for the attack of other chlorinated compounds. Although hexachlorobenzene (HCB), a very well known porphyrinogenic drug, and lindane are chemically related and generate similar metabolites, the last one produces a small and qualitatively different alteration of haem biosynthesis. This may be related with the absence or scarce formation of the reactive metabolite that accounts for the porphyrinogenic ability of HCB.

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Saccharomyces cerevisiae mutants bearing mutations at the cyc4 locus are partially deficient in cytochrome synthesis. Although the mutation is not in the structural gene for delta-aminolevulinic acid (Alv) synthase, the mutants are deficient in Alv synthesis in vivo as indicated by abnormally low intracellular Alv concentrations. The cyc4 mutation causes cells to grow very slowly in minimal glucose medium, but not in yeast extract-peptone-glucose medium. A simple nutritional defect caused by the cyc4 mutation is not involved because cytochrome deficiency is enhanced by growing cyc4 cells in yeast extract-peptone medium. A regulatory role for CYC4 is indicated. Evidence for negative feed-back control of Alv synthase by heme is provided by the observation of enhanced intracellular Alv accumulation in yeast mutants partially deficient in decarboxylation of uroporphyrinogen and coproporphyrinogen, respectively.

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A patient who had hereditary tyrosinemia was observed during two illnesses to have characteristics of acute intermittent porphyria with associated hypertension. Metabolic studies revealed elevated levels of urinary aminolevulinic acid but normal levels of porphyrin metabolites associated with, and possibly explained by, decreased red blood cell activity of the zinc-dependent enzyme, aminolevulinic acid dehydratase. Zinc deficiency could not be directly associated with the diminished enzyme activity. The patient's hypertension appeared to be related to increased urinary excretion of catecholamines and to elevated renin activity in peripheral venous blood.

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