Spectrofluorimetric and HPLC Determination of Morin in Human Serum

Morin is a flavonol antioxidant. In ethanol-water mixtures (70 wt% of ethanol) it reacts with Al3+ to give Al(Morin)(2) in the pH range 3-6. The conditional stability constant of this complex at 298 K was found to be log beta(2) = 16.96 +/- 0.02 at pH 4.40. The complex shows strong fluorescence emission at 500 nm upon excitation at 410 nm. The fluorescence intensity is pH dependent with maximum emission at pH 4.40. Since the complexation reaction enhances the fluorescence of morin, this property was used for the determination of morin in human serum. A linear dependence of the intensity of fluorescence of the complex on the concentration of morin was obtained in morin concentration range from 1.5-30.5 ng mL(-1), relative standard error of measurements was 1.4%. The LOD was 0.02 ng mL(-1) while LOQ was 1.0 ng mL(-1). Serum concentration of morin was also determined using HPLC as a reference method. A C-18 Hypersil Gold AQ column was used with acetonitrile-0.1% v/v phosphoric acid (30:70% v/v) as the mobile phase at 1.0 mL min(-1) flow rate and UV detection at 250 nm. Acceptable relative standard errors (less than 5%) between determinations obtained by the two methods indicate that the fluorescence method is reliable

COMPARISON OF DIFFERENT MATHEMATICAL MODELS IN DECONVOLUTION OF LIGNIN FLUORESCENCE SPECTRA

We analysed fluorescence spectra of poplar lignin and lignin model compound, using gaussian, log-normal and exponential power model. Asymmetric models show wavelength positions of components in a complex molecule. Gaussian model indicates only discrete origin of the emission, but fails in defining component positions

DETERMINATION OF COUMAPHOS RESIDUES IN HONEY AFTER VARROA TREATMENT USING FLUORESCENCE SPECTROSCOPY

Varroa infestation is considered a major threat to honeybees due to its strong impact on the colony, attacking larval cells, increasing bee mortality, and serving as a vector of pathogens. Coumaphos is an organophosphate-based acaricide insecticide which is frequently used to control Varroa (Varroa destructor) mites in the honey bee (Apis melifera) colonies. The accumulation of these lipophilic contaminants in honey and beeswax may affect honey bee health, mainly honey bee larvae which are the most sensitive, and negatively affect queen quality. In this study, we analyzed honey samples from hives of honey bees that were treated with Coumaphos, as well as the samples of pure Coumafos. The front-face fluorescence measurements of analyed honey samples were recorded using an Fl3-221 P spectrofluorimeter, with a 450 W Xenon lamp. Obtained results showed an emission peak with maximum around 400 nm, after excitation at 340 nm. It could be related to the presence of Coumaphos in honey samples, which showed the characteristic position of emission maximum at the same wavelength. These results showed that fluorescence spectroscopy may be useful as a fast and sensitive method, for the efficient and specific determination of toxic substances such as Coumaphos residues in honey. Due to the proven toxic effects on honey bees, as well as on humans, monitoring hazardous contaminants in honey is of crucial importance in the field of food and agriculture

COMPARISON OF CELL WALL STRUCTURE OF DIFFERENT WESTERN BALKAN PLANT SPECIES AS A SOURCE FOR BIOFUELS

Understanding of composition and connections between the building macromolecules of plant biomass, such as cellulose, hemicellulose and lignin, is main key for their better utilization in biofuels industry. We compared four different plant species which are abundant in the region of the Western Balkans. We investigated the structure of the cell walls, as the main constituent of plant biomass, isolated from branches of softwood (Picea omorika (Pancic) Purkine), hardwood (Acer platanoides L.), maize stem (Zea mays L.) as examples of crop species, and Paulownia tomentosa tree as a fast-growing species with a huge biomass yield. For our investigation, we combined Fluorescence-detected linear dichroism (FDLD) method and X-ray Diffraction. We obtained data for anisotropy and crystallography which are a base for prediction of the best and appropriate plant species for easy deconstruction of its biomass. Our results show that Acer branch as a hardwood shows the highest anisotropy and the lowest crystallinity compared to the other species while Picea Omorika needles show opposite results as the lowest anisotropy and the higher crystallinity. The results for maize show that the stems are easier for utilization than leaves. The isolated cell walls from leaves of Paulownia tomentosa show similar results and good correlation between anisotropy and crystallinity, thus we can conclude that this plant is easy to use in biofuel industries

ORIENTATION OF CELL WALL POLYMERS IN THE Arabidopsis thaliana STEM

Mechanical and physical propreties of plant fibres are dependent on the orientation of constituent polymers (cellulose, hemicellulose, lignin). Fourier transform infrared (FTIR) microscopy was used to examine the orientation of the main plant polymers in transversal and longitudinal direction of the isolated cell wall of the Arabidopsis thaliana stem. The polarised FTIR measurements indicated that xylan and glucomannan have parallel orientation with regard to the orientation of cellulose, as well as lignin

The use of fluorescence microscopy for classification of pollen grains

In this pilot research, we showed that pollen autofluorescence varied between the pollen of the different botanical species. Our findings suggest that classification of pollen grains may be obtained based on their fluorescence images

BENEFICIAL EFFECTS OF UV-A RADIATION ON MUNG BEAN (VIGNA RADIATA L.) SEEDS

Mung bean (Vigna Radiata L.) seeds are an important source of both nutrients (such as proteins, fibers, vitamins) and a variety of bioactive compounds (like phenolic compounds). Ultraviolet (UV) light has an important function as a major environmental signal important for plant growth and development, but at the same time it may cause certain damaging effects on macromolecules and other cellular components. Plants respond to the production of reactive oxygen species (ROS), caused by UV irradiation, by activating changes in morphology, physiology, or production of secondary metabolites. Some of these mechanisms increase antioxidant capacity in order to reduce the harmful effect of produced ROS. In the present research, the duration of exposure to UV-A irradiation and its influence on antioxidant activity of mung bean seeds were studied. Seeds were exposed to constant irradiation for 1 or 3 hours using a UV-A lamp (93μW/cm2) at a 50 cm distance. Antioxidant activity was tested using DPPH (2,2-Diphenyl-1-picrylhydrazyl) assay. The results indicate a significant increase in antioxidant activity of the UV-A irradiated seeds after 1 hour (74,45 % ± 0,40), compared to the control (72,85 % ± 1,55). After prolonged exposure, measured antioxidant activity significantly increased (77,99 % ± 0,71). Our results show a correlation between duration of UV-A irradiation and the increase in antioxidant activity. This could be beneficial in agriculture for producing fortified food

Fluorescence Spectroscopy in Structural Studies of Plant Cell Walls

Plant cell walls represent the most abundant, renewable and biodegradable composite on Earth. Its highly complex structure consists mainly of three organic compounds: cellulose, hemicelluloses, and lignin. Cell walls have wide applications in different industries, especially for biofuels and biomaterials. Fluorescence spectroscopy is the method allowing investigation of cell wall structure thought monitoring of lignin autoflorescence and thus interactions of lignin with the other cell wall constituents. Deconvolution of fluorescence spectra reveals the number and location of spectral component peaks by calculation of the approximation of the probability density (APD) of component positions. A characteristic of complex CW fluorescence is that the emission spectrum contains multiple log–normal components originating from different fluorophores, shorter wavelengths corresponding to phenolic structures and longer wavelengths to conjugated structures in lignin. Fluorescence spectroscopy has been used for fast screening of the cell wall properties from plants of different origin (hardwood, softwood and herbaceous plant), that may be important for selection of plants for possible applications. Fluorescence spectroscopy may be applicable in the investigation of the effect of stress on the cell wall. Lignin fluorescence emission spectra, peak intensities and shifts in the positions of the long-wavelength spectral components may be indicators of changes in cell wall structure during the stress. There is an increasing application of quantum dots (QDs) in plant science, as fluorescent markers. The isolated cell wall is an appropriate object for study of the interactions with nanoparticles. The results of different physico-chemical techniques including fluorescence spectroscopy combined with spectral deconvolution, show that in the cell walls, CdSe QDs predominantly bind to cellulose, via OH groups, and to lignin, via the conjugated C=C/C–C chains. Variability of bond types in lignin is related to the involvement of this polymer in plant response to various types of stress, by introducing local structural modifications in the cell wall. Different lignin model compounds have been used in order to reveal spectroscopic properties of lignin. Lignin model polymers were synthesized from three monomers, coniferyl alcohol, ferulic acid and p-coumaric acid mixed in various ratios, simulating lignin synthesis in the real cell walls. Further, by using fluorescence spectroscopy and appropriate mathematical methods, it is possible to get deeper insight into the structural characteristics of the molecule. Future investigations will be based on synthetic cell walls and on variation in a portion of all three main components: cellulose, hemicelluloses and lignin, also having in mind results of fine structural modifications in lignin model compounds

Primena fluorescentne spektroskopije u kombinaciji sa metodom dekonvolucije u analizi semena kukuruza (Zea mays L.) kontaminiranih aflatoksinom

Semena kukuruza (Zea mais L.) mogu biti kontaminirana sa jednim od najpasnijih prirodnih kancerogenih kontaminanata kao što su aflatoksini. U ovoj studiji, proučavan je uticaj aflateritoksina na strukturne karakteristike strukture. Korišćena je fluorescentna spektropija u kombinaciji sa matematičkom analizom spektra. Primenjen je matematički model za analizu spektra asimetričnog modela (Log-normal) koji koristi Nelder-Mead algoritam (Matlab 6.5). Rezultati razlaganja fluorescentnih spektara pokazuju četiri glavne fluorescentne fluorescentne emisije sa 054 emisiona približna 050 komponenta. Ovaj rezultat može da se koristi za praćenje promene u semenu usled prisustva aflatoksina. Glavni pokazatelj uticaja aflatoksina na strukturu kukuružnog semena je pomeraj dugo-talasne emisione komponente (520 nm)

Determination of Flavonoids and Total Polyphenol Contents in Commercial Apple Juices

We propose a sensitive and selective spectrofluorimetric method for the determination of flavonoids as expressed in 'quercetin equivalent' in apple juices. The method is based on the strong emission of the aluminium(III)-quercetin complex at 480 nm with excitation at 420 nm, and it is successfully applied for the determination of flavonoids in commercial apple juices and compared with results obtained in reference spectrophotometric determination. The flavonoid content in commercial apple juices was found to range from 5.53 to 15.55 mg/l quercetin equivalent. The very good agreement between the two methods indicates the suitability of the proposed spectrofluorimetric method for the precise and accurate determination of flavonoids. In addition, the total polyphenol content was determined spectrophotometrically using the Folin-Ciocalteu (FC) method and the antioxidative activity of the tested juices was tested in a DPPH assay and these values were correlated with each other. The obtained profiles of compounds with antioxidative ability lead us to conclude that fruit juice labels based only on fruit % might sometimes misinform consumers