Macedonian Journal of Chemistry and Chemical Engineering, 2009
Spectrophotometric assays of total anthocyanins, total phenolics, total catechins, total flavonoi... more Spectrophotometric assays of total anthocyanins, total phenolics, total catechins, total flavonoids, color intesity and hue were performed on Merlot wines obtained with 3, 6 and 10 days of maceration, containing 30 and 70 ppm SO2. Changes of phenolic contents were observed during three stages of the wines: after maceration, after 6 and 16 months in order to check the effect of maceration time, SO2 and storage of the wines. Wines were stored at low and higher temperature to check also the influence of storage temperature on the studied parameters. It was found that maceration time influences the content of polyphenol compounds, observing increasing of their concentrations with increased maceration time, while lower contents were measured in the wines after 16 months of storage (3006, 1732 and 1602 mg/l total phenolics and 478, 188 and 98.5 mg/l total anthocyanins, after maceration, after 6 and 16 months of storage, respectively, in wine with 30 ppm SO2). SO2 had not a significant eff...
A simple, rapid and precise HPLC method has been developed for the assay of hydrochlorothiazide i... more A simple, rapid and precise HPLC method has been developed for the assay of hydrochlorothiazide in urine. The clean-up of the urine samples was carried out by solid-phase extraction using HLB cartridges. Extraction recovery was 94.00-100.28 %. HPLC separation was performed with isocratic elution on Hypersil BDS C18 column (100 x 4.0 mm I.D., 3 µm particle size) protected with appropriate guard column. The mobile phase was 18 % acetonitrile and 0.025 mol/L solution of KH2PO4, pH 4 at flow rate of 0.3 mL/min. Detection of the substances was performed at 220 nm. The calibration curves were linear in the range of 2-50 µg/mL. The developed method is validated by checking its accuracy, precision and stability. The detection limit is 2 µg/mL hydrochlorothiazide. The method is proved to be convenient for routine analysis of hydrochlorothiazide in urine.
ABSTRACT Permethrin is an insecticide with various uses because of its high bioactivity, stabilit... more ABSTRACT Permethrin is an insecticide with various uses because of its high bioactivity, stability, and low toxicity to mammals. It is usually present in preparations for applications in agronomy, textile industry, medicine, veterinary medicine and pest control in households. A simple high-performance liquid chromatographic (HPLC) method has been developed for its fast identification and quantification in raw material and in various formulations. Cis- and trans-isomers of the active substance have been separated and analyzed isocraticaly using a reversе-phase column. Identification was performed by comparing retention times of isomers to the ones of a standard substance, and the presence of permethrin was confirmed by characteristic ultraviolet (UV) spectra obtained by diode array detector (DAD). The method was validated and employed for permethrin analysis in samples of raw material and in various formulations (medical shampoo, veterinary powder, powder and gel for domestic use). K...
The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl ace... more The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl acetate and n-butanol) obtained from Teucrium species (T. chamaedrys, T. montanum, T. polium) were investigated in this work. Phytochemical screening of the plant extracts proved the presence of flavonoids: luteolin, apigenin and/or diosmetin. The chemical composition of extracts was evaluated by HPLC and spectrophotometry. Antioxidant activities of the extracts were evaluated using three complementary in vitro assays: inhibition of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical, inhibition of hydroxyl radicals and protection of beta-carotene-linoleic acid model system. In the first two assays, strong inhibitory activity was shown by T. montanum and T. chamaedrys extracts. In the beta-carotene-linoleic acid model system, extracts from T. polium showed remarkable activity. These findings demonstrated that Teucrium species possess free radical and hydroxyl radical scavenging activity as well as...
The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl ace... more The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl acetate and n-butanol) obtained from Teucrium species (T. chamaedrys, T. montanum, T. polium) were investigated in this work. Phytochemical screening of the plant extracts proved the presence of flavonoids luteolin, apigenin and/or diosmetin. The chemical composition of extracts was evaluated by HPLC and spectrophotometry. Antioxidant activities of the extracts were evaluated using three complementary in vitro assays: inhibition of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical, inhibition of hydroxyl radicals and protection of beta-carotene-linoleic acid model system. In the first two assays, strong inhibitory activity was shown by T. montanum and T. chamaedrys extracts. In the beta-carotene-linoleic acid model system, extracts from T. polium showed remarkable activity. These findings demonstrated that Teucrium species possess free radical and hydroxyl radical scavenging activity as well as ...
The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl ace... more The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl acetate and n-butanol) obtained from Teucrium species (T. chamaedrys, T. montanum, T. polium) were investigated in this work. Phytochemical screening of the plant extracts proved the presence of flavonoids luteolin, apigenin and/or diosmetin. The chemical composition of extracts was evaluated by HPLC and spectrophotometry. Antioxidant activities of the extracts were evaluated using three complementary in vitro assays: inhibition of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical, inhibition of hydroxyl radicals and protection of beta-carotene-linoleic acid model system. In the first two assays, strong inhibitory activity was shown by T. montanum and T. chamaedrys extracts. In the beta-carotene-linoleic acid model system, extracts from T. polium showed remarkable activity. These findings demonstrated that Teucrium species possess free radical and hydroxyl radical scavenging activity as well as ...
Your article is protected by copyright and all rights are held exclusively by Springer Science+Bu... more Your article is protected by copyright and all rights are held exclusively by Springer Science+Business Media, LLC. This e-offprint is for personal use only and shall not be selfarchived in electronic repositories. If you wish to self-archive your article, please use the accepted manuscript version for posting on your own website. You may further deposit the accepted manuscript version in any repository, provided it is only made publicly available 12 months after official publication or later and provided acknowledgement is given to the original source of publication and a link is inserted to the published article on Springer's website. The link must be accompanied by the following text: "The final publication is available at link.springer.com".
Hypericum perforatumL. is a medicinal plant considered as an important natural source of secondar... more Hypericum perforatumL. is a medicinal plant considered as an important natural source of secondary metabolites with a wide range of pharmacological attributes. Hairy roots (HR) were induced from root segments ofin vitrogrown seedlings fromH. perforatumafter cocultivation withAgrobacterium rhizogenesA4. Investigations have been made to study the production of phenolic compounds in dark-grown (HR1) and photoperiod-exposed (HR2) cultures. The chromatographic analysis of phenolic acids, flavonols, flavan-3-ols, and xanthones revealed marked differences between HR1 and HR2 cultures. The production of quinic acid, kaempferol, and seven identified xanthones was increased in HR2. Moreover, HR2 showed a capability forde novobiosynthesis of two phenolic acids (3-p-coumaroylquinic acid and 3-feruloylquinic acid), three flavonol glycosides (kaempferol hexoside, hyperoside, and quercetin acetylglycoside), and five xanthones (tetrahydroxy-one-methoxyxanthone, 1,3,5-trihydroxy-6-methoxyxanthone, 1...
Hypericum perforatum L. cell suspensions were evaluated for their growth, antioxidant activity, p... more Hypericum perforatum L. cell suspensions were evaluated for their growth, antioxidant activity, phenolic compound productions and xanthone profile after elicitation with Agrobacterium tumefaciens and Agrobacterium rhizogenes. Secondary metabolite production in H. perforatum elicited cells was enhanced without loss of biomass. A. tumefaciens elicited cells showed largely increased amounts of total phenolics and flavonoids, whereas total flavanol contents were slightly enhanced. The production of phenolic compounds was clearly less marked in A. rhizogenes treated cells. Antioxidant activity of elicited cells was remarkably elevated throughout the post-elicitation period. A significant correlation between antioxidant activity and phenolic production in elicited cells was found. Xanthone profile of H. perforatum cells was notably changed after bacterial elicitation. H. perforatum elicited cells yielded about 17-fold higher levels of xanthones compared to control cells. Among the twenty-one detected xanthones, five of them identified as 1,3,5,6-tetrahydroxyxanthone C-prenyl isomer, toxyloxanthone, 1,3,7-trihydroxy-6methoxy-8-prenyl xanthone, 1,3,6-trihydroxy-7-methoxy-8prenyl xanthone and 1,3,6,7-tetrahydroxyxanthone 2-prenyl xanthone were de novo synthesized in elicited cells. Altogether, these results indicated that H. perforatum cells elicited with Agrobacterium represent promising experimental system for enhanced production of xanthones. Keywords Agrobacterium rhizogenes Á Agrobacterium tumefaciens Á Antioxidant activity Á Hypericum perforatum Á Phenolic compounds Á Xanthones Abbreviations CE Catechin equivalents DAD Diode-array detection DPPH 1,1-Diphenyl-2-picrylhydrazyl DW Dry weight ESI Electrospray ionization GAE Gallic acid equivalents HPLC High-performance liquid chromatography MS Mass spectrometry TP Total phenolics TF Total flavonoids TFL Total flavanols 2,4-D 2,4-Dichlorophenoxyacetic acid BA N 6-benzyladenine Electronic supplementary material The online version of this article (
Vranec is one of the most important red grape varieties in Republic of Macedonia, grown in all vi... more Vranec is one of the most important red grape varieties in Republic of Macedonia, grown in all vineyards, mostly in the Tikveš wine region. In this study, Vranec wines produced with different maceration times (4, 7, 14 and 30 days) in presence of enzyme and oak chips during fermentation were studied in order to determine the influence of vinification conditions on the aroma profile. The volatile compounds were determined using headspace solid phase microextraction (HS-SPME) with a PDMS/Carboxen/DVB fibre, coupled with gas chromatography-mass spectrometry (GC-MS). In total 63 aroma compounds were detected revealing a complex aroma profile of Vranec wines composed of esters, alcohols, fatty acids, aldehydes, ketones and sulphur compounds. The content of aroma compounds was related mostly to maceration time, observing increased relative amount of alcohols, esters and fatty acids from the fourth to seventh day of maceration and the presence of oak chips during the fermentation enhanced their formation. The Student-Newman-Keuls test has been applied to ascertain possible significant differences between the studied wines, and principal component analysis has been employed, showing separation and grouping of the wines according to maceration time and oak chips treatment.
Wines and grapes contain a large array of phenolic compounds belonging to non-flavonoids and flav... more Wines and grapes contain a large array of phenolic compounds belonging to non-flavonoids and flavonoids. This study evaluates the polyphenolic contents of six commercial red and white Macedonian wines and four grape varieties. Spectrophotometric methods were applied for the determination of the total phenolics, the total flavonoids, the total anthocyanins and the total catechins. The efficiency of acetone/water (80/20) and methanol/water (80/20) solutions for the extraction of polyphenols from grape pulp, seeds and skins were compared. The best extraction efficiency was achieved using acetone/ water. The obtained results showed that Macedonian grapes are rich in polyphenols, whereby the highest concentration of total phenolics was found for Vranec grapes. The analyzed wines contained high contents of polyphenol; the highest contents were found for Disan wine produced from the Vranec variety of grapes (1515 mg/L total phenolics, 1103 mg/L total flavonoids, 237 mg/L total anthocyanins...
Coenzyme Q10 (CoQ10) is one of the essential components of the mitochondrial electron-transport c... more Coenzyme Q10 (CoQ10) is one of the essential components of the mitochondrial electron-transport chain (ETC) with the primary function to transfer electrons along and protons across the inner mitochondrial membrane (IMM). The concomitant proton gradient across the IMM is essential for the process of oxidative phosphorylation and consequently ATP production. Cytochrome P450 (CYP450) monoxygenase enzymes are known to induce structural changes in a variety of compounds and are expressed in the IMM. However, it is unknown if CYP450 interacts with CoQ10 and how such an interaction would affect mitochondrial function. Using voltammetry, UVÀvis spectrometry, electron paramagnetic resonance (EPR), nuclear magnetic resonance (NMR), fluorescence microscopy and high performance liquid chromatographyÀmass spectrometry (HPLCÀMS), we show that both CoQ10 and its analogue CoQ1, when exposed to CYP450 or alkaline media, undergo structural changes through a complex reaction pathway and form quinone structures with distinct properties. Hereby, one or both methoxy groups at positions 2 and 3 on the quinone ring are replaced by hydroxyl groups in a time-dependent manner. In comparison with the native forms, the electrochemically reduced forms of the new hydroxylated CoQs have higher antioxidative potential and are also now able to bind and transport Ca 2þ across artificial biomimetic membranes. Our results open new perspectives on the physiological importance of CoQ10 and its analogues, not only as electron and proton transporters, but also as potential regulators of mitochondrial Ca 2þ and redox homeostasis.
Journal of Biochemical and Biophysical Methods, 2008
A simple, rapid and precise HPLC method has been developed for the assay of verapamil in human pl... more A simple, rapid and precise HPLC method has been developed for the assay of verapamil in human plasma. The clean up of the plasma samples was tested using several adsorbents for solid-phase extraction and best recovery was obtained using mixed-mode cartridges (HLB-hydrophilic-lipophilic balance) ranging between 94.70 and 103.71%. HPLC separation was performed with isocratic elution on Lichrospher 60 RP-select B column (250 mm × 4 mm I.D., 5 μm particle size). The mobile phase was 40% acetonitrile and 0.025 mol/L KH 2 PO 4 with pH 2.5 at flow rate of 1 mL/min. Diltiazem was used as internal standard and the detection wavelength was 200 nm. The calibration curves were linear in the range of 10-500 ng/mL. The developed method is convenient for routine analysis of verapamil in human plasma.
The application of an ion trap mass spectrometer, usually employed for identification, has been h... more The application of an ion trap mass spectrometer, usually employed for identification, has been here systematically evaluated for quantitative analysis of various conjugated forms of flavonoids and compared with UV quantification. Three MS methods were tested to assess the potential and limits of the ion trap for quantification of flavonoids: full-scan experiment MS(2) , isolated ion experiment MS, and full-scan experiment MS. The test was performed using nine reference standards of flavonoids with six different aglycones: luteolin, apigenin, hypolaetin, 4'-O-methylhypolaetin, isoscutellarein and 4'-O-methylisoscutellarein in the form of 7-O-glucosides and diglucosides, mono or diacetylated, isolated from Sideritis scardica. The analytical characteristics of the tested MS methods were shown to be comparable to UV with regards to precision and accuracy, and superior for selectivity and sensitivity especially when using extracted ion chromatograms. Detection limits did not differ significantly between the MS methods but were significantly lower than those obtained with UV detection by one order of magnitude. Another issue addressed by these results was the choice of most suitable standard substances for quantification of flavonoids with various substituents attached when using MS. In UV detection, the nature of the aglycone is crucial for the absorbance properties, and various derivatives can be quantified with the available one with the same aglycone. Here, it was shown that in MS detection, one flavone derivative can be quantified using other available derivatives with similar substitution pattern with regards to attached and acetylated sugars, whereas the nature of the aglycone is not crucial.
Journal of Liquid Chromatography & Related Technologies, 2007
A non‐parameter method based on the assumption of additive substituent increments on retention is... more A non‐parameter method based on the assumption of additive substituent increments on retention is employed for estimation of the effects of the substituents in the flavone ring on the retention in the reversed phase HPLC. The influence of OH‐ and OCH3‐groups in the positions: 3, 5, 6, 7, 8, 3′, 4′, and 5′ on reversed phase HPLC retention is studied
Journal of Liquid Chromatography & Related Technologies, 2001
Assay of flavone aglycones in extracts of Origanum vulgare from Macedonia including identificatio... more Assay of flavone aglycones in extracts of Origanum vulgare from Macedonia including identification and isolation is performed. Hydrolyzed extracts were analyzed by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC). Three aglycones were identified and then isolated by column chromatography and TLC. Further TLC, HPLC, and UV/VIS spectrophotometric analysis showed that the isolated compounds are luteolin (the most
Your article is protected by copyright and all rights are held exclusively by Association of Food... more Your article is protected by copyright and all rights are held exclusively by Association of Food Scientists & Technologists (India). This e-offprint is for personal use only and shall not be self-archived in electronic repositories. If you wish to self-archive your work, please use the accepted author's version for posting to your own website or your institution's repository. You may further deposit the accepted author's version on a funder's repository at a funder's request, provided it is not made publicly available until 12 months after publication.
Macedonian Journal of Chemistry and Chemical Engineering, 2009
Spectrophotometric assays of total anthocyanins, total phenolics, total catechins, total flavonoi... more Spectrophotometric assays of total anthocyanins, total phenolics, total catechins, total flavonoids, color intesity and hue were performed on Merlot wines obtained with 3, 6 and 10 days of maceration, containing 30 and 70 ppm SO2. Changes of phenolic contents were observed during three stages of the wines: after maceration, after 6 and 16 months in order to check the effect of maceration time, SO2 and storage of the wines. Wines were stored at low and higher temperature to check also the influence of storage temperature on the studied parameters. It was found that maceration time influences the content of polyphenol compounds, observing increasing of their concentrations with increased maceration time, while lower contents were measured in the wines after 16 months of storage (3006, 1732 and 1602 mg/l total phenolics and 478, 188 and 98.5 mg/l total anthocyanins, after maceration, after 6 and 16 months of storage, respectively, in wine with 30 ppm SO2). SO2 had not a significant eff...
A simple, rapid and precise HPLC method has been developed for the assay of hydrochlorothiazide i... more A simple, rapid and precise HPLC method has been developed for the assay of hydrochlorothiazide in urine. The clean-up of the urine samples was carried out by solid-phase extraction using HLB cartridges. Extraction recovery was 94.00-100.28 %. HPLC separation was performed with isocratic elution on Hypersil BDS C18 column (100 x 4.0 mm I.D., 3 µm particle size) protected with appropriate guard column. The mobile phase was 18 % acetonitrile and 0.025 mol/L solution of KH2PO4, pH 4 at flow rate of 0.3 mL/min. Detection of the substances was performed at 220 nm. The calibration curves were linear in the range of 2-50 µg/mL. The developed method is validated by checking its accuracy, precision and stability. The detection limit is 2 µg/mL hydrochlorothiazide. The method is proved to be convenient for routine analysis of hydrochlorothiazide in urine.
ABSTRACT Permethrin is an insecticide with various uses because of its high bioactivity, stabilit... more ABSTRACT Permethrin is an insecticide with various uses because of its high bioactivity, stability, and low toxicity to mammals. It is usually present in preparations for applications in agronomy, textile industry, medicine, veterinary medicine and pest control in households. A simple high-performance liquid chromatographic (HPLC) method has been developed for its fast identification and quantification in raw material and in various formulations. Cis- and trans-isomers of the active substance have been separated and analyzed isocraticaly using a reversе-phase column. Identification was performed by comparing retention times of isomers to the ones of a standard substance, and the presence of permethrin was confirmed by characteristic ultraviolet (UV) spectra obtained by diode array detector (DAD). The method was validated and employed for permethrin analysis in samples of raw material and in various formulations (medical shampoo, veterinary powder, powder and gel for domestic use). K...
The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl ace... more The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl acetate and n-butanol) obtained from Teucrium species (T. chamaedrys, T. montanum, T. polium) were investigated in this work. Phytochemical screening of the plant extracts proved the presence of flavonoids: luteolin, apigenin and/or diosmetin. The chemical composition of extracts was evaluated by HPLC and spectrophotometry. Antioxidant activities of the extracts were evaluated using three complementary in vitro assays: inhibition of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical, inhibition of hydroxyl radicals and protection of beta-carotene-linoleic acid model system. In the first two assays, strong inhibitory activity was shown by T. montanum and T. chamaedrys extracts. In the beta-carotene-linoleic acid model system, extracts from T. polium showed remarkable activity. These findings demonstrated that Teucrium species possess free radical and hydroxyl radical scavenging activity as well as...
The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl ace... more The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl acetate and n-butanol) obtained from Teucrium species (T. chamaedrys, T. montanum, T. polium) were investigated in this work. Phytochemical screening of the plant extracts proved the presence of flavonoids luteolin, apigenin and/or diosmetin. The chemical composition of extracts was evaluated by HPLC and spectrophotometry. Antioxidant activities of the extracts were evaluated using three complementary in vitro assays: inhibition of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical, inhibition of hydroxyl radicals and protection of beta-carotene-linoleic acid model system. In the first two assays, strong inhibitory activity was shown by T. montanum and T. chamaedrys extracts. In the beta-carotene-linoleic acid model system, extracts from T. polium showed remarkable activity. These findings demonstrated that Teucrium species possess free radical and hydroxyl radical scavenging activity as well as ...
The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl ace... more The chemical composition and antioxidant activity of different extracts (diethyl ether, ethyl acetate and n-butanol) obtained from Teucrium species (T. chamaedrys, T. montanum, T. polium) were investigated in this work. Phytochemical screening of the plant extracts proved the presence of flavonoids luteolin, apigenin and/or diosmetin. The chemical composition of extracts was evaluated by HPLC and spectrophotometry. Antioxidant activities of the extracts were evaluated using three complementary in vitro assays: inhibition of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical, inhibition of hydroxyl radicals and protection of beta-carotene-linoleic acid model system. In the first two assays, strong inhibitory activity was shown by T. montanum and T. chamaedrys extracts. In the beta-carotene-linoleic acid model system, extracts from T. polium showed remarkable activity. These findings demonstrated that Teucrium species possess free radical and hydroxyl radical scavenging activity as well as ...
Your article is protected by copyright and all rights are held exclusively by Springer Science+Bu... more Your article is protected by copyright and all rights are held exclusively by Springer Science+Business Media, LLC. This e-offprint is for personal use only and shall not be selfarchived in electronic repositories. If you wish to self-archive your article, please use the accepted manuscript version for posting on your own website. You may further deposit the accepted manuscript version in any repository, provided it is only made publicly available 12 months after official publication or later and provided acknowledgement is given to the original source of publication and a link is inserted to the published article on Springer's website. The link must be accompanied by the following text: "The final publication is available at link.springer.com".
Hypericum perforatumL. is a medicinal plant considered as an important natural source of secondar... more Hypericum perforatumL. is a medicinal plant considered as an important natural source of secondary metabolites with a wide range of pharmacological attributes. Hairy roots (HR) were induced from root segments ofin vitrogrown seedlings fromH. perforatumafter cocultivation withAgrobacterium rhizogenesA4. Investigations have been made to study the production of phenolic compounds in dark-grown (HR1) and photoperiod-exposed (HR2) cultures. The chromatographic analysis of phenolic acids, flavonols, flavan-3-ols, and xanthones revealed marked differences between HR1 and HR2 cultures. The production of quinic acid, kaempferol, and seven identified xanthones was increased in HR2. Moreover, HR2 showed a capability forde novobiosynthesis of two phenolic acids (3-p-coumaroylquinic acid and 3-feruloylquinic acid), three flavonol glycosides (kaempferol hexoside, hyperoside, and quercetin acetylglycoside), and five xanthones (tetrahydroxy-one-methoxyxanthone, 1,3,5-trihydroxy-6-methoxyxanthone, 1...
Hypericum perforatum L. cell suspensions were evaluated for their growth, antioxidant activity, p... more Hypericum perforatum L. cell suspensions were evaluated for their growth, antioxidant activity, phenolic compound productions and xanthone profile after elicitation with Agrobacterium tumefaciens and Agrobacterium rhizogenes. Secondary metabolite production in H. perforatum elicited cells was enhanced without loss of biomass. A. tumefaciens elicited cells showed largely increased amounts of total phenolics and flavonoids, whereas total flavanol contents were slightly enhanced. The production of phenolic compounds was clearly less marked in A. rhizogenes treated cells. Antioxidant activity of elicited cells was remarkably elevated throughout the post-elicitation period. A significant correlation between antioxidant activity and phenolic production in elicited cells was found. Xanthone profile of H. perforatum cells was notably changed after bacterial elicitation. H. perforatum elicited cells yielded about 17-fold higher levels of xanthones compared to control cells. Among the twenty-one detected xanthones, five of them identified as 1,3,5,6-tetrahydroxyxanthone C-prenyl isomer, toxyloxanthone, 1,3,7-trihydroxy-6methoxy-8-prenyl xanthone, 1,3,6-trihydroxy-7-methoxy-8prenyl xanthone and 1,3,6,7-tetrahydroxyxanthone 2-prenyl xanthone were de novo synthesized in elicited cells. Altogether, these results indicated that H. perforatum cells elicited with Agrobacterium represent promising experimental system for enhanced production of xanthones. Keywords Agrobacterium rhizogenes Á Agrobacterium tumefaciens Á Antioxidant activity Á Hypericum perforatum Á Phenolic compounds Á Xanthones Abbreviations CE Catechin equivalents DAD Diode-array detection DPPH 1,1-Diphenyl-2-picrylhydrazyl DW Dry weight ESI Electrospray ionization GAE Gallic acid equivalents HPLC High-performance liquid chromatography MS Mass spectrometry TP Total phenolics TF Total flavonoids TFL Total flavanols 2,4-D 2,4-Dichlorophenoxyacetic acid BA N 6-benzyladenine Electronic supplementary material The online version of this article (
Vranec is one of the most important red grape varieties in Republic of Macedonia, grown in all vi... more Vranec is one of the most important red grape varieties in Republic of Macedonia, grown in all vineyards, mostly in the Tikveš wine region. In this study, Vranec wines produced with different maceration times (4, 7, 14 and 30 days) in presence of enzyme and oak chips during fermentation were studied in order to determine the influence of vinification conditions on the aroma profile. The volatile compounds were determined using headspace solid phase microextraction (HS-SPME) with a PDMS/Carboxen/DVB fibre, coupled with gas chromatography-mass spectrometry (GC-MS). In total 63 aroma compounds were detected revealing a complex aroma profile of Vranec wines composed of esters, alcohols, fatty acids, aldehydes, ketones and sulphur compounds. The content of aroma compounds was related mostly to maceration time, observing increased relative amount of alcohols, esters and fatty acids from the fourth to seventh day of maceration and the presence of oak chips during the fermentation enhanced their formation. The Student-Newman-Keuls test has been applied to ascertain possible significant differences between the studied wines, and principal component analysis has been employed, showing separation and grouping of the wines according to maceration time and oak chips treatment.
Wines and grapes contain a large array of phenolic compounds belonging to non-flavonoids and flav... more Wines and grapes contain a large array of phenolic compounds belonging to non-flavonoids and flavonoids. This study evaluates the polyphenolic contents of six commercial red and white Macedonian wines and four grape varieties. Spectrophotometric methods were applied for the determination of the total phenolics, the total flavonoids, the total anthocyanins and the total catechins. The efficiency of acetone/water (80/20) and methanol/water (80/20) solutions for the extraction of polyphenols from grape pulp, seeds and skins were compared. The best extraction efficiency was achieved using acetone/ water. The obtained results showed that Macedonian grapes are rich in polyphenols, whereby the highest concentration of total phenolics was found for Vranec grapes. The analyzed wines contained high contents of polyphenol; the highest contents were found for Disan wine produced from the Vranec variety of grapes (1515 mg/L total phenolics, 1103 mg/L total flavonoids, 237 mg/L total anthocyanins...
Coenzyme Q10 (CoQ10) is one of the essential components of the mitochondrial electron-transport c... more Coenzyme Q10 (CoQ10) is one of the essential components of the mitochondrial electron-transport chain (ETC) with the primary function to transfer electrons along and protons across the inner mitochondrial membrane (IMM). The concomitant proton gradient across the IMM is essential for the process of oxidative phosphorylation and consequently ATP production. Cytochrome P450 (CYP450) monoxygenase enzymes are known to induce structural changes in a variety of compounds and are expressed in the IMM. However, it is unknown if CYP450 interacts with CoQ10 and how such an interaction would affect mitochondrial function. Using voltammetry, UVÀvis spectrometry, electron paramagnetic resonance (EPR), nuclear magnetic resonance (NMR), fluorescence microscopy and high performance liquid chromatographyÀmass spectrometry (HPLCÀMS), we show that both CoQ10 and its analogue CoQ1, when exposed to CYP450 or alkaline media, undergo structural changes through a complex reaction pathway and form quinone structures with distinct properties. Hereby, one or both methoxy groups at positions 2 and 3 on the quinone ring are replaced by hydroxyl groups in a time-dependent manner. In comparison with the native forms, the electrochemically reduced forms of the new hydroxylated CoQs have higher antioxidative potential and are also now able to bind and transport Ca 2þ across artificial biomimetic membranes. Our results open new perspectives on the physiological importance of CoQ10 and its analogues, not only as electron and proton transporters, but also as potential regulators of mitochondrial Ca 2þ and redox homeostasis.
Journal of Biochemical and Biophysical Methods, 2008
A simple, rapid and precise HPLC method has been developed for the assay of verapamil in human pl... more A simple, rapid and precise HPLC method has been developed for the assay of verapamil in human plasma. The clean up of the plasma samples was tested using several adsorbents for solid-phase extraction and best recovery was obtained using mixed-mode cartridges (HLB-hydrophilic-lipophilic balance) ranging between 94.70 and 103.71%. HPLC separation was performed with isocratic elution on Lichrospher 60 RP-select B column (250 mm × 4 mm I.D., 5 μm particle size). The mobile phase was 40% acetonitrile and 0.025 mol/L KH 2 PO 4 with pH 2.5 at flow rate of 1 mL/min. Diltiazem was used as internal standard and the detection wavelength was 200 nm. The calibration curves were linear in the range of 10-500 ng/mL. The developed method is convenient for routine analysis of verapamil in human plasma.
The application of an ion trap mass spectrometer, usually employed for identification, has been h... more The application of an ion trap mass spectrometer, usually employed for identification, has been here systematically evaluated for quantitative analysis of various conjugated forms of flavonoids and compared with UV quantification. Three MS methods were tested to assess the potential and limits of the ion trap for quantification of flavonoids: full-scan experiment MS(2) , isolated ion experiment MS, and full-scan experiment MS. The test was performed using nine reference standards of flavonoids with six different aglycones: luteolin, apigenin, hypolaetin, 4'-O-methylhypolaetin, isoscutellarein and 4'-O-methylisoscutellarein in the form of 7-O-glucosides and diglucosides, mono or diacetylated, isolated from Sideritis scardica. The analytical characteristics of the tested MS methods were shown to be comparable to UV with regards to precision and accuracy, and superior for selectivity and sensitivity especially when using extracted ion chromatograms. Detection limits did not differ significantly between the MS methods but were significantly lower than those obtained with UV detection by one order of magnitude. Another issue addressed by these results was the choice of most suitable standard substances for quantification of flavonoids with various substituents attached when using MS. In UV detection, the nature of the aglycone is crucial for the absorbance properties, and various derivatives can be quantified with the available one with the same aglycone. Here, it was shown that in MS detection, one flavone derivative can be quantified using other available derivatives with similar substitution pattern with regards to attached and acetylated sugars, whereas the nature of the aglycone is not crucial.
Journal of Liquid Chromatography & Related Technologies, 2007
A non‐parameter method based on the assumption of additive substituent increments on retention is... more A non‐parameter method based on the assumption of additive substituent increments on retention is employed for estimation of the effects of the substituents in the flavone ring on the retention in the reversed phase HPLC. The influence of OH‐ and OCH3‐groups in the positions: 3, 5, 6, 7, 8, 3′, 4′, and 5′ on reversed phase HPLC retention is studied
Journal of Liquid Chromatography & Related Technologies, 2001
Assay of flavone aglycones in extracts of Origanum vulgare from Macedonia including identificatio... more Assay of flavone aglycones in extracts of Origanum vulgare from Macedonia including identification and isolation is performed. Hydrolyzed extracts were analyzed by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC). Three aglycones were identified and then isolated by column chromatography and TLC. Further TLC, HPLC, and UV/VIS spectrophotometric analysis showed that the isolated compounds are luteolin (the most
Your article is protected by copyright and all rights are held exclusively by Association of Food... more Your article is protected by copyright and all rights are held exclusively by Association of Food Scientists & Technologists (India). This e-offprint is for personal use only and shall not be self-archived in electronic repositories. If you wish to self-archive your work, please use the accepted author's version for posting to your own website or your institution's repository. You may further deposit the accepted author's version on a funder's repository at a funder's request, provided it is not made publicly available until 12 months after publication.
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Papers by Marina Stefova