Development of novel optical pH sensors based on coumarin 6 and nile blue A encapsulated in resin particles and specific support materials.

Inspired by traditional pH papers, two types of ratiometric fluorescent pH sensors for neutral and alkaline pH ranges were developed in this study by using two fluorescent dyes, coumarin 6 (C6) and nile blue A (NB). These dyes were encapsulated into melamine-formaldehyde (MF) resin particles, which were then incorporated with polymer Nafion (Nf) or polyurethane hydrogel (PU) to prepare pH-sensing membranes. MF-C6 particles immobilized into polymer Nafion (i.e., MF-C6-Nf membrane) showed a dynamic ratiometric pH detection range of 4.5-7.5 through shift in fluorescence emission spectra at acidic and neutral pH solutions, as well as distinct color transition under normal visual sense from pink to yellow color. By contrast, MF-NB particles immobilized into polyurethane hydrogel (i.e., MF-NB-PU membrane) displayed a dynamic ratiometric fluorescence detection range of pH 9 - pH 12 via change in ratiometric fluorescence intensity at two emission band edges. The membrane also showed a clear change from blue to purple color under sunlight at high pH values. These pH-sensing membranes also exhibited high sensitivity, good reversibility, and stability. They were then applied to measure pH values in real urine samples and fermentation media.

FTIR analysis of cellulose treated with sodium hydroxide and carbon dioxide.

Cellulose samples treated with sodium hydroxide (NaOH) and carbon dioxide in dimethylacetamide (DMAc) were analyzed by FTIR spectroscopy. Absorbance of hydrogen-bonded OH stretching was considerably decreased by the treatment of NaOH and carbon dioxide. The relative absorbance ratio (A(4000-2995)/A(993)) represented the decrease of absorbance as a criterion of hydrogen-bond intensity (HBI). The absorbance of the band at 1430cm(-1) due to a crystalline absorption was also decreased by NaOH treatment. The absorbance ratio of the bands at 1430 and 987-893cm(-1) (A(1430)/A(900)), adopted as crystallinity index (CI), was closely related to the portion of cellulose I structure. With the help of FTIR equipped with an on-line evacuation apparatus, broad OH bending due to bound water could be eliminated. FTIR spectra of the carbon dioxide-treated cellulose samples at 1700-1525cm(-1) were divided into some bands including 1663, 1635, 1616, and 1593cm(-1). The broad OH bending due to bound water at 1641-1645cm(-1) was resolved to two bands at 1663 and 1635cm(-1). As a trace of DMAc, the band at 1616cm(-1) is disappeared by washing for the cellulose treated with carbon dioxide (Cell 1-C and Cell 2/60-C). The decrease of HBI, the easy removal of DMAc, and the band at 1593cm(-1) supported the introduction of new chemical structure in cellulose. The bands shown at 1593 and 1470cm(-1) was assigned as hydrogen-bonded carbonyl stretching and O-C-O stretching of the carbonate ion.

Crystalline structure analysis of cellulose treated with sodium hydroxide and carbon dioxide by means of X-ray diffraction and FTIR spectroscopy.

Crystalline structures of cellulose (named as Cell 1), NaOH-treated cellulose (Cell 2), and subsequent CO2-treated cellulose (Cell 2-C) were analyzed by wide-angle X-ray diffraction and FTIR spectroscopy. Transformation from cellulose I to cellulose II was observed by X-ray diffraction for Cell 2 treated with 15-20 wt% NaOH. Subsequent treatment with CO2 also transformed the Cell 2-C treated with 5-10 wt% NaOH. Many of the FTIR bands including 2901, 1431, 1282, 1236, 1202, 1165, 1032, and 897 cm(-1) were shifted to higher wave number (by 2-13 cm(-1)). However, the bands at 3352, 1373, and 983 cm(-1) were shifted to lower wave number (by 3-95 cm(-1)). In contrast to the bands at 1337, 1114, and 1058 cm(-1), the absorbances measured at 1263, 993, 897, and 668 cm(-1) were increased. The FTIR spectra of hydrogen-bonded OH stretching vibrations at around 3352 cm(-1) were resolved into three bands for cellulose I and four bands for cellulose II, assuming that all the vibration modes follow Gaussian distribution. The bands of 1 (3518 cm(-1)), 2 (3349 cm(-1)), and 3 (3195 cm(-1)) were related to the sum of valence vibration of an H-bonded OH group and an intramolecular hydrogen bond of 2-OH ...O-6, intramolecular hydrogen bond of 3-OH...O-5 and the intermolecular hydrogen bond of 6-O...HO-3', respectively. Compared with the bands of cellulose I, a new band of 4 (3115 cm(-1)) related to intermolecular hydrogen bond of 2-OH...O-2' and/or intermolecular hydrogen bond of 6-OH...O-2' in cellulose II appeared. The crystallinity index (CI) was obtained by X-ray diffraction [CI(XD)] and FTIR spectroscopy [CI(IR)]. Including absorbance ratios such as A1431,1419/A897,894 and A1263/A1202,1200, the CI(IR) was evaluated by the absorbance ratios using all the characteristic absorbances of cellulose. The CI(XD) was calculated by the method of Jayme and Knolle. In addition, X-ray diffraction curves, with and without amorphous halo correction, were resolved into portions of cellulose I and cellulose II lattice. From the ratio of the peak area, that is, peak area of cellulose I (or cellulose II)/total peak area, CI(XD) were divided into CI(XD-CI) for cellulose I and CI(XD-CII) for cellulose II. The correlation between CI(XD-CI) (or CI(XD-CII)) and CI(IR) was evaluated, and the bands at 2901 (2802), 1373 (1376), 897 (894), 1263, 668 cm(-1) were good for the internal standard (or denominator) of CI(IR), which increased the correlation coefficient. Both fraction of the absorbances showing peak shift were assigned as the alternate components of CI(IR). The crystallite size was decreased to constant value for Cell 2 treated at >or= 15 wt% NaOH. The crystallite size of Cell 2-C (cellulose II) was smaller than that of Cell 2 (cellulose I) treated at 5-10 wt% NaOH. But the crystallite size of Cell 2-C (cellulose II) was larger than that of Cell 2 (cellulose II) treated at 15-20 wt% NaOH.

Production of natural indirubin from indican using non-recombinant Escherichia coli.

Indirubin is an important natural substance and has positive effects on various diseases. However, the current process of producing indirubin is inefficient, making it difficult to produce indirubin of high purity; thus, it is commercially unavailable. In this study, a method of indirubin using non-recombinant Escherichia coli as a whole cell enzyme with indican as a substrate was developed. After confirming that indirubin was produced from indican by non-recombinant E. coli under general conditions, attempts to compare the yield and purity of indirubin were conducted under various pH, temperature and culturing media conditions. Under the optimum conditions, the yield was reliably determined to be about 25-35%, and it was further increased (1.8-2.1 fold) by replenishing the catalyst with freshly prepared whole cells. Since the established method was simple and reproducible, high purity indirubin would expected to be produced efficiently through improvement of whole cell enzymes and development of scale-up processes.