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Analysis of alkyl gallates and nordihydroguaiaretic acid using plastic capillary electrophoresis - Microchips
Abstract
A simple and fast method for the simultaneous determination of the antioxidants propyl gallate (PG), octyl gallate (OG), lauryl gallate (LG), and nordihydroguaiaretic acid (NDGA) has been established by using microchip micellar electrokinetic chromatography with pulsed amperometric detection. Under the optimum conditions (30 mM borate buffer, pH 9.7, 30 mM sodium dodecyl sulfate, separation voltage of 1200 V and 5 s injection time) the analytes were baseline separated. Linear relationships were found between the concentration and peak current for all the selected antioxidants. The measured detection limits (S/N ≥ 3) of PG, OG, LG, and NDGA were 2.2, 1.4, 2.3, and 4.6 μM, respectively, which corresponds to 2–6 fmol of analyte. This approach has remarkable advantages with respect to other methodologies involving separations and electrochemical detection including minimal sample consumption, higher analysis speed, lower cost, and portability. Additionally, a highly reproducible signal (migration time and peak current) was obtained for a series of injections (n = 30). In order to demonstrate the capabilities of the method, the determination of antioxidants in a commercial food sample is also presented.
... It was also observed that if no preconcentration step is performed and sample volumes in the nL range are used, limits of detection in the low µM range can be obtained. [1][2][3] With only a few exceptions, 189,190 these values are of the same magnitude than previously reported for similar devices and analytes. 22,118,145,162,169,191 2. Electrophoretic separation using microchips. ...
... Sodium dodecyl sulfate (SDS) was used for the separation of a group of alkyl gallates and nordihydroguaiaretic acid. 2 In Figure 5, the dependence of the migration time of these phenolic antioxidants as a function of SDS concentration is shown. As can be observed, a clear decrease in the migration time (tM) was obtained for all the selected compounds upon the first addition of SDS (10 mM). ...
... 193 Figure 6b shows the electropherograms corresponding to a standard mixture of antioxidants and the analysis of a commercial food sample containing propyl gallate. 2 As can be observed, the baseline separation of the four selected antioxidants was achieved in less than 2 min. Using the optimized conditions, a gravy mix sample was also analyzed (b), and the corresponding amount of propyl gallate was calculated to be 45 mg of PG/kg of sample. ...
In this report, the most recent results regarding the use of microchip-capillary electrophoresis and pulsed electrochemical detection are reviewed. This article is particularly focused on the analysis of three groups of compounds: phenolic contaminants, phenolic acids, and phenolic antioxidants. Background information and a brief discussion covering other related analytical strategies are also included.
... 33 16,34 Tween 20, cetyltrimethylammonium bromide, 16 sodium dodecyl sulfate, 35 didodecyldimethylammonium bromide, 25,36,37 1,2-di-lauroylsn-phosphatidylcholine, Triton X-100, [38][39][40] and palmityl sulfobetaine 41 have all been shown to strongly influence both the m EOF and separation efficiency. 42 Different aspects of adsorption of surfactants to the capillary surfaces 16,30,[43][44][45][46][47][48][49][50][51][52][53][54][55][56] have been previously reported. It is generally accepted that surfactants can adsorb to the capillary wall by a combination of hydrophobic and electrostatic interactions. ...
... 52,62 The improvement in electrochemical signal is particularly significant when anionic surfactants are used in combination with pulsed amperometric detection (PAD). 30,[52][53][54]56,63,64 The main purpose of our work was to simultaneously study the electrophoretic and electrochemical effects of the addition of a series of alkyl sulfates to the background electrolyte. Experiments were conducted using a microchip-CE device and pulsed electrochemical detection. ...
... 66-70 A 25-mm gold wire was used as the working electrode, which has been previously described. 30,52,53,56,[66][67][68][69]71 The detection area was estimated to be 0.0039 mm 2 (25 mm electrode diameter, 50 mm channel width). Ag wire and Pt electrodes were used as pseudo-reference and auxiliary electrodes, respectively, and placed in the waste reservoir. ...
The effects of different surfactants (sodium 2-ethylhexyl sulfate, sodium decyl sulfate, sodium dodecyl sulfate and sodium tetradecyl sulfate) on the analysis of phenolic compounds by microchip-CE with pulsed amperometric detection were investigated. Using sodium decyl sulfate as a model surfactant, the effects of concentration and pH were examined. Under the optimized conditions, the analysis of six phenolic compounds was performed and compared with control runs performed without surfactant. When these surfactants were present in the run buffer, decreases in the migration time and increases in the run-to-run reproducibility were observed. Systematic improvements in the electrochemical response for the phenolic compounds were also obtained. According to the results presented, surfactants enhance the analyte-electrode interaction and facilitate the electron transfer process. These results should allow a more rational selection of the surfactants based on their electrophoretic and electrochemical effects.
... Owing to the advantageous features of microchip-CE technology, including negligible consumption of reagents and samples, and the capability for fast and automatized analysis insitu, the emergent development of microchips was already expected to see important applications in the food analysis field [1] as corroborated by the many recent applications of microchips CE in food analysis. These applications of microchips-CE include the analysis of DNA [64][65][66], proteins [110], amines [111], seleno-amino acids [112], antioxidants [113,114] and sulfite [115]. Besides, among the current developments in microchip-CE, different detection schemes have been developed to be used together with these microdevices. ...
... To avoid this, pulsed amperometric detection (PAD) has been proposed as a sensitive detection system suitable for the detection of a large number of analytes. Ding et al. [114] used this analytical strategy for the detection of antioxidants in 2 min above the low micromolar range. ...
This review covers the application of capillary electromigration methods to analyze foods and food components, including amino acids, biogenic amines, peptides, proteins, DNAs, carbohydrates, phenols, polyphenols, pigments, toxins, pesticides, vitamins, additives, small organic and inorganic ions, chiral compounds, and other compounds in foods, as well as those applications of CE for monitoring food interactions and food processing. The use of microchips as well as other foreseen trends in food analysis by CE are discussed. Papers that were published during the period June 2005-March 2007 are included following the previous review by Cifuentes (Electrophoresis 2006, 27, 283-303).
... In order to evaluate the performance of microchip, an initial analysis was performed using FA (106 mM in 5 mM phosphate buffer, 1 mM SDS, pH 12.0) [5,15,18,19,[45][46][47][48][49][50] as probe. After dispensing the sample, the test (injection, separation, and detection) was activated wirelessly. ...
We report the design of a digital system to wirelessly control microchip capillary electrophoresis (CE) equipment and a mobile unit for chemical analysis. The digital system consists of an embedded processor designed for digital control, decoding and applying of wirelessly-transmitted test parameters, data acquisition, and mobility control. The design is implemented on a field programmable gate array (FPGA) and its development board interfaces with four digital-to-analog converters on a newly-designed 3-channel high voltage power supply, electrochemical detector, wireless modems for communications with a base unit, mobile platform motor controllers, GPS sensor, and a micropump. The FPGA allows for all the interfacing hardware to perform CE and transmission of the data acquired from the interfacing electrochemical detector.
... Those studies also allowed to demonstrate that electrochemical detection can be effectively used for the analysis of phenolic compounds, avoiding electrode fouling [4][5][6][7]. Similar conditions were used to perform the analysis of other phenolic compounds including acids [8], antioxidants [9], and pharmaceuticals [10]. It is also worth highlighting that the use of carbon-ink electrodes (see Figure 1) enabled the analysis of catecholamines down to nM concentrations, without preconcentration, and using only 1.5 nL of sample [11]. ...
Recent developments in materials, surface modifications, separation schemes, detection systems and associated instrumentation have allowed significant advances in the performance of lab-on-a-chip devices. These devices, also referred to as micro total analysis systems (µTAS), offer great versatility, high throughput, short analysis time, low cost and, more importantly, performance that is comparable to standard bench-top instrumentation. To date, µTAS have demonstrated advantages in a significant number of fields including biochemical, pharmaceutical, military and environmental. Perhaps most importantly, µTAS represent excellent platforms to introduce students to microfabrication and nanotechnology, bridging chemistry with other fields, such as engineering and biology, enabling the integration of various skills and curricular concepts. Considering the advantages of the technology and the potential impact to society, our research program aims to address the need for simpler, more affordable, faster and portable devices to measure biologically active compounds. Specifically, the program is focused on the development and characterization of a series of novel strategies towards the realization of integrated microanalytical devices. One key aspect of our research projects is that the developed analytical strategies must be compatible with each other; therefore, enabling their use in integrated devices. The program combines spectroscopy, surface chemistry, capillary electrophoresis, electrochemical detection and nanomaterials. This article discusses some of the most recent results obtained in two main areas of emphasis: capillary electrophoresis, microchip-capillary electrophoresis, electrochemical detection and interaction of proteins with nanomaterials.
... A schematic drawing of the microchip is given in Figure 1. As previously reported, the detection electrode was then aligned at the end of the separation channel in an ad hoc perpendicular channel [29,31,41,42,44]. The two PDMS layers were then placed in an air plasma cleaner (Harrick, PDC-32G, Ithaca, NY), oxidized for 20 s and immediately brought into conformal contact to form an irreversible seal. ...
In this work, the development of a packed immobilized enzyme reactor (IMER) and its integration to a capillary electrophoresis microchip is described. The present microchip design differs from others, in the fact that the same design could be used with or without the particles and, just by changing the material used to pack the IMER, different analytes can be detected. The applied procedure involves the separation of the target analyte by capillary electrophoresis (CE), which is then coupled to a post-column IMER that produces H2O2. The H2O2 produced is finally detected downstream at the surface of a working electrode. Glucose was detected above 100 μM by packing particles modified with glucose oxidase at the end of the separation channel. The analytical performance of the microchip-CE has been demonstrated by performing the separation and detection of glucose and noradrenaline. Additions of fructose showed no effect on either the peak position or the peak magnitude of glucose. The microchip-CE-IMER was also used to quantify glucose in carbonated beverages with good agreement with other reports.
... They also developed analysis method for levoglucosan from smoke samples that can be used to fast locate forest fire or air pollution sources [115]. In the presence of 30 mM sodium dodecylsulfate surfactant, pulsed amperometric detection method successfully detect the antioxidants propyl gallate , octyl gallate, lauryl gallate (LG), and nordihydroguaiaretic acid (NDGA) [116] . Four synthetic antioxidants were analyzed within 2 min and the limit of detection was recorded 1- 4 m by microfluidic device–MEKC (Micellar electrokinetic chromatography) with pulsed amperometric detection mode. ...
Rashid, M. et. al. (2010) 'Recent developments in polymer microfluidic devices with capillary electrophoresis and electrochemical detection', Micro and Nanosystems, 2(2), pp.108-136. Abstract: We review recent work on the development of polymer microfluidic devices using capillary electrophoresis (CE) for fluid driving and separation and electrochemical detection (ED). A variety of commonly used off-chip and inte-grated electrochemical detection approaches, including amperometric, conductimetric and potentiometric detection ap-proaches, have been given. We describe fabrication approaches for the creation of microfluidic architecture in a polymer substrate and electrode integration. The methods for polymer surface modification to enhance the microfluidic device properties are also described as well as developments in power supply for the creation of a robust CE-ED microfluidic de-vice system. A variety of applications are described for the CE-ED systems.
... Details about the fabrication of PDMS microchannels can be found in the literature242526. Briefly, a master mold was produced using a 100 mm silicon wafer (Silicon; Santa Clara, CA, USA) and a 50 mm layer of SU-8 2035 photoresist (MicroChem, Newton, MA, USA). ...
This paper reports a simple procedure for coating fused-silica capillaries with poly(diallyldimethyl ammonium chloride) and montmorillonite. The coated capillaries were characterized by performing EOF measurements as a function of buffer pH, number of layers of coating, and number of runs (stability). The coated capillaries showed a highly stable μEOF (run-to-run RSD less than 1.5%, n = 20), allowing continuous use for several days without conditioning. The coated capillaries were then used for the effective separation of nine environmentally important phenolic compounds showing a significant improvement in the resolution, when compared to bare fused-silica capillaries. The EOF of the coated capillaries was constant in alkaline solutions (pH ≥ 7), allowing the optimization of the separation conditions of phenolic compounds without significantly affecting the μEOF.
... For instance, to extract BHA, BHT, PG, OG and DG in oil samples Sin et al. (2006) used n-hexane, AcN and a methanolic ascorbic acid solution while Aparicio et al. (2000) used only hexane and AcN for BHT and petroleum ether and ethanol for the others. The gallates were extracted with methanol and AcN with ammonium acetate solution from food samples by Ding, Mora, and Garcia (2006). ...
Antioxidants are an important group of food additives that have the ability to protect against detrimental change of oxidizable nutrients and consequently they extend shelf-life of foods.The present paper is an updated review on the analysis of ten antioxidants (both synthetic and natural). Three types of gallates, propyl (PG), octyl (OG) and dodecyl (DG), tert-Butylhydroquinone (TBHQ), Butylated Hydroxytoluene (BHT), tert-Butylhydroxyanisole (BHA), α-tocopherol, ascorbic acid, erythorbic acid and sodium d-isoascorbate were the antioxidants selected. The physico-chemical properties, sample preparation procedures and analytical methods for their determination are compared and discussed.
... MEKC can be advantageous over LC in terms of simplicity, resolution, and economy. Additionally, neutral analytes, which cannot be separated by capillary zone electrophoresis, are readily separated by MEKC (Ding, Mora, & Garcia, 2006;Hompesch, Garcia, Weiss, Vivanco, & Henry, 2005;Terabe, 2004). However, it typically suffers from low concentration sensitivity as a consequence of the limited sample volume and short path length for absorbance based detection (Molina & Silva, 2002). ...
A simple and fast method, based on the use of micellar electrokinetic capillary chromatography in combination with UV detection, was developed for the determination of Sudan dyes (I, II, III and IV). The separation of a mixture of the four standards was achieved using a background electrolyte consisting of 5 mM borate (pH 9.3), 20 mM sodium dodecyl sulfate and 20% acetonitrile. Under optimized conditions, the four azo-dyes were baseline separated in 20 min with limits of detection ranging from 96 to 610 μg/L (S/N > 3). The applicability of the method for rapid screening and determination of Sudan dyes (I, II and III) was studied by analyzing spiked chili powder samples from India, Pakistan, Mexico, United States, Canada, and China.
... In order to evaluate the performance of microchip, an initial analysis was performed using FA (106 mM in 5 mM phosphate buffer, 1 mM SDS, pH 12.0) [5,15,18,19,[45][46][47][48][49][50] as probe. After dispensing the sample, the test (injection, separation, and detection) was activated wirelessly. ...
In this paper, the fabrication of a wireless mobile unit containing an electrochemical detection module and a 3-channel high-voltage power supply (HVPS) designed for microchip CE is described. The presented device consists of wireless global positioning system controlled robotics, an electrochemical detector utilizing signal conditioning analog circuitry and a digital feedback range controller, a HVPS, an air pump, and a CE microchip. A graphical user interface (LabVIEW) was also designed to communicate wirelessly with the device, from a distant personal computer communication port. The entire device is integrated and controlled by digital hardware implemented on a field programmable gate array development board. This lab-on-a-robot is able to navigate to a global position location, acquire an air sample, perform the analysis (injection, separation, and detection), and send the data (electropherogram) to a remote station without exposing the analyst to the testing environment.
... 3 Capillary electrophoresis (CE) with high resolution and a small sample volume has been proved to be a powerful separation technique in the fields of analytical chemistry and bioanalytical chemistry. 10 CE 11,12 and microchip micellar electrokinetic chromatography 13 were developed for the resolution of other antioxidants in edible grain derivatives, other phenolic compounds in extra-virgin olive oil and PG, respectively. The amperometric detection technique is a popular method for the determination of various species with electrochemical activity. ...
Capillary electrophoretic separation coupled with end-column amperometric detection for the simultaneous quantification of butylated hydroxyanisole (BHA) and propyl gallate (PG) in food was developed. Important factors affecting separation and detection, such as the running buffer, separation voltage, and detection potential, were investigated in detail. An improved working electrode preparation method was used, where a carbon disk of 33 microm in diameter was sealed in a tip and positioned opposite the outlet of a capillary. The experiments indicated that the preparation method was simple, and the obtained electrode exhibited good flexibility and stability for the determination of phenolic antioxidants. The separation was carried out within 5 min using a 50 cm length capillary, with a solution containing 5 mM phosphate and 5 mM borax of pH 8.84 as a separation buffer, and a separation potential of 20 kV. Amperometric detection was achieved with an applied potential of 0.70 V versus Ag|AgCl| saturated KCl. There was excellent linearity between the peak current and the concentrations of the analytes in the range of 1.8 - 180.2 microg/mL for BHA and 10.6 - 212.2 microg/mL for PG, respectively. Relative standard deviations of 4.92% for BHA and 5.27% for PG were obtained, respectively. The developed method was successfully applied for the determination of antioxidants in several commercial foods.
... While determining and confirming trace quantities of 5 antioxidants in foods, Tsuji et al. (9) demonstrated that addition of ascorbic acid to the extraction solvent stabilized the target compounds, including NDGA. Use of microchip micellar electrokinetic chromatography with pulsed amperometric detection was recently reported (10). We have developed a single-laboratory validated method for the determination of NDGA in dietary supplement products. ...
Nordihydroguaiaretic acid (NDGA) occurs naturally in chaparral (Larrea tridentate Coville), a plant which commonly grows in the Southwest United States and has been used for medicinal purposes by Native Americans indigenous to that region. In addition to its traditional use as a tea, manufacturers of dietary supplements have marketed chaparral-containing products in a variety of formulations. Because of the hepatotoxicity of NDGA, and its occurrence in regulated products, we have developed a method for the determination of NDGA in dietary supplements and have tested this method in several dietary supplement formulations. Products were extracted with 80% methanol, filtered, and analyzed by high-performance liquid chromatography. NDGA was detected and determined with both a diode array detector and negative-ion electrospray. Fragmentation in the triple-quadrupole mass spectrometer was obtained by collisional activation of the [M-H](-) ion. Collisional activation produced sufficient fragmentation to provide unambiguous identification. Lack of a stable isotope labeled internal standard has led us to compare quantitations based on UV detection with quantitations based on tandem mass spectrometry (MS/MS). Presence of NDGA was confirmed in several dietary supplement products. Quantitative results from the 2 detection methods were comparable for most products. The limit of quantitation using MS/MS was lower and fewer interferences were observed, although UV detection provided better linearity.
This reference describes recent advances and applications of capillary electrophoresis in the field of food science. The first two chapters are devoted to the fundamentals of capillary electrophoresis, and to the main sample preparation techniques used for food analysis using this miniaturized separation technique, respectively. These two introductory chapters are followed by several chapters focused on the different strategies for analyzing specific food components, including lipids, carbohydrates, proteins, peptides, amino acids, vitamins, polyphenols, and food additives.
The information provided in these chapters helps readers to understand and develop appropriate methods to carry out a deep characterization of food samples. Relevant concepts such as food authentication, chemical food safety or the control of the quality and safety of dietary supplements, and food metabolomics are also covered, where appropriate. The big potential of capillary electrophoresis to achieve chiral separations and the determination of enantiomers in food samples or to develop targeted and non-targeted metabolomics strategies to ensure food safety and quality is also described. As an additional step towards analytical miniaturization, a chapter devoted to food analysis by microchip electrophoresis is also included in this book. All 14 chapters are contributed by highly experienced researchers in the field.
Capillary Electrophoresis in Food Analysis is a key source of information for food chemists and analytical chemists in industry (quality control laboratories) and academia (research labs and training courses).
The present study describes a simple strategy to integrate electrochemical detection with an assembled microchip-capillary electrophoresis platform. The electrochemical cell was integrated with a microfluidic device consisting of five plastic squares interconnected with fused silica capillaries, forming a four-way injection cross between the separation channel and three side-arms (each of 15 mm in length) acting as buffer/sample reservoirs. The performance of the system was evaluated using electrodes made with either carbon ink, carbon nanotubes, or gold and under different experimental conditions of pH, capillary length, and injection time. Using this system it was possible to separate the neurotransmitters dopamine and cathecol and to quantify phenol from a real sample using a linear calibration curve with a calculated LOD of 0.7 µM. A similar concept was applied to determine glucose, by including a pre-reactor filled with beads modified with glucose oxidase (GOx). The latter system was used to determine glucose in a commercial sample, with a recovery of 95.2 %. Overall, the presented approach represents a simple, inexpensive, and versatile approach to integrate electrochemical detection with CE separations without requiring access to microfabrication facilities.
Monitoring of synthetic phenolic antioxidants in food is very important due to their potential harmfulness to human beings. Octyl gallate (OG) is a commonly used phenolic antioxidant as a preservative in food stuff such as margarine, lard, sunflower oil etc. Herein, a glassy carbon electrode (GCE) chemically modified with gold nanoparticles (AuNPs) followed by self-assembled monolayer of dodecane thiol (DDT) was fabricated and employed for the determination of OG. An anodic peak corresponds to the oxidation of OG was observed at a potential of 0.22 V at DDT/AuNPs/GCE by square wave voltammetry. Based on this, a novel electrochemical sensor was developed for the determination of OG in oils and fats. Linear calibration graph was obtained within the range 1.2 to 0.2 μM. The detection limit was found to be 8.3 nM which is remarkably lower than those reported previously. The applicability of the method was demonstrated by analyzing OG in commercially available food samples such as margarine, butter and sunflower oil.
As described throughout this book, capillary electrophoresis (CE) has many advantages, but some shortcomings as well. The objective of this chapter is to describe the versatile roles of surfactants in solving problems encountered in CE. First, background information will be given for the structure and properties of surfactant monomers and their aggregates. Then, the use of surfactants as capillary wall coatings to increase separation efficiency, as a separation buffer modifier to tune selectivity, and as a pseudostationary phase to preconcentrate analytes to improve sensitivity and limits of detection will be described. Finally, the impact of surfactants on detection techniques coupled to CE will be discussed.
A simple and rapid capillary zone electrophoretic method is proposed for the analysis of antioxidants and preservatives in food. The important factors affecting separation and detection, for example pH, and concentration of the buffer electrolyte and organic modifier, were investigated in detail. Separation of five antioxidants (propyl gallate, gallic acid isoamyl ester, gallic acid n-octyl ester, nordihydroguaiaretic acid, and trihydroxybutyrophenone) and one preservative (benzoic acid) was achieved in a 50.5 cm (effective length) × 75 μm i.d fused-silica capillary, with 15 mmol L-1 borate buffer, pH 9.18, containing 25% (v/v) acetonitrile as separation buffer. UV detection was at 219 nm and the applied potential was 25 kV. Regression analysis revealed linear relationships between peak area and amount of each additive from 10 to 1000 μg mL-1 (R = 0.9992-1.0000). RSD of retention time and peak area were 0.44-0.74% and 1.25- 4.31%, respectively. The method was successfully used for simultaneous analysis of the six compounds in food with the recoveries from 89.3 to 115.8%.
The relevance of the development of microchip electrophoresis applications in the field of food analysis is considered in this work. A novel method to determine important phenolic compounds in extra virgin olive oil samples using a miniaturized chemical analysis system is presented in this paper. Three interesting phenolic compounds in olive oil and fruit (tyrosol, hydroxytyrosol and oleuropein glucoside) were studied by end-channel amperometric detection using a 100 μm gold wire as working electrode in glass microchip electrophoresis. The electrochemical behavior of these compounds was studied and the medium to carry out their detection was selected (0.1 M aqueous sulfuric acid). The best conditions for the separation were achieved in sodium tetraborate (10% methanol, pH 9.50) with different concentrations for the sample and the running buffer in order to allow the sample stacking phenomenon. The injection was carried out using 600 V for 3 s and the separation voltage was set at 1000 V. The quality of the method was evaluated through its analytical figures of merit and by its performance on real extra virgin olive oil samples. Determination of these compounds was carried out using the standard addition calibration method with good recoveries.
We report the design of a digital system to wirelessly control microchip capillary electrophoresis (CE) equipment and a mobile
unit for chemical analysis. The digital system consists of an embedded processor designed for digital control, decoding and
applying of wirelessly-transmitted test parameters, data acquisition, and mobility control. The design is implemented on a
field programmable gate array (FPGA) and its development board interfaces with four digital-to-analog converters on a newly-designed
3-channel high voltage power supply, electrochemical detector, wireless modems for communications with a base unit, mobile
platform motor controllers, GPS sensor, and an air micropump. The FPGA allows for all the interfacing hardware to perform
CE and transmission of the data acquired from the interfacing electrochemical detector. The work described herein extends
the utilization of microchip capillary electrophoresis to include remotely controlled field applications.
The addition of surfactants to the separation electrolyte is one of the most convenient ways to modify the electro‐osmotic flow (μEOF) in capillary electrophoresis. However, surfactants spontaneously adsorb to most surfaces; therefore, their presence in the running electrolyte may also affect the electrochemical detection. Changes in selectivity and sensitivity due to dynamic coating of electrode surfaces have been systematically reported during the last 30 years. In this review, some pertinent papers related to the use of surfactants to perform dynamic coatings of the capillary surface are discussed. The proposed mechanisms to explain the enhancements produced by surfactants at the detection step are also discussed along with some specific applications. A particular emphasis was placed on recent reports from our group dealing with the effects of anionic surfactants on the separation and detection of phenolic compounds.
A simple and rapid method has been developed for the analysis of four nonsteroidal anti-inflammatory drugs (NSAIDs) in serum using microchip capillary electrophoresis with pulsed amperometric detection. The selected NSAIDs (salicylic acid, acetaminophen, diflunisal, and diclofenac) are among the most commonly used drugs to treat fever, inflammation, and pain. Used above the therapeutic levels, these drugs can cause a wide variety of adverse effects and their fast analysis could have a significant impact in treatment and recovery of the patients. Several conditions, including separation potential, pH, and concentration of the electrolyte solution were studied to optimize the separation and detection. In this study, salicylic acid, acetaminophen, diflunisal, and diclofenac were separated in less than 2 minutes using a 5 mM borate buffer at pH 11.5 and a separation potential of +1200 V. Linear relationships were obtained between the concentration and peak current in the 0.5–15.3 μg/mL range and detection limits around 0.26 μg/mL. After 30 consecutive injections, the stability of both the response and migration time of the analytes showed relative related deviations of less than 4.6% and 1.0%, respectively. The potential of this method was verified by spiking a bovine serum sample with the four NSAIDs and analyzing the recovery ratio.
Antioxidants are one of the most common active ingredients of nutritionally functional foods which can play an important role in the prevention of oxidation and cellular damage inhibiting or delaying the oxidative processes. In recent years there has been an increased interest in the application of antioxidants to medical treatment as information is constantly gathered linking the development of human diseases to oxidative stress. Within antioxidants, phenolic molecules are an important category of compounds, commonly present in a wide variety of plant food materials. Their correct determination is pivotal nowadays and involves their extraction from the sample, analytical separation, identification, quantification and interpretation of the data. The aim of this review is to provide an overview about all the necessary steps of any analytical procedure to achieve the determination of phenolic compounds from plant matrices, paying particular attention to the application and potential of capillary electroseparation methods. Since it is quite complicated to establish a classification of plant food material, and to structure the current review, we will group the different matrices as follows: fruits, vegetables, herbs, spices and medicinal plants, beverages, vegetable oils, cereals, legumes and nuts and other matrices (including cocoa beans and bee products). At the end of the overview, we include two sections to explain the usefulness of the data about phenols provided by capillary electrophoresis and the newest trends.
Seven food grade antioxidants were resolved by microemulsion electrokinetic chromatography. The antioxidants included propyl gallate, octyl gallate, dodecyl gallate, butylated hydroxyanisole, butylated hydroxytoluene, tert-butylhydroquinone and alpha-tocopherol. They were resolved in 11 min using a low pH microemulsion containing 20% propan-2-ol. Theoretical plates in the range 39,000-95,000 per length of column (38.5 cm) were recorded for the antioxidants. The LODs were calculated to be in the range 9-44 microg/mL (30 s hydrostatic injection and a 50 microm id capillary column). The method can also be used to simultaneously analyse for the synthetic preservatives sorbic acid, benzoic acid, salicylic acid and the parabens. The method was successfully applied to noodles for the determination of butylated hydroxytoluene and propyl gallate and to pharmaceutical supplement tablets for alpha-tocopherol. The in-capillary concentration of butylated hydroxytoluene by sweeping was also demonstrated.
In this work, capillary electrophoresis chips with electrochemical detection have been assessed as creative and selective microfluidic platforms to integrate and simplify on a microscale the traditional methods for complex natural antioxidants determination. Depending on the acid-base properties of the analytes, two approaches (class-selective electrochemical index determination (CSEID) and individual antioxidant determination (IAD)) were investigated for the analysis of nine antioxidants ((+)-catechin, rutin, quercetin, chlorogenic, ferulic, caffeic, protocatechuic, vanillic, and gallic acids) in food samples. First, the novel concept of a class-selective electrochemical index is proposed allowing a fast and reliable determination of the main antioxidant classes (flavonoids and phenolic acids) in less than 100 s. In addition, an impressive separation of nine antioxidants is also offered in less than 260 s with the individual antioxidant determination approach. Qualitative and quantitative performances of both approaches were studied. The analytical figures of merit (i.e., electroosmotic flow (EOF) precision as relative standard deviation (RSD), resolution, signal precision as RSD, limit of detection, limit of quantification, and accuracy as recovery) of both approaches were <4%, approximately 1, < or = 5%, <8 microM, 30 microM, between 91% and 104%, and <4%, < or = 2%, <9%, < or = 6 microM, < or = 20 microM (with the exception of protocatechuic acid, which shows values of 40 and 130 microM, respectively), between 80% and 107% for the CSEID and IAD concepts, respectively, which are excellent for food samples analysis. A set of representative samples was analyzed including apple and pear skins and pulps, red and white wines, and green tea tablets. High agreement was observed between the results of the sample analyses from the two microchip-based approaches, and good correlation was observed with results obtained from traditional methods. Although the prominent phenolic antioxidant classes and compounds were successfully determined, some nonprominent peaks were not detected in the samples when applying the IAD approach. Different integration strategies on microchip platforms were further explored, looking for a simplification of the overall analytical process without losing the excellent analytical characteristics obtained in both approaches. The results are promising and indicative of the progress of analytical microfluidics toward the "plateau of productivity" and the routine laboratory application.
Micro-total analysis systems (microTAS) integrate different analytical operations like sample preparation, separation and detection into a single microfabricated device. With the outstanding advantages of low cost, satisfactory analytical efficiency and flexibility in design, highly integrated and miniaturized devices from the concept of microTAS have gained widespread applications, especially in biochemical assays. Electrochemistry is shown to be quite compatible with microanalytical systems for biochemical assays, because of its attractive merits such as simplicity, rapidity, high sensitivity, reduced power consumption, and sample/reagent economy. This review presents recent developments in the integration of electrochemistry in microdevices for biochemical assays. Ingenious microelectrode design and fabrication methods, and versatility of electrochemical techniques are involved. Practical applications of such integrated microsystem in biochemical assays are focused on in situ analysis, point-of-care testing and portable devices. Electrochemical techniques are apparently suited to microsystems, since easy microfabrication of electrochemical elements and a high degree of integration with multi-analytical functions can be achieved at low cost. Such integrated microsystems will play an increasingly important role for analysis of small volume biochemical samples. Work is in progress toward new microdevice design and applications.
In this paper we used a chromatographic response function (CRF) that included an output for each of the two performance parameters (resolution and analysis time) to optimize the separation of five bisphenols by micellar electrokinetic chromatography (MEKC) using multivariate response surface methodology (RSM). To validate the real utility of this approach, we have also compared the efficiency of the proposed optimization method with a traditional univariate analysis. For both methods, the selected variables of the analysis were: buffer concentration, pH, amount of organic solvent, and concentration of surfactant. The predictive nature of a validated response surface design allowed for the elucidation of a strong interactive effect and resulted in a more labor-efficient optimization when compared to the univariate approach.
In this paper, we describe the separation and detection of six phenolic acids using an electrophoretic microchip with pulsed amperometric detection (PAD). The selected phenolic acids are particularly important because of their biological activity. The analysis of these compounds is typically performed by chromatography or standard CE coupled with a wide variety of detection modes. However, these methods are slow, labor intensive, involve a multistep solvent extraction, require skilled personnel, or use bulky and expensive instrumentation. In contrast, microchip CE offers the possibility of performing simpler, less expensive, and faster analysis. In addition, integrated devices can be custom-fabricated and incorporated with portable computers to perform on-site analysis. In the present report, the effect of the separation potential, buffer pH and composition, injection time and PAD parameters were studied in an effort to optimize both the separation and detection of these phenolic acids. Using the optimized conditions, the analysis can be performed in less than 3 min, with detection limits ranging from 0.73 microM (0.10 microg/mL) for 4-hydroxyphenylacetic acid to 2.12 microM (0.29 microg/mL) for salicylic acid. In order to demonstrate the capabilities of the device, the degradation of a mixture of these acids by two aquatic plants was followed using the optimized conditions.
A simple method to fabricate cylindrical carbon electrodes for use in capillary electrophoresis (CE) microchips is described. The electrodes were fabricated using a metallic wire coated with carbon ink. Several experimental variables were studied in order to establish the best conditions to fabricate the electrode. Finally, the electrodes were integrated in a poly(dimethylsiloxane) microchip and used for the analysis of phenolic compounds. Using the optimum conditions, the analysis of a mixture of dopamine, epinephrine, catechol, and 4-aminophenol was achieved in less than 240 s, showing good linear responses (R(2)=0.999) in the 0.1-190 microM range, and limits of detection (without the use of stacking or a decoupler) of 140 and 105 nM for dopamine and epinephrine, respectively.
Poly(dimethylsiloxane) (PDMS) is one of the most convenient materials to construct capillary electrophoresis microchips. Even though PDMS has many advantages, its use is often limited by its hydrophobicity. Although it is well-known that the surface properties of PDMS can be modified by anionic surfactants, very little is known regarding the driving forces or the electrophoretic consequences of the adsorption of anionic surfactants. In this work, the adsorption of alkyl surfactants on PDMS was studied by performing electroosmotic flow (microEOF) measurements. In order to mimic the behavior of PDMS microchannels, fused-silica capillaries were coated with PDMS and used for the microEOF measurements. This approach allowed using standard CE instrumentation and provided significant advantages over similar experiments performed on microchips. The change in the microEOF in the presence of surfactants was correlated to the surfactant adsorbed amount which, plotted versus surfactant concentration, gives an adsorption isotherm. The adsorption isotherms were obtained using alkyl surfactants with different chain lengths and head groups. According to our results, the interaction of alkyl surfactants with the PDMS surface is determined by a combination of hydrophobic and electrostatic interactions, where the former is more significant than the latter. The affinity of each surfactant for the PDMS surface was calculated by fitting the adsorption profiles with a Langmuir equation and, in the case of single-charged surfactants, correlated to the corresponding cmc value.
The application of CE to the analysis of additives and organic contaminants in food since 2001 has been reviewed. Additives included in the review are dyes, preservatives, antioxidants and sweeteners. Organic contaminants included in the review are pesticides, antibiotics and those produced during processing or cooking food. The review highlights the strategies and approaches adopted in making CE suitable for the determination of analytes at residue level and in complicated matrices.
Voltammetry on ultramicroelectrodes was used to determine antioxidants tert-butylhydroxyanisole (BHA), tert-butylhydroxytoluene (BHT) and propyl gallate (PG) in vegetable oils or lard. The present work demonstrates the advantages of using not only the conventional three electrode configuration but also the non-conventional two electrode configuration cell system, with respect to the determination of antioxidants. Different ways of performing the determinations are compared either through step by step extractive procedures or through direct measurements using standard addition. Detection limits in the range of (0.20–1.25)×10−5 mol dm−3, (1.00–1.25)×10−5 mol dm−3 and (0.60–1.00)×10−5 mol were obtained for BHA and BHT in vegatable oil and PG in lard determinations, respectively, depending upon the medium composition. Relative standard deviations for antioxidants concentration were about 6–7% for most of media studied and can be compared with those reported for HPLC from collaborative studies, i.e. 4.54, 6.61 and 3.18% for BHT, BHA and PG, respectively.
Antioxidants can be divided into two broad classes, primary and secondary, depending upon their mode of action. The most broadly used primary antioxidants are hindered phenolics. Phenolic antioxidants have traditionally been based on 2,6-di-t-butyl-4-methylphenol (BHT). This functional moiety has been incorporated into larger molecules affording products with lower volatility or greater polymer compatibility. More recently, molecules have been introduced which vary in steric hindrance about the phenol and also the way in which multiple phenol functional groups are linked to form larger molecules. This has led to structures which have lower colour contribution. Surprisingly, in some cases these molecules have shown higher levels of efficiency relative to other antioxidants with similar phenol/molecular weight ratios. This has included enhanced levels of synergism with secondary antioxidants. An attempt is made to correlate structural features of these molecules to the enhanced performance and/or lower colour.
A flow-through optosensor with solid phase UV spectroscopic detection is proposed for the direct determination of single antioxidants, namely butylated hydroxyanisole (BHA) and n-propyl gallate (n-PG), without previous derivatization. The methods are based on the transient retention behaviour of these compounds in a flow-through cell packed with C-18 silica using ethanol–water mixtures as a carrier, and on the intrinsic absorbance monitored at 290 and 283 nm, respectively. After recording the analytical signal, the antioxidants were easily and quickly desorbed from the solid support by the same carrier. For BHA, calibration graphs were linear over the range 1.0–300.0 mg L−1 using area as the analytical parameter. The relative standard deviation (RSD) was between 0.5 and 1.6%. For n-PG, calibration graphs were linear over the range 1.0–300.0 mg L−1 in area and the RSD was between 1.4 and 1.5% .The methods were applied to the
determination of these antioxidants in several food and cosmetics samples, and were validated using the standard additions method and an HPLC reference method.
The potential of antioxidants to reduce the cellular damage induced by ionizing radiation has been studied in animal models for more than 50 years. The application of antioxidant radioprotectors to various human exposure situations has not been extensive although it is generally accepted that endogenous antioxidants, such as cellular non-protein thiols and antioxidant enzymes, provide some degree of protection. This review focuses on the radioprotective efficacy of naturally occurring antioxidants, specifically antioxidant nutrients and phytochemicals, and how they might influence various endpoints of radiation damage. Results from animal experiments indicate that antioxidant nutrients, such as vitamin E and selenium compounds, are protective against lethality and other radiation effects but to a lesser degree than most synthetic protectors. Some antioxidant nutrients and phytochemicals have the advantage of low toxicity although they are generally protective when administered at pharmacological doses. Naturally occurring antioxidants also may provide an extended window of protection against low-dose, low-dose-rate irradiation, including therapeutic potential when administered after irradiation. A number of phytochemicals, including caffeine, genistein, and melatonin, have multiple physiological effects, as well as antioxidant activity, which result in radioprotection in vivo. Many antioxidant nutrients and phytochemicals have antimutagenic properties, and their modulation of long-term radiation effects, such as cancer, needs further examination. In addition, further studies are required to determine the potential value of specific antioxidant nutrients and phytochemicals during radiotherapy for cancer.
We have found that phenolic antioxidants specifically induce expression of the c-fos and c-jun protooncogenes. After treatment of quiescent human hepatoma HepG2 cells with butylated hydroxytoluene, butylated hydroxyanisole, or other phenolic antoxidants, the levels of c-fos and c-jun mRNAs are substantially increased. This response is antioxidant specific, dose dependent, and transient, with maximal levels at 3-6 h. The antioxidant-specific induction of c-fos/CAT promoter constructs in transient transfections indicates that at least a portion of this response is transcriptional. Deletions and point mutations map sequences required for the antioxidant response of the c-fos promoter to the serum response element. The antioxidant-specific induction of expression directed by a reporter plasmid containing four AP-1 sites and the induction of AP-1 DNA-binding activity confirm previous results indicating that antioxidant treatment increases AP-1 activity.
Copper (Cu) is an integral part of many important enzymes involved in a number of vital biological processes. Although normally bound to proteins, Cu may be released and become free to catalyze the formation of highly reactive hydroxyl radicals. Data obtained from in vitro and cell culture studies are largely supportive of Cu's capacity to initiate oxidative damage and interfere with important cellular events. Oxidative damage has been linked to chronic Cu-overload and/or exposure to excess Cu caused by accidents, occupational hazards, and environmental contamination. Additionally, Cu-induced oxidative damage has been implicated in disorders associated with abnormal Cu metabolism and neurodegenerative changes. Interestingly, a deficiency in dietary Cu also increases cellular susceptibility to oxidative damage. A number of nutrients have been shown to interact with Cu and alter its cellular effects. Vitamin E is generally protective against Cu-induced oxidative damage. While most in vitro or cell culture studies show that ascorbic acid aggravates Cu-induced oxidative damage, results obtained from available animal studies suggest that the compound is protective. High intakes of ascorbic acid and zinc may provide protection against Cu toxicity by preventing excess Cu uptake. Zinc also removes Cu from its binding site, where it may cause free radical formation. Beta-carotene, alpha-lipoic acid and polyphenols have also been shown to attenuate Cu-induced oxidative damage. Further studies are needed to better understand the cellular effects of this essential, but potentially toxic, trace mineral and its functional interaction with other nutrients.
Electrosynthesis of benzoquinone from phenol was investigated in acetonitrile–water mixtures on α-PbO2 and β-PbO2 electrodes. The influence of water concentration, electrolysis potential, initial phenol concentration and temperature on benzoquinone production was also studied. Different electrocatalytic activities of α and β structures of PbO2 were determined.
This paper reports on a study by reflectometry of the adsorption process of phenolic compounds electrochemically oxidized on glassy carbon electrodes. This technique provides information about the adsorption rate and the maximum adsorbed amount. The adsorption process depends on several variables like adsorbent concentration, solution pH and working potential, while ionic strength has no significant influence. The best adsorption conditions were obtained at high phenolic concentrations (0.1 M), high pH values (>10) and working potential values higher than 0.600 V. The adsorption process on glassy carbon electrodes showed irreversible behavior, reaching the maximum adsorbed amount at relatively short times (less than 100 s).
Electrochemical oxidation of phenolic compounds leads to a deposition of polymeric films on the surfaces of Pt, glassy carbon, pyrolytic graphite and carbon fiber electrodes, decreasing the electrode activity (passivation). Addition of EDTA to the medium produces a significant enhancement in the electrochemical behavior of phenolic compounds, avoiding electrode passivation. An improvement in the selectivity and sensitivity for the quantitative voltammetric determination of dopamine, catechol and 4-aminophenol was also verified. The electroactivity of the electrode was maintained even after long periods of use in the presence of EDTA. The effect of the EDTA concentration on the electrochemical behavior of some phenolic compounds is reported.
A new poly (vinyl choloride) (PVC)-composite electrode is described for the determination of propyl gallate (PG), octyl gallate (OG) and tert-butylhydroxyanisole (BHA) used as antioxidants in processed foods. This electrode has favourable properties with respect to its mechanical resistance, very low cost, simple preparation and easy renewal of the electrode surface. A new method to determine antioxidants is developed in order to demonstrate the applicability of this PVC-graphite electrode as an amperometric detector in a flow-injection system using a wall-jet flow cell. The samples are injected into the carrier stream consisting of a KNO3 solution and 10 or 20% (v/v) methanol. Linear calibration graphs are obtained within the ranges 0.1–30, 0.1–13 and 0.1–35mgl−1 for PG, OG and BHA, respectively. The applicability of the method was demonstrated by analysing synthetic samples. Finally, the method was applied to the determination of the antioxidants in sunflower oils and soups.
A flow injection method for the determination of the antioxidant BHT using polymer modified electrodes in connection with amperometric detection is proposed. The procedure is based on the electrochemical oxidation at 0.700 V (vs. Ag/AgCl[3M NaCl]) in phosphate buffer solution (pH 7.2). BHT is determined over the range 0.001–0.020 or 0.005–0.030 gL ¹1 . Other analytical parameters, such as electrode working potential, flow rate, electrodeposited charge and solution pH were optimized. The proposed method was applied to the determination of BHT in electrical transformer and vegetable oils samples. The results were compared with the HPLC-UV detection standard technique.
MEKC belongs to a mode of CE but also to micro-LC
An amperometric flow-injection method for the determination of the antioxidants BHA, TBHQ and a mixture of them, using a 4-hydroxybenzaldehyde/formaldehyde polymer modified electrode is proposed. The procedure is based on the electrochemical oxidation in phosphate buffer solution (pH 7.2). Different analytical parameters and electrode stability were analyzed to obtain the best electrode performance, being the optimal conditions: working potentials 0.200 V and 0.700 V and flow rate 0.74 mLmin–1. The proposed method was used for antioxidants quantification in a cosmetic sun tanning lotion, without any previous sample treatment and mean recoveries of 104% were obtained. Interferences from other antioxidants (BHT and PG) were also analyzed and in all the cases, no synergic behavior was observed.
There is a need to develop analytical methods that are capable of rapidly measuring small biological markers in the field of metabolomics. Among others, carbohydrates play an important role biologically yet are traditionally hard to detect since they have no chromophore or fluorophore. In the present report, the first application of integrated pulsed amperometric detection (iPAD) coupled with microchip electrophoresis to the analysis of glucose, mannose, sucrose, maltose, glucosamine, lactose, maltotriose and galactose is demonstrated. iPAD is an electrochemical detection mode that can be used for direct detection of carbohydrates, amines and sulfur containing compounds. The effect of different solution parameters, including the buffer concentration, pH and the concentration of SDS on both separation and detection response was analyzed. In addition, a comparison study between PAD and iPAD was performed using glucose, glucosamine, sucrose and maltose as model carbohydrates.
Pulsed electrochemical detection (PED) is an excellent method for detection of analytes that normally foul electrodes. In PED, the detection electrode is first cleaned at a high positive potential, then reactivated at a negative potential dissolving the surface oxide, and finally used to oxidize the analyte at a moderate positive potential. Due to the advantages and versatility of PED, many different variations of the detection waveform can be found in literature. This review focuses on application of PED to CE and in particular, the most commonly used modes: pulsed amperometric detection (PAD) and integrated pulsed amperometric detection (iPAD).
Pulsed electrochemical detection (PED) has been applied to the direct (i.e., requires no derivatization), sensitive and reproducible detection of numerous polar aliphatic compounds (e.g., carbohydrates, amines, and thiols). These compounds, many of which have biological significance, typically have been classified as non-electroactive for detection under constant applied potentials and have poor optical detection properties. PED exploits the electrocatalytic activity of noble metal (e.g., Au and Pt) electrode surfaces to oxidize various polar functional groups using multi-step potential-time waveforms to realize amperometric/coulometric detection while maintaining uniform and reproducible electrode activity. The response mechanisms in PED are dominated by the surface properties of the electrode, and as a consequence, members of each chemical class of compounds produce virtually identical voltammetric responses. Thus, the full analytical potential is achieved when combined with an a priori separation. Although popularized in combination with high performance liquid chromatography, the combination of PED with highly efficient microseparation techniques offer the analyst unique advantages. This paper reviews the fundamental aspects of PED especially at microelectrodes, and its application in microchromatographic and electrophoretic separation techniques, including microchip devices.
Electrochemistry detection offers considerable promise for capillary-electrophoresis (CE) microchips, with features that include remarkable sensitivity, portability, independence of optical path length or sample turbidity, low cost and power requirements, and high compatibility with modern micromachining technologies. This article highlights key strategies in controlled-potential electrochemical detectors for CE microchip systems, along with recent advances and directions. Subjects covered include the design of the electrochemical detection system, its requirements and operational principles, common electrode materials, isolation from the separation voltage, derivatization reactions, typical applications, and future prospects. It is expected that electrochemical detection will become a powerful tool for CE microchip systems and will lead to the creation of truly portable (and possibly disposable) devices.
The use of a composite graphite–Teflon–tyrosinase biosensor for the determination of the additive propyl gallate in fatty food samples is reported. The compatibility of this composite biosensor with predominantly nonaqueous media allowed the use of reversed micelles—formed with ethyl acetate as the continuous phase, a 5% of a phosphate buffer solution of pH 7.4 as the dispersed phase, and 0.10 mol L−1 dioctyl sulfosuccinate (AOT) as the emulsifying agent—as a suitable working media for the determination of propyl gallate. Monitoring of the enzyme reaction involving the additive was performed by electrochemical reduction (at −0.10 V) of the corresponding o-quinone formed during the catalytic oxidation of propyl gallate. Composite bioelectrodes allow the regeneration of the electrode surface by polishing and exhibit long-term operation (70 days). Moreover, the behaviour of PG as an inhibitor-like of the phenol oxidation reaction catalysed by tyrosinase was studied from an analytical point of view. This process is due to a competition of the two substrates for the enzyme active centers. The analytical characteristics were very similar using both measurement methodologies (the direct amperometric and the inhibition-like responses). Performance of the composite biosensor for the analysis of propyl gallate in foodstuffs was checked using lard as a sample. An aliquot of the analyte extract in ethyl acetate was transferred directly to the electrochemical cell containing the reversed micellar medium.
During vulcanization, rubber is treated with different substances that have a marked influence on the properties of the end product. Identifying and determining the chemical species responsible for such properties is rendered difficult by the fact that some are incorporated into the vulcanizate or decompose during the process. In this work, we developed a new method for the determination of vulcanization accelerators and antioxidants based on Fourier Transform Infrared spectra of the CCl4 extracts of vulcanized rubber specimens. Calibration is performed by using Partial Least-Squares Regression (PLSR); the additive concentrations prior to vulcanization are related to the spectra of the fragments or decomposition products extracted from the polymer matrix following vulcanization. The effects of the spectral mode (absorbance or first-derivative) and wavenumber range were assessed by constructing various models to ensure accurate determinations. The proposed method was applied to two vulcanizates obtained by two different procedures; the results were quite satisfactory in both cases.
Glycoconjugates were separated by capillary electrophoresis at alkaline pH and the migration order was investigated using different complex-forming agents in the electrolyte. Addition of cationic micelles, formed by cetrimide hydroxide, made the separation of anomers and element analogues possible. Boric acid gave the highest selectivity but the peak efficiency was slightly lower owing to slow kinetics of the complexation mechanism. Although the background electrolyte had very high pH values, no indications of instability of the fused-silica capillary or the analytes were observed under the separation conditions used.
The performance of a graphite–Teflon composite amperometric tyrosinase biosensor for the determination of the food additive propyl gallate (PG) in different types of foodstuffs is reported. The enzyme reaction involves the catalytic oxidation of PG to the corresponding o-quinone, and the electrochemical reduction of this o-quinone was employed to monitor the enzyme reaction. Depending on the nature of the food sample analysed and on the presence of other phenolic antioxidants in these samples, aqueous buffer solutions or predominantly nonaqueous acetonitrile–Tris buffer mixtures were employed as working media. Experimental conditions such as the aqueous solution percentage in the predominantly nonaqueous medium, pH, and the potential to be applied were optimised. Control charts constructed showed a useful lifetime for the biosensor of 40 days when working in phosphate buffer of pH 6.5, and of 50 days in 80:20 acetonitrile–Tris buffer (pH 7.4) mixture. The limits of detection obtained for PG in these media were 9.0×10−7 and 1.1×10−6 mol L−1, respectively. The composite bioelectrode also performed well in the flow-injection mode. PG was determined in dehydrated broth bars using the phosphate buffer solution of pH 6.5 as working medium. However, PG was determined in spiked olive oil in the working medium formed by the 80:20 acetonitrile–Tris buffer mixture, because a liquid–liquid extraction step was carried out. Comparison of the results with those obtained by applying reference methods showed that no significant differences existed at a significance level of 0.05.
The partial electrochemical oxidation of aqueous phenol, as a model organic pollutant, has been studied using a flow-through electromembrane reactor with a novel ceramic-based membrane as the anode material. The ceramic-based membranes were modified to exhibit electrocatalytic activities by depositing nano-sized electrocatalytic inks on the membrane surfaces. Carbon black-supported antimony doped tin oxide, prepared by sol–gel a method, was used as the electrocatalyst. The experiments were carried out at room temperature at flow rates of 250 l m−2 h−1. It was found that phenol is readily oxidized to benzoquinone and organic acids. The amount of energy used for oxidation of phenol was calculated in kWh per gram of phenol converted.
The voltammetric behavior of synthetic food antioxidants, such as butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), propyl gallate (PG) and tert-butylhydroquinone (TBHQ), at a glassy carbon electrode in solution of 0.1 mol l−1 perchloric acid containing 1% methanol has been investigated. The obtained linear sweep voltammograms (LSV) showed that all these four compounds have well-defined oxidation waves with peak potentials of 629, 818, 599 and 501 mV for BHA, BHT, PG and TBHQ, respectively. Linear calibration graphs were also obtained in the concentration ranges of 0.5–15.0, 0.5–8.0, 1.0–15.0 and 1.0–15.0 mg l−1 for BHA, BHT, PG and TBHQ, respectively. However, the voltammetric peaks of these antioxidants seriously overlap and it is difficult to determine them individually from a mixture without prior separation. In this work, a method for simultaneous determination of these antioxidants, based on their oxidation at the glassy carbon electrode, with the aid of chemometric approaches, such as classical least squares (CLS), principal component regression (PCR) and partial least squares (PLS), has been developed. The method was applied to determine the four antioxidants in a set of synthetic mixtures and several commercial food samples; in general, satisfactory results were obtained.
Continuous flow injection and UV spectrophotometric detection have been proposed for simultaneous determination of the two binary mixtures, butylated hydroxytoluene (BHT)/n-propyl gallate (n-PG) and butylated hydroxytoluene (BHT)/butylated hydroxyanisole (BHA), in food and cosmetics samples. The method is based on the different residence times of each antioxidant when the flow cell is packed to a height of 25 mm with silica C18 using methanol–water 50:50% (v/v) as a carrier with a flow rate of 1.25 and 1.10 ml min−1, respectively. The determination of each antioxidant is based on the measurement of its absorbance at its maximum wavelengths using a DAD detector at 30 and 180 s for the mixture n-PG–BHT and 90 and 220 s for BHA–BHT. Calibration graphs were linear over the range 10.0–300.0 μg ml−1 for each antioxidant in both mixtures. The relative standard deviations were 2.5% for BHT and 2.0% for the co-existing antioxidant. Resolution of the n-PG–BHT mixture in ratios between 1:8 and 8:1 and the BHA–BHT mixture in ratios between 1:10 and 10:1 is possible. The method was applied to the determination of both antioxidants in fat foods and cosmetics samples with recoveries ranging between 101 and 105%.
Browning reactions represent an interesting research area for the implications in food technology, nutrition and health. The development of some non-enzymatic browning reactions, such as Maillard reaction, has been recently associated to the formation of compounds with strong antioxidant capacity. In this paper, the relation between colour changes due to non-enzymatic browning and the formation of compounds with antioxidant activity is discussed. Simple positive or complex correlation between colour and antioxidant properties can be found depending on composition and technological history of the product. Complex relations between these variables are generally obtained in multi-component and in formulated foods, where the simultaneous development of a number of reactions, interacting or prevailing Maillard reaction itself, can affect in opposite ways the overall antioxidant properties and colour of the product.
This review deals with HPLC method to be used for the determination of synthetic phenolic antioxidants added to various foods. Sample preparation, isolation techniques, separation systems as well as detection methods used in applied food analysis procedures are discussed.
The present report describes a new analysis strategy for microchip capillary electrophoresis with pulsed amperometric detection and its application to the determination of glucose. The addition of sodium dodecyl sulfate (SDS) to the mobile phase and detection reservoir stabilized flow rates and enhanced the detection signal for glucose. A higher pH (compared to the running buffer) was used at the waste reservoir in order to improve the detection performance while maintaining good separations. To our knowledge, this is the first report describing the use of post-column pH modification using microchip electrophoresis. Under optimum conditions, a linear relationship between the peak current and the concentration of glucose was found between 10−2–10−5 M, with a limit-of-detection of 1.2 μM. In addition, the separation of glucosamine and glucose was performed at pH 7.1 while the detection was performed at pH 11 to demonstrate the ability to use post-column pH modification.
A new and efficient method for the determination of synthetic phenolic antioxidants (SPAs) has been developed by using micellar electrokinetic capillary chromatography (MECC) with electrochemical detection. Under the optimum conditions, all analytes were successfully separated within 13 min at the separation voltage of 18 kV in a 20 mmol/L borate running buffer (pH 7.4) containing 25 mmol/L sodium dodecyl sulfate. The excellent linearity was obtained in the concentration range from 5.0 × 10−4 to 2.0 × 10−6 mol/L and the detection limits (S/N = 3) of propyl gallate (PG), tert-butylhydroquinone (TBHQ), butylated hydroxyanisole (BHA), and butylated hydroxytoluene (BHT) range from 2.9 × 10−7 to 2.7 × 10−6 mol/L. This method has been proved to be effective and successfully applied for the determination of SPA in food products, providing a promising and convenient entry to monitor the superscale use of phenolic antioxidants.
This review covers the applications of mass spectrometry with sample introduction in the liquid phase to the analysis of the oxidation products of DNA, proteins and lipids in vitro and in vivo. The analysis of antioxidant xenobiotics in biological systems is also covered. The review covers the period 1997–late 2002. The most commonly used mass spectrometric mode used in this area of study is electrospray mass spectrometry that may also include collision-induced dissociation (CID). The review summarises for each paper the important methodological aspects of the techniques used and, where appropriate, the importance of the observations made within the context of biology.
A preliminary study of the antioxidant content of hand-produced grain derivatives from the Canary Islands, named gofio, is developed by means of a capillary electrophoresis (CE) method. Protocatechuic acid, salicylic acid, p-hydroxybenzoic acid, vanillic acid, syringic acid, p-coumaric acid, ferulic acid and sinapic acid were extracted from corn, wheat, barley and pea gofio samples by sonication with methanol and were successfully separated by CE using a polycation, hexadimetrine bromide (HDB), as electrosmotic flow modifier. The separation was achieved using a running buffer consisting of 125 mM boric acid, 49 mM disodium hydrogen phosphate, 0.002% (w/v) HDB and 2.5 mM α-cyclodextrin at pH 7.5. The developed procedure allowed the simultaneous determination of the selected antioxidants in less than 3.5 min. The electrophoretic profile obtained for each kind of flour (corn, wheat, barley and pea) allowed differentiation of the type of gofio. Among the analyzed samples, corn gofio showed the highest antioxidant content. Furthermore, highly roasted corn or wheat gofio samples were also analyzed and their antioxidant contents were lower than those of less roasted samples.
The voltammetric behaviour of the antioxidants 3-tert-butyl-4-hydroxyanisole (BHA) and tert-butylhydroquinone (TBHQ), at a polymer electrode modified with nickel phthalocyanine as electron mediator, is described, and an electroanalytical method for the determination of these antioxidants based on their electrochemical oxidation on the modified electrode is proposed. Cyclic voltammograms showed well-defined oxidation peaks slightly shifted towards less positive potentials with respect to those obtained at the platinum disk electrode for both antioxidants. The peak currents measured at the modified electrode are considerably higher than those obtained at the unmodified electrode. A scan rate of 100 mV s(-1), a minimum methanol percentage of 0.5% and a 0.1-M Britton-Robinson medium were chosen as working conditions. The obtained results suggest that the BHA oxidation produces the TBHQ reduction product. This product is oxidised during the second cyclic scan, to generate the mentioned TBHQ as final product. These studies also reveal that sensitive response for both antioxidants can be obtained by using the differential pulse voltammetry (DPV) technique. Responses observed are dependent on the nature of the control solution (% methanol and pH) and the nature of voltammetric conditions (potential amplitude, DeltaE, and scan rate, v(b)). These factors have been modified in order to find the best analytical conditions. The mutual interferences between both antioxidants and the measurement reproducibility were tested. Using cyclic voltammetry (CV) and differential pulse voltammetry, BHA and TBHQ linear calibration graphs were obtained. The detection limits were 2.1 ppm for both when CV was used, and 18.7 ppm (BHA) and 1.23 ppm (TBHQ) for DPV. The developed methods were applied to the determination of BHA in spiked glaze biscuits and TBHQ in spiked mushroom cream.
We have found that phenolic antioxidants specifically induce expression of the c-fos and c-jun protooncogenes. After treatment of quiescent human hepatoma HepG2 cells with butylated hydroxytoluene, butylated hydroxyanisole, or other phenolic antoxidants, the levels of c-fos and c-jun mRNAs are substantially increased. This response is antioxidant specific, dose dependent, and transient, with maximal levels at 3-6 h. The antioxidant-specific induction of c-fos/CAT promoter constructs in transient transfections indicates that at least a portion of this response is transcriptional. Deletions and point mutations map sequences required for the antioxidant response of the c-fos promoter to the serum response element. The antioxidant-specific induction of expression directed by a reporter plasmid containing four AP-1 sites and the induction of AP-1 DNA-binding activity confirm previous results indicating that antioxidant treatment increases AP-1 activity.
A micellar electrokinetic chromatography method was developed to simultaneously analyse commonly used food additives. The additive mixture, comprising propyl gallate, octyl gallate, dodecyl gallate, butylated hydroxyanisole, butylated hydroxytoluene, tertiary butylhydroquinone, p-hydroxybenzoic acid methyl ester, p-hydroxybenzoic acid ethyl ester, benzoic acid, sorbic acid, saccharin, aspartame and acesulfame-K, was not resolved using single surfactant micellar systems consisting of sodium dodecyl sulfate (SDS), sodium cholate (SC) or sodium deoxycholate (SDC). The separation of these additives using mixed micellar systems, involving SDS/SC, SDS/SDC and SC/SDC, was investigated. Organic solvents were added to the mixed micellar phases to optimise the separation. The mixture was successfully separated using a 20 mM borate buffer with 35 mM SC, 15 mM SDS and 10% methanol added at pH 9.3. Additives in cola beverages and low-joule jam were investigated and quantified using this method.
Paracetamol (acetaminophen) and hydroxyurea were found to inhibit DNA synthesis in a dose-dependent manner in tissue slices in vitro, with little effect on protein synthesis. Considerable variation in the sensitivity of the different tissues was also observed with an order of least sensitive to most sensitive tissue of liver < testis < spleen. The phenolic antioxidant properties of paracetamol are thought to be the mechanism by which paracetamol inhibits DNA synthesis, which led us to study other phenolic antioxidant molecules and flavonoids for specific inhibition of DNA synthesis. (+)-catechin, m-aminophenol, p-aminophenol and p-cresol all displayed a highly specific inhibition of DNA synthesis. Quercetin displayed a preferential inhibition of DNA synthesis but a significant level of inhibition of protein synthesis was also seen. Nordihydroguaiaretic acid (NDGA) and n-propyl gallate showed preferential inhibition of DNA synthesis at the lower doses tested, but at higher doses showed significant inhibition of protein synthesis, presumably because of cytotoxicity. Caffeic acid and naringenin did not display any specific inhibition of DNA synthesis as protein synthesis was equally inhibited at all doses tested. This study demonstrates that certain phenolic antioxidants can inhibit DNA synthesis specifically but this is not a property shared by all phenolic antioxidants; and that these inhibitors show considerable variation in effectiveness between different tissues.
Capillary electrokinetic chromatography is suitable for the separation of mixtures of uncharged and charged solutes. In the present work the behavior of six synthetic food antioxidants--2[3]-tert.-butyl-4-hydroxyanisole, 2,6-di-tert.-butyl-p-cresol, tercbutylhydroquinone, 3,4,5-trihydroxybenzoic acid propyl ester, 3,4,5-trihydroxybenzoic acid octyl ester and 3,4,5-trihydroxybenzoic acid dodecyl ester--was studied in a capillary electrophoresis system using capillary electrokinetic chromatography with vesicles of the surfactant bis(2-ethylhexyl)sodium sulfosuccinate (AOT). Several studies aimed at calculating the critical aggregation concentration of the surfactant were conducted to check that under the conditions used the AOT was in a state of aggregation. Having checked the association shown by the surfactant, we then explored the greater or lesser capacity of the antioxidants to interact with this compound. We followed the evolution of the molecular absorption spectra of each of the antioxidants in the presence of the surfactant at different concentrations and the retention factors were calculated at different pH values. Additionally, in order to determine which species--anionic or neutral--was present at the pH of the buffer used (boric/borate), the pKa values in acetonitrile-water (20:80) were obtained. Resolution and quantification of the antioxidants demand optimization of the variables involved in the system, such as the percentage of acetonitrile, the concentration of AOT and boric/borate buffer, pH, voltage, etc. When this part of the study had been completed, calibrations were obtained for each of the antioxidants, obtaining good linear correlation coefficients in all cases. Finally, we propose a method that allows the resolution of the six most employed antioxidants in a capillary electrophoretic system in 15 min, using electrokinetic chromatography with AOT as the pseudostationary phase.
The world-wide aviation jet fuel used for civil and military aircraft is of a kerosene type. To avoid peroxide production after the refinery process a specific antioxidant additive should be added on fuel. The antioxidants generally used are based on hindered phenols in a range of concentration 10-20 microg/ml. In the present work a specific method to measure the concentration of phenolic antioxidants is shown. The method is based on a liquid chromatographic technique with electrochemical detection. The technique, because of its selectivity, does not require sample pre-treatments. The analysis of a 5-10 ml fuel sample can be performed in less than 10 min with a sensitivity of 0.1 microg/ml and a RSD=2.5%. A comparison with another highly selective gas chromatographic technique with mass spectrometric detection with selected ion monitoring (GC-MS-SIM) is reported. The sensitivity of GC-MS-SIM method was 2 microg/ml with a RSD=3.1%.
Both carcinogenic and anticarcinogenic properties have been reported for the synthetic antioxidants butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT). The association between dietary intake of BHA and BHT and stomach cancer risk was investigated in the Netherlands Cohort Study (NLCS) that started in 1986 among 120,852 men and women aged 55 to 69 years. A semi-quantitative food frequency questionnaire was used to assess food consumption. Information on BHA or BHT content of cooking fats, oils, mayonnaise and other creamy salad dressings and dried soups was obtained by chemical analysis, a Dutch database of food additives (ALBA) and the Dutch Compendium of Foods and Diet Products. After 6.3 years of follow-up, complete data on BHA and BHT intake of 192 incident stomach cancer cases and 2035 subcohort members were available for case-cohort analysis. Mean intake of BHA or BHT among subcohort members was 105 and 351 microg/day, respectively. For consumption of mayonnaise and other creamy salad dressings with BHA or BHT no association with stomach cancer risk was observed. A statistically non-significant decrease in stomach cancer risk was observed with increasing BHA and BHT intake [rate ratio (RR) highest/lowest intake of BHA = 0.57 (95% confidence interval (CI): 0.25-1.30] and BHT = 0.74 (95% CI: 0.38-1.43). In this study, no significant association with stomach cancer risk was found for usual intake of low levels of BHA and BHT.
Chronic pulmonary inflammatory diseases predispose towards lung cancer by unknown mechanisms. Butylated hydroxytoluene (BHT) administration to mice causes lung injury and a subsequent inflammatory response, and when administered chronically to certain inbred strains following carcinogen treatment, increases lung tumor multiplicity. We hypothesize that inflammation promotes lung tumor growth in this model system and have begun to examine this hypothesis by assessing inflammatory parameters in inbred strains that vary in their susceptibility to promotion. Positive correlations were found between susceptibilities to tumor promotion and BHT induction of alveolar macrophage and lymphocyte infiltration into alveolar airspaces, and increased vascular permeability (P < .03, P < .04, and P < .005, respectively). The amounts of pulmonary cyclooxygenase (COX)-1 and COX-2 did not strongly correlate with promotion. Because persistent elevation of macrophage content is the hallmark of a chronic inflammatory response, the alveolar macrophage population was depleted by adding chlorine to the drinking water prior to carcinogenesis. This treatment reduced lung tumor multiplicity following 2-stage carcinogenesis (P < .05). These correlations between inflammatory and tumorigenic responses to BHT, along with decreased tumorigenesis after macrophage depletion, are consistent with a role of inflammation in promotion. Inflammatory mediators may provide targets for early diagnosis and chemoprevention.
Cosmetics are commercially available products that are used to improve the appearance of the skin. Since the late 1980s, consumer demand for more effective products that more substantively beautify the appearance has resulted in increased basic science research and product development in the cosmetics industry. The result has been more ingredients that may actually improve not just the appearance of the skin, but the health of the skin as well. We now have products that renew, restore, and rejuvenate—not just cleanse, protect, and moisturize. There is probably no greater focus of interest currently than the incorporation of vitamins and antioxidants in skin care products. There are considerable data to suggest the benefits of such ingredients in cosmetics. This article reviews the published data that support the usefulness of vitamins and antioxidants in cosmetics. Our job as dermatologists is to digest and assess the data so we can give knowledgeable recommendations to our patients. The ingestion and absorption of vitamins and antioxidants, most importantly through diet, and secondarily through intake of manufactured supplements, is critical to the health of human beings. The skin is the largest organ; as our primary external barrier, it is on the forefront of the battle with external causes of damaging free radicals. Ultraviolet light and environmental pollutants are known initiators of free radicals. Free radicals are highly reactive molecules with an unpaired electron that result in damage to surrounding molecules and tissues. The most significant damage by free radicals is to biomembranes and to DNA. It is thought that additional, topical use of vitamins and antioxidants in cosmetics can better protect and possibly correct the damage by neutralizing these free radicals. In addition, some vitamins may be beneficial to the skin because of other actions such as effects of suppression of pigmentation and bruising, stimulation of collagen production, refinement of keratinization, or anti-inflammatory effects. Vitamin A
The antioxidant 3,5-di-tert-butyl-4-hydroxy-toluene (BHT) is widely used as an additive to increase the tenability of food and plastics. BHT is degraded to 3,5-di-tert-butyl-4-hydroxybenzaldehyde (BHT-CHO) in mammals, as well as in the natural environment such as in water and soils. BHT-CHO has been studied extensively in terms of their potential toxicities. The present investigation was carried out to quantify BHT and BHT-CHO in river, ground, rain and drinking water obtained from several locations in Germany. Apart from the compounds mentioned above, 1,2-bis-(3,5-di-tert-butyl-4-hydroxyphenyl)ethane (2-BHT), which is a dimer of BHT, was also detected in the extracts of some ground water samples. The applied analytical method is based on solid phase extraction (SPE) to concentrate trace compounds from water samples followed by gas chromatography/mass spectrometry (GC/MS) of the extracts. A total of 51 of the respective water samples were used for extraction purposes and analyte recoveries were all > or = 80%. The determination limit for BHT was 5 ng l(-1) and for BHT-CHO 16 ng l(-1). The standard deviations for the analytical procedure were 6% for BHT and 10% for BHT-CHO. The use of the antioxidant BHT in Germany has resulted in water concentrations of 7-791 ng l(-1) in the rivers Rhine, Elbe, Main, Oder, Nidda and Schwarzbach. The degradation product BHT-CHO was also detected in the river water samples at concentrations between 29 and 223 ng l(-1). The concentrations of BHT measured in German rivers are lower compared to values measured in the USA and Japan 20 years ago. In ground water, levels for BHT varied from non-detectable up to 2156 ng l(-1) and for BHT-CHO from non-detectable up to 674 ng l(-1). Both compounds were also detected in rain water in Frankfurt/Main at a concentration of 1797 ng l(-1) for BHT and 59 ng l(-1) for BHT-CHO.
Artichoke leaf is an herbal medicine known for a long time. A systematic antioxidant activity-directed fractionation procedure was used to purify antioxidative components from the aqueous methanol extractions of artichoke heads and leaves in this study. Seven active polyphenolic compounds were purified from artichoke, and structural elucidation of each was achieved using MS and NMR. Two of these compounds, apigenin-7-rutinoside and narirutin, were found to be unique to artichoke heads, this represents the first report of these compounds in the edible portion of this plant. The contents of these antioxidants and total phenols in dried artichoke samples from leaves and immature and mature heads of three varieties, Imperial Star, Green Globe, and Violet, were then analyzed and compared by colorimetric and validated HPLC methods. Significant differences by variety and plant organ were observed.
Cerebral ischemia and recirculation cause delayed neuronal death in rodents, such as Mongolian gerbils and stroke-prone spontaneously hypertensive rats (SHRSP), which were used as an experimental stroke model. It was documented that an enhanced nitric oxide production, the occurrence of apoptosis, and an attenuated redox regulatory system contribute to the development of delayed neuronal death. Many studies have suggested the beneficial antioxidant effects of antioxidant nutrients such as vitamin E, green tea extract, ginkgo biloba extract, resveratrol and niacin in cerebral ischemia and recirculation brain injury. These results are important in light of an attenuation of the deleterious consequences of oxidative stress in ischemia and recirculation injury.
Solar radiation induces acute and chronic reactions in human and animal skin. Chronic repeated exposures are the primary cause of benign and malignant skin tumors, including malignant melanoma. Among types of solar radiation, ultraviolet B (290-320 nm) radiation is highly mutagenic and carcinogenic in animal experiments compared to ultraviolet A (320-400 nm) radiation. Epidemiological studies suggest that solar UV radiation is responsible for skin tumor development via gene mutations and immunosuppression, and possibly for photoaging. In this review, recent understanding of DNA damage caused by direct UV radiation and by indirect stress via reactive oxygen species (ROS) and DNA repair mechanisms, particularly nucleotide excision repair of human cells, are discussed. In addition, mutations induced by solar UV radiation in p53, ras and patched genes of non-melanoma skin cancer cells, and the role of ROS as both a promoter in UV-carcinogenesis and an inducer of UV-apoptosis, are described based primarily on the findings reported during the last decade. Furthermore, the effect of UV on immunological reaction in the skin is discussed. Finally, possible prevention of UV-induced skin cancer by feeding or topical use of antioxidants, such as polyphenols, vitamin C, and vitamin E, is discussed.
Analytical strategies dealing with bioactive phenols in plants and foods are reviewed. These depend on the purpose of the analysis which may be classified as studies where the principal purpose is biological screening, phytochemical and/or chemical screening. Nevertheless, extraction of the phenol from the sample matrix is common and methods of achieving a suitable extract are assessed. Advances in the separation sciences and spectrometry are exploited for identification and quantification of isolated phenols. The various procedures are summarized and some typical "case studies" are presented. Two important areas are introduced briefly. Thus, plant phenols are reactive species and their ultimate fate has been relatively neglected. Studies of bioactive compounds generate a considerable volume of data making data handling and informatics important topics that warrant a separate review.
Olive oil is the main source of fat in the Mediterranean diet, and its consumption has been related to a low incidence of coronary heart disease and certain cancers. Recent findings demonstrate that olive oil phenolics are powerful in vitro and in vivo antioxidants and display other biological activities that could partially account for the observed healthful effects of the Mediterranean diet. A detailed method optimization plan was carried out to separate the most popular phenols in olive oil for four separation parameters: buffer concentration, buffer pH, applied voltage and temperature. Consequently, an analytical method capable of separating 21 different phenols and polyphenols by capillary zone electrophoresis was developed; the separation was performed within 10 min, using a 40 cm x 50 microm capillary, with a 45 mM sodium tetraborate buffer (pH 9.60), at 27 kV and 30 degrees C. The optimized method was applied to methanolic extracts of several Italian extra-virgin olive oils obtained by different technologies in order to characterize and to compare their antioxidant profile. Positive correlations of phenolic compounds found by capillary zone electrophoresis (CZE) and two colorimetric indexes (total polyphenols and o-diphenols) were found and discussed.
The separation and detection of underivatized carbohydrates, amino acids, and sulfur-containing antibiotics in an electrophoretic microchip with pulsed amperometric detection (PAD) is described. This report also describes the development of a new chip configuration for microchip electrophoresis with PAD. The configuration consists of a layer of poly(dimethylsiloxane) that contains the microfluidic channels, reservoirs, and a gold microwire, sealed to a second layer of poly(dimethylsiloxane). Example separations of carbohydrates, amino acids, and sulfur-containing antibiotics are shown. The effect of the separation and injection potentials, buffer pH and composition, injection time, and PAD parameters were studied in an effort to optimize separations and detection. Detection limits ranging from 6 fmol (5 microM) for penicillin and ampicillin to 455 fmol (350 microM) for histidine were obtained.
The antibacterial activity of a series of alkyl gallates (3,4,5-trihydroxybenzoates) against Gram-positive bacteria was tested using a broth dilution method. All of the Gram-positive bacteria tested were susceptible to alkyl gallates, and this activity was found to correlate with the alkyl chain length. The antibacterial activity of alkyl gallates against Bacillus subtilis was a parabolic function of their lipophilicity and maximized with alkyl chain length between C(8) and C(11). Notably, alkyl gallates were found to be bactericidal against B. subtilis ATCC 9372, but this activity was significantly affected by the endospore formation in the culture. The antibacterial activity of alkyl gallates likely comes at least in part from their ability to inhibit the membrane respiratory chain but is not due to the prooxidant action.
Creatinine, creatine, and uric acid are three important compounds that are measured in a variety of clinical assays, most notably for renal function. Traditional clinical assays for these compounds have focused on the use of enzymes or chemical reactions. Electrophoretic microchips have the potential to integrate separation power of capillary electrophoresis with devices that are small, portable, and have the speed of conventional sensors. The development of a microchip CE system for the direct detection of creatinine, creatine, and uric acid is presented. The device uses pulsed amperometric detection (PAD) to detect the nitrogen-containing compounds as well as the easily oxidizable uric acid. Baseline separation of creatinine, creatine and uric acid was achieved using 30 mM borate buffer (pH = 9.4) in less than 200 s. Linear calibration curves were obtained with limits of detection of 80 microM, 250 microM and 270 microM for creatinine, creatine and uric acid respectively. An optimization of the separation conditions and a comparison of PAD with other amperometric detection modes is also shown. Finally, analysis of a real urine sample is presented with validation of creatinine concentrations using a clinical assay kit based on the Jaffé reaction.
A method was developed for simultaneous analysis of natural antioxidants in beer using multichannel electrochemical detection with a CoulArray detector, which enables selective and sensitive antioxidant detection in gradient HPLC and facilitates the identification of analytes based on the ratios of signals recorded at different potentials applied to the detection cells arranged in series. The separation conditions were optimised for 27 phenolic compounds including derivatives of benzoic and cinnamic acids, flavones, and a few related glycosides identified in beer samples. Separation selectivities of 11 columns with different stationary phase chemistries were compared, and the pH and gradient programs were optimised for the individual columns to provide best resolution and high number of resolved peaks, using the window-diagram approach. The effects of pH on the sensitivity of electrochemical coulometric detection were considered in the optimisation approach. The optimised conditions were applied to the analysis of real beer samples.