Ultrafiltration of Thin Stillage from Conventional and E-Mill Dry Grind Processes

We used ultrafiltration (UF) to evaluate membrane filtration characteristics of thin stillage and determine solids and nutrient compositions of filtered streams. To obtain thin stillage, corn was fermented using laboratory methods. UF experiments were conducted in batch mode under constant temperature and flow rate conditions. Two regenerated cellulose membranes (10 and 100 kDa molecular weight cutoffs) were evaluated with the objective of retaining solids as well as maximizing permeate flux. Optimum pressures for 10 and 100 kDa membranes were 207 and 69 kPa, respectively. Total solids, ash, and neutral detergent fiber contents of input TS streams of dry grind and E-Mill processes were similar; however, fat and protein contents were different (p < 0.05). Retentate obtained from conventional thin stillage fractionation had higher mean total solids contents (27.6% to 27.8%) compared to E-Mill (22.2% to 23.4%). Total solids in retentate streams were found similar to those from commercial evaporators used in industry (25% to 35% total solids). Fat contents of retentate streams ranged from 16.3% to 17.5% for the conventional process. A 2% increment in fat concentration was observed in the E-Mill retentate stream. Thin stillage ash content was reduced 60% in retentate streams.     

Synthesis and spectroscopic characterisation of BODIPY based fluorescent off-on indicators with low affinity for calcium

Two fluorescent off-on Ca2+ indicators based on APTRA (o-aminophenol-N,N,O-triacetic acid) as low-affinity ligand for Ca2+ and BODIPY(4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) as a fluorophore were synthesized. The new BODIPY-APTRA compounds absorb in the visible spectrum, with absorption maxima from 505 nm to 570 nm, and have fluorescence spectra that span the visible spectrum, with emission maxima ranging from 525 nm to 625 nm dependent on the substituents at the alpha-positions to the nitrogen atoms. The indicators show a large increase of the fluorescence quantum yield upon increasing Ca2+ concentration. The ground-state dissociation constants Kd estimated at 20 degrees C in 100 mM KCl aqueous buffered solution, pH 7.20, for the two complexes with Ca2+ were found to be around 100 microM.     

BODIPY-based hydroxyaryl derivatives as fluorescent pH probes

[structures: see text] Seven new 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) dyes with phenolic or naphtholic subunits on position 8 and with substituents having different electron driving forces on positions 3 and 5 were synthesized. Their absorption and steady-state fluorescence properties were investigated as a function of solvent. The novel compounds, with the exception of 4,4-difluoro-8-(4-hydroxyphenyl)-3,5-bis-(4-methoxyphenyl)-4-bora-3a,4a-diaza-s-indacene, are characterized by absorption maxima in the range 493-515 nm and small (400-600 cm(-1)) Stokes shifts. The exceptional dye has absorption maxima in the 570-580 nm region and fluorescence emission maxima around 610-620 nm, depending on the solvent. In aqueous solution, the dyes show a large fluorescent enhancement upon increasing the acidity of the solution. They can be used in aqueous solution as fluorescent pH probes excitable with visible light, with pKa values ranging from 7.5 to 9.3, depending on the substitution pattern on positions 3, 5, and 8.     

Characterisation of a synthesised fluorescent ligand (4-acridinol-1-sulphonic acid) using nuclear magnetic resonance spectroscopy

The synthesis and complete characterisation of the fluorescent ligand, 4-acridinol-1-sulphonic acid (the acridine analogue of 8-quinolinol-5-sulfonic acid) is described. Using a judicious array of nuclear magnetic resonance spectroscopy experiments, the structural elucidation and full assignment of all proton and carbon chemical shifts were afforded. The 4-acridinol-1-sulphonic acid was found to behave in a similar manner to 8-quinolinol-5-sulphonic acid, forming fluorescent complexes with magnesium(II) and zinc(II). The uncorrected emission maxima for the metal-acridinol complexes were found to be at around 620 nm compared to 505 nm for the respective quinolinol complexes. Unfortunately, preliminary spectrofluorimetric analytical figures of merit revealed that the detection limits of the new acridinol metal complexes were one and a half orders of magnitude poorer than those attained with the corresponding quinolinol ligand. However, in contrast to 8-quinolinol-5-sulphonic acid, the 4-acridinol-1-sulphonic acid ligand showed considerable selectivity for magnesium(II) and zinc(II) over aluminium(III).     

新型5-[10-(9-羧基蒽基)]-间苯二甲酸的合成及荧光性质

以5-氨基-间苯二甲酸二甲酯为原料,经重氮化溴取代、Miyaura硼酸酯化反应生成3,5-二甲氧羰基苯硼酸频哪醇酯(3)。此外,以9-蒽甲酸为原料,经溴化、酯化反应生成10-溴-9-蒽甲酸甲酯(6)。最后以化合物3和6为原料,经Suzuki偶联、水解反应得到目标化合物5-[10-(9-羧基蒽基)]-间苯二甲酸。其结构经1H NMR、13C NMR和高分辨质谱表征。研究结果表明,该化合物钠盐水溶液具有发蓝光的性质,最大发射波长为425nm,同时也有较好的荧光量子效率。     

Improved determination of milk oligosaccharides using a single derivatization with anthranilic acid and separation by reversed-phase high-performance liquid chromatography

An improved analytical scheme for human milk neutral oligosaccharides determination was developed, in which, the oligosaccharides were pooled in two fractions (pools 1 and 2) after gel filtration, and then were quantitatively derivatized with a single fluorescent reagent, 2-anthranilic acid. Separation was by reversed-phase HPLC on an ODS-100Z column with a mobile phase of 50 mM ammonium acetate pH 4.0 and 150 mM citrate buffer pH 4.5 and monitored by a fluorescence detector at 360 nm excitation and 425 nm emission wavelengths. The method improved on the separation of neutral tetra- and hexa-saccharide isomers, namely, lacto-N-tetraose (LNT) and lacto-N-neotetraose (LNnT) as well as of lacto-N-difucohexaose I (LNDFH I) and lacto-N-difucohexaose II (LNDFH II). The separation of trisacccharide isomers, 3-fucosyllactose (3-FL) and 2′-fucosyllactose (2′-FL) was also successful. Limits of detection and quantification were in the range of 1–10 ng/l and 2–30 ng/l, respectively. The methods’ accuracy was good with its precision at <20% RSD and <1% RSD, respectively, for oligosaccharide concentration and retention time. The recoveries were in the range of 80–100%. This method was successfully applied to the separation and determination of representative neutral oligosaccharide contents in Samoa women milk.     

1,3,5,7-Tetramethyl-8-aminozide-difluoroboradiaza-s-indacene as a new fluorescent labeling reagent for the determination of aliphatic aldehydes in serum with high performance liquid chromatography

A BODIPY-based fluorescent derivatization reagent with a hydrazine moiety, 1,3,5,7-tetramethyl-8-aminozide-difluoroboradiaza-s-indacene (BODIPY-aminozide), has been designed for aldehyde labeling. An increased fluorescence quantum yield was observed from 0.38 to 0.94 in acetonitrile when it reacted with aldehydes. Twelve aliphatic aldehydes from formaldehyde to lauraldehyde were used to evaluate the analytical potential of this reagent by high performance liquid chromatography (HPLC) on C₁₈ column with fluorescence detection. The derivatization reaction of BODIPY-aminozide with aldehydes proceeded at 60 °C for 30 min to form stable corresponding BODIPY hydrazone derivatives in the presence of phosphoric acid as a catalyst. The maximum excitation (495 nm) and emission (505 nm) wavelengths were almost the same for all the aldehyde derivatives. A baseline separation of all the 12 aliphatic aldehydes (except formaldehyde and acetaldehyde) is achieved in 20 min with acetonitrile–tetrahydrofuran (THF)–water as mobile phase. The detection limits were obtained in the range from 0.43 to 0.69 nM (signal-to-noise = 3), which are better than or comparable with those obtained by the existing methods based on aldehyde labeling. This reagent has been applied to the precolumn derivatization followed with HPLC determination of trace aliphatic aldehydes in human serum samples without complex pretreatment or enrichment method.     

ESIPT inspired fluorescent 2-(4-benzo[d]oxazol-2-yl)naphtho[1,2-d]oxazol-2-yl)phenol: experimental and DFT based approach to photophysical properties

ESIPT inspired fluorescent 2-(4-benzo[d]oxazol-2-yl)naphtho[1,2-d]oxazol-2-yl)phenol was synthesized from 1-amino-3-(1,3-benzoxazol-2-yl)naphthalen-2-ol. Photophysical behavior of the synthesized compound was studied using UV–visible and fluorescence spectroscopy in polar and non-polar solvents. The synthesized naphthoxazolyl benzoxazole is fluorescent and very sensitive to the micro-environment. It shows a single absorption and dual emission in non-polar solvents with large Stokes shift originating from Excited State Intramolecular Proton Transfer while in polar solvents only a single short wavelength emission is observed. Experimental absorption and emission wavelengths are in good agreement with those predicted using the Time-Dependent Density Functional Theory (TD-DFT) [B3LYP/6-31G(d)]. The largest wavelength difference between the experimental and computed absorption maxima was 16 nm (acetonitrile) and 7 nm (ethyl acetate, THF, and 1,4-dioxane) in the short and long wavelength regions, respectively. A largest difference of 25 nm was observed for the short wavelength emission in DMF and 22 nm for the longer wavelength emission in chloroform.     

Syntheses of fluorescent dyes. XIV. Standards for fluorescence measurements in the near neutral pH-range

Highly fluorescent cationic indicators have been obtained by reaction of heterocyclic nitrogen bases with 1,3-propane sultone. Unlike the bases themselves, the fluorescence intensity of the cationic salts is practically pH-independent in the neutral pH-range, which makes them useful reference substances in measuring fluorescence intensities in physiological studies. Furthermore, they are sufficiently water soluble, the fluorescence maxima covering the wavelength range from 440 to 530 nm. 2-(3-Sulfonatopropyl)harmanium, ( 5 ) appears to be the most useful compound for intra- and extracellular liquids, since its fluorescence is not quenched by chloride ion.     

An improved analytical method for determination of heterocyclic amines in chicken legs

Summary Several extraction, separation and detection methods for heterocyclic amines (HAs) in chicken legs were evaluated by liquid chromatography. Results showed that the most appropriate extraction method includes the removal of macrosubstances by centrifugation and subsequent purification using a PRS (propylsulfonic acid silica gel) and a C₁₈ cartridge, and the recovery obtained ranged between 51 and 89 %. For HPLC separation, a binary solvent system consisting of acetonitrile and 0.05 M ammonium acetate solution (pH 3.6) with gradient elution with flow rate of 1.0 mL min⁻¹ and detection at 258 nm was used to resolve 16 HAs. With fluorescence nine HAs could be detected by employing a programmable wavelength, and the sensitivity was 100–400 times higher than that by UV detection. The detection limits for UV and fluorescence detection were 0.02≈0.5 ng and 0.05≈3 pg respectively, with a signal-to-noise ratio 3. The presence of HAs in fried chicken legs was also determined.     

Synthesis of water-soluble, ring-substituted squaraine dyes and their evaluation as fluorescent probes and labels

A series of ring-substituted squaraines absorbing and emitting in the red and NIR spectral region was synthesized and their spectral and photophysical properties (quantum yields, fluorescence lifetimes) and photostabilities were measured and compared to Cy5, a commonly used fluorescent label. The absorption maxima in aqueous media were found to be between 628 and 667 nm and the emission maxima are between 642 and 685 nm. Squaraine dyes exhibit high extinction coefficients (163,000–265,000 M⁻¹ cm⁻¹) and lower quantum yields (2–7%) in aqueous buffer but high quantum yields (up to 45%) and long fluorescence lifetimes (up to 3.3 ns) in presence of BSA. Dicyanomethylene- and thio-substituted squaraines exhibit an additional absorption around 400 nm with extinction coefficients between 21,500 and 44,500 M⁻¹ cm⁻¹. These dyes are excitable not only with red but also with blue diode lasers or light emitting diodes. Due to the favourable spectral and photophysical properties these dyes can be used as fluorescent probes and labels for intensity- and fluorescence lifetime-based biomedical applications.     

Synthesis and fluorescence properties of side-chain carboxylated 5,9-diaminobenzo[a]phenoxazinium salts

The efficient synthesis of a series of novel side-chain carboxylated 5,9-diaminobenzo[a]phenoxazinium salts is described. The ring system was prepared by the reaction of 5-alkylamino-2-nitrosophenol hydrochlorides with the appropriate N-alkylated-naphthylamine. Evaluation of the visible and fluorescence properties of the cationic dyes was carried out in ethanol and water at physiological pH. In both solvents they showed intense visible absorption maxima in the range 500-638 nm (ethanol) and 625-650 (water), and fluoresced strongly, with fluorescence maxima from 612 to 669 nm (ethanol) and from 654 to 685 nm (water). A wide variation in fluorescence quantum yields is observed, ranging from 0.051 to 0.50 and 0.065 to 0.32 in ethanol and water, respectively.     

Coumarinyl(thienyl)thiazoles as new fluorescent molecular photoswitches

A number of new photochromic 3-(4-phenylthiazol-5-yl)-and 3-(4-hetarylthiazol-5-yl)- coumarins has been synthesized. These compounds possess properties of molecular photo- switches providing a reversible change of the fluorescence intensity in the visible region of the spectrum upon alternating irradiation of their solutions with the visible and UV light. Irradiation with the UV light (λ < 400 nm) leads to their electrocyclization and loss of fluorescence, whereas irradiation of the cyclic form with the visible light (λ < 400 nm) returns the system to the state with the original absorption and fluorescence spectra. Switching of fluorescence is also observed in polymer matrices.     

Novel B,O-chelated fluorescent probe for nitric oxide imaging in Raw 264.7 macrophages and onion tissues

A novel fluorescent probe based on B,O-chelated dipyrromethene chromophore in far-visible and near-infrared spectral region (600–900 nm), boron chelated 8-(3,4-diaminophenyl)-3,5-bis(2-hydroxyphenyl)-4-bora-3a,4a-diaza-s-indancene (BOPB), has been first developed for nitric oxide (NO) imaging. BOPB, a turn-on fluorescent probe, can react with NO rapidly under physiological condition. The reaction product of BOPB with NO, BOPB-T, emits bright red fluorescence at 643 nm when excited at 622 nm. Meanwhile, BOPB-T displays high fluorescent quantum yield of 0.21 and good photostability. The selectivity for NO over other reactive oxygen/nitrogen species and ascorbic acid has been investigated and BOPB has good specificity for the detection of NO. MTT assay shows that the toxicity of BOPB (below 10 μM) to living cells can be neglected. Based on these investigations, BOPB has been used for NO imaging in Raw 264.7 cells and onion tissues. Meanwhile, mechanical injury to onion tissues results in a brighter fluorescence around the wound, which indicates that more NO has been produced in plant tissues in response to external stimuli. Our studies illustrate that BOPB has advantages of high sensitivity, low background interference and little photo damage on fluorescence imaging of NO.     

Fluorescence properties of indolenine semi-squarylium dyes

3-Butoxy-4-(1-butyl-3,3-dimethyl-3H-indol-2-ylidenemethyl)-3-cyclobut-1,2-dione exhibited the most intense florescence at fluorescence maxima 536 and 563 nm with fluorescence quantum yield 0.21 among any indolenine semi-squarylium dyes in the crystalline form due to the isolated dimer-type molecular packing and its suitable melting point. This compound showed aggregation-induced emission enhancement.     

Application of polyfluorophenazines to dichroic fluorescent dyes in guest-host liquid crystal displays

Novel dichroic fluorescent 2- and 2,7-disubstituted perfluorophenazines exhibit emission maxima in the range 570-600 nm in MLC-2039. The order parameter of fluorescence was observed in the range 0.29-0.49.     

Unnatural amino acid mutagenesis of green fluorescent protein

Unnatural amino acid mutagenesis has been used to selectively substitute tyrosine 66 of green fluorescent protein (GFP) with five novel amino acids: p-amino-L-phenylalanine, p-methoxy-L-phenylalanine, p-iodo-L-phenylalanine, p-bromo-L-phenylalanine, and L-3-(2-naphthyl)alanine. The absorbance and emission maxima of the resulting mutant GFPs span the range from 375 to 435 nm and 428 to 498 nm, respectively. The spectral properties of the mutant GFPs, including the absorbance and fluorescence maxima and quantum yields, correlate with the structural and electronic properties of the substituents on the amino acids.     

Some observations on the use of 1,4-dihydroxyanthraquinone as a fluorometric reagent for traces of lithium

A new fluorometric method for the determination of lithium based on the formation of a fluorescent species with 1,4-dihydroxyanthraquinone (Quinizarine), in acetone-water (90-10%) medium and 10⁻³M in NaOH is described. The fluorescent species has two excitation maxima at 560 and 596 nm and one emission maxima at 620 nm. It is stable at least 7 hr and the calibration graph is linear over the range 50–700 ppb Li. The method has a relative error over the average of measurements of 1.62%.     

Preparation of a fluorescent sensor based on BODIPY-functionalized hydroxyapatite nanoparticles and spectroscopic study of the Cd2+ and Zn2+ complex formation

A new fluorescent chemosensor 1, which based on hydroxyapatite (HA) nanoparticles covalently functionalized with a difluoroboron dipyrromethene, has been prepared by nucleophilic substitution of the fluorescent dye 3-chloro-4,4-difluoro-8-(4-tolyl)-5-[bis(pyridine2-ylmethyl)amino]-4-bora-3a,4a-diaza-s-indacene (2) with surface-modified HA nanoparticles. The HA particles were prepared by using SiO₂ as templates (THA) with 3-(aminopropyl)triethoxysilane (THA-APTES). Substitution of the electron-withdrawing chlorine in 2 by an electron-donating amino group of HA changes the properties of the nanoparticles 1 and the corresponding fluorescent dye 2. Absorption and emission maxima of 1 in ethanol are red-shifted by 75 and 30?nm, respectively, in comparison with those of 2. In contrast to no selectivity of dye 2 for Cd²⁺ or Zn²⁺ in EtOH/H₂O (95/5?V/V) solutions, the nanofluorescent probe 1 forms 1?:?1 complexes with Cd²⁺ or Zn²⁺, producing an instant color change along with large hypsochromic shifts in the absorption and fluorescence spectra by 70 and 35?nm, respectively, and large cation-induced fluorescence amplifications.     

Treatment of silk production wastewaters by membrane processes for sericin recovery

Sericin protein, although a valuable resource for many industries including cosmetics, pharmaceutical and biomedical, has been discarded as a waste in silk industry, causing environmental pollution. This paper describes determination of a membrane-based process for sericin recovery from cocoon cooking wastewaters (CCW) that will enable value-added utilization of waste sericin. The iso-electric point (pI) of sericin was found as 5–6, whose MW was distributed as 180–200, 70–80, 30–40 and 10–25 kDa. Prior to membrane filtration, sericin was separated from other impurities via centrifugation (CFG) followed by microfiltration (MF) in the pre-treatment stage, which also helped minimize post membrane fouling. Ultrafiltration (UF) and nanofiltration (NF) were adopted at a pH equal to pI of sericin. UF achieved partial recovery of sericin polypeptides at 37–60%, which was attributed to increased transmission of uncharged sericin polypeptides at their pI. On the other hand, NF achieved sericin recovery as high as 94–95%, containing all MW fractions. Severe flux decline was the major problem due to protein–membrane interactions and high sericin concentrations, where concentration polarization mainly had a dominant effect. Although flux declines were as high as 58–88% in UF and 70–75% in NF, flux recovery by at least 83% was achieved by chemical cleaning using NaOH and free chlorine.