Derivatization With 2,4-dinitrophenylhydrazine Research Articles

Volatile Carbonyl Compounds Emission in Dry-Process Fibreboard: Identification through a Selective GDME-HPLC-DAD-MS/MS Method

This work presents a novel application of gas-diffusion microextraction (GDME) for the extraction and identification of volatile carbonyl compounds from a dry-process fibreboard (MDF) by combining high-performance liquid chromatography with diode array detection (HPLC–DAD) and mass spectrometry (MS). GDME is a simple, inexpensive, and environmentally friendly technique that allows the simultaneous extraction and derivatization with 2,4-dinitrophenylhydrazine (DNPH) of selected volatiles. The extraction conditions were optimized using design of experiments through a 24 full fractional design followed by a Box–Behnken design, resulting in 35 min of extraction at 45 °C using 500 µL of DNPH 0.15%. The analysis of commercial MDF samples with different characteristics, such as thickness or colour, showed a distinct emission profile of volatile carbonyls. The principal emitting compounds found were formaldehyde, acetaldehyde, acetone, butanal, pentanal, hexanal, heptanal, octanal, and nonanal. A total of 25 compounds were identified using MS, including saturated and unsaturated aldehydes, ketones, dicarbonyls, and benzaldehyde-derivatives. This method can be a valuable tool for the qualitative evaluation of VOCs released from wood-based panels and for the assessment of indoor-air quality.

4-Hydroxynonenal - A Toxic Leachable from Clinically Used Administration Materials.

The migration of chemicals from processing materials into biopharmaceuticals can lead to various problems. Leachables from administration materials, with no possibility of further clearance, are of particular concern. Released chemicals can be toxic or react with formulation components, thereby impacting product safety. Therapeutic proteins, which are susceptible to chemical modifications, have highest risk to be affected. The aim of this study was to identify a previously unknown leachable compound from clinical administration sets, which was present above the applied generic safety threshold. Extracts of commonly used clinical administration sets were analyzed using a recently established specific assay allowing the identification and quantification of the α,β-unsaturated aldehyde 4-hydroxynonenal (HNE) in a drug product surrogate solution. HNE was quantified after derivatization with 2,4-dinitrophenylhydrazine (DNPH) and liquid extraction of the formed hydrazone by LC-MRM analysis. Potentially genotoxic HNE was a leachable compound from all tested administration sets, in parts exceeding safety thresholds for genotoxicants. The HNE-releasing polymer was identified as PVC. Clinical administration sets should be, like manufacturing materials and container closure systems, in the focus of routine leachables studies. Manufacturers of clinical administration sets should show responsibility to avoid the presence of safety concerning chemicals, like HNE.

4-Hydroxynonenal is An Oxidative Degradation Product of Polysorbate 80.

Polysorbates (PS) are used in biopharmaceuticals to stabilize therapeutic proteins. Oxidative degradation of (poly)unsaturated fatty acids (PUFAs) contained in PS was shown to lead to α,β-unsaturated carbonyls. The n-6-PUFA linoleic acid accounts for up to 18% of all FAs contained in multi-compendial grade PS80. 4-Hydroxynonenal (HNE) is highly reactive towards nucleophilic amino acids, potentially leading to covalent protein modifications. This study tests whether HNE may be a pharmaceutically relevant PS80 peroxidation product. Since HNE was not directly detectable in the PS80 matrix by UV and MS, a new quantification method was established. After derivatization with 2,4-dinitrophenyl hydrazine (DNPH) and extraction of the formed hydrazone with a salting-out assisted liquid-liquid extraction, the HNE-DNPH adduct was analyzed by multiple reaction monitoring. Kinetic oxidation studies were conducted incubating PS80 in presence and absence of the antioxidant butylhydroxytoluene (BHT). HNE was confirmed as PS80 degradant in oxidatively stressed samples. BHT was shown to prevent its formation. HNE is a detectable PS80 degradation product raising questions about the potential impact on critical quality attributes of biopharmaceuticals formulated with PS80. Addition of BHT prevented HNE formation under oxidative stress. Consequently, BHT might be a valuable additive in PS used in biopharmaceuticals.

Spectrophotometric Determination of Dopamine in Bulk and Dosage Forms Using 2,4-Dinitrophenylhydrazine.

Dopamine (DA) hydrochloride is a sympathomimetic agent used therapeutically for the correction of hemodynamic disorders associated with shock episodes. Although several analytical methods have been described, a spectroscopic assay of DA after chemical derivatization with 2,4-dinitrophenylhydrazine (DNP) is still unexamined. Therefore, the optimization of the reaction parameters and validation of developed method were required. The method is based on coupling of DA as a phenolic compound with a diazonium salt to produce an intensely colored azo derivative. DNP was oxidized with potassium periodate to produce a diazonium salt that coupled with DA in basic media. The experimental parameters were then optimized. The developed method was validated according to International Conference on Harmonisation Guidelines and was applied to dosage forms. The results were compared with the data of a reference method. The method was linear in a concentration range between 5 and 50 μg/mL. The regression line equation was Y=0.042±0.0003X+0.0672±0.0015 with a regression coefficient of 0.9944 (n=5). The limit of detection and limit of quantification were 0.32 and 0.97 μg/mL, respectively. The precision was satisfactory; the percentage relative standard deviation did not exceed 2%. The average values of the recovery study were in the range 98.90- 100.40±0.31-1.21%. The developed method was applied successfully for the determination of DA in injection and infusion fluid. The method is accurate, sensitive, and practical for DA analysis in quality control laboratories.

A Method to Purify a Sample Derivatized with 2,4-Dinitrophenylhydrazine (DNPH) Using Solid Phase Extraction

A Method to Purify a Sample Derivatized with 2,4-Dinitrophenylhydrazine (DNPH) Using Solid Phase Extraction

Extraction of acetaldehyde from mineral water using 2,4- dinitrophenylhydrazine as derivatizing agent

Acetaldehyde generated, when PET bottles of water are deposited at high temperature for long period of time. The acetaldehyde produced, transferred into the mineral water and alter the organoleptic properties of water. Acetaldehyde is also a possible carcinogen and mutagen. In the present research work, acetaldehyde in mineral water was derivatized with 2, 4-dinitrophenyl hydrazine (DNPH), extracted by solid phase extraction (SPE) using octadecyl silane oxide (C18) and finally investigated by gas chromatography (GC). Acetaldehyde-2, 4-dinitropheyl hydrazone (A-DNPH) formed after derivatization which were adsorbed on the surface of C18. Solvent acetonitrile was used for the elution of A-DNPH from the solid phase extraction (SPE) column. Effect of storage time and light on the migration of acetaldehyde was investigated and found the mineral water stored for longer period of time in sun light change their organoleptic properties due to migration of acetaldehyde into water. Detection limit of this method was 9.8µg/L while the efficiency of the method was 87.6%, which was higher than solid phase micro-extraction. The concentration of acetaldehyde determined by this method was in the range of 9.8 to 61 µg/L. The results indicate that C18 packed in column had good efficiency to hold the acetaldehyde-DNPH, hence it is good adsorbent for acetaldehyde extraction from bottled mineral water. Thus, it has been concluded that bottled water should not be stored at elevated temperature for longer period of time. High temperature and long storage duration increase the acetaldehyde concentration in bottled water which changes the organoleptic properties of water. Water with acetaldehyde concentration greater than 20 g/L have changed organoleptic properties but is acceptable for consumption. Key words: Acetaldehyde, Octadecyl silane oxide, Acetonitrile, Poly (ethylene terephthalate), Gas chromatography

Method development of levoglucosenone analysis by UHPLC-UV-MS in fast pyrolysis samples and aspects of its degradation in aqueous samples.

Levoglucosenone ((1S,5R)-6,8-dioxabicyclo[3.2.1]oct-2-en-4-one or LGO; CAS number: 37112-31-5) is an extensively studied biomass based chemical and its potential applications in organic synthesis, as well as in the extraction of various natural substances, have been established; therefore developing dependable, yet fast and simple analytical methods for determining LGO in the products of thermochemical biomass processing is of great importance. In this work different UHPLC methods have been developed and compared for the determination of LGO in wood pyrolysis products. In addition to UV detection, LGO determination conditions by single quadrupole mass spectrometry have been adjusted with and without chemical derivatization. The ionization of LGO in the positive and negative electrospray ionisation mode and the effect of a mobile phase additive on LGO and its degradation product ionisation have been evaluated. Using the UHPLC-UV system, a method for the quantification of levoglucosenone was developed and the analytical parameters of the method were determined. Utilising the developed UHPLC-UV method, studies of LGO stability in standard solution and in fast pyrolysis products were carried out in various solvent systems at different temperatures and pH levels. The main LGO primary degradation product has been confirmed to be furfural. It has been concluded that the degradation rate of LGO is significantly affected by the solvent and storage temperature, so it is essential that LGO standard solutions and samples are analysed when freshly prepared. Using a UHPLC-MS system and derivatization with 2,4-dinitrophenylhydrazine (DNPH), a method for the determination of LGO was developed and the method analytical parameters were tested. To sum up, quantitative determination of LGO in pyrolysis products can be performed by reversed phase UHPLC-UV without any particular sample preparation, but in the case of UHPLC-MS it is necessary to derivatize LGO with DNPH to ensure ionisation of LGO. The additional work, requiring MS detection, can be outweighed by the greater selectivity compared to that of the more straightforward UV detection method in cases of incomplete chromatographic separation.

Applicability of FTIR-ATR Method to Measure Carbonyls in Blood Plasma after Physical and Mental Stress.

Introduction Oxidative stress is a state of imbalance between the production of reactive oxygen species and antioxidant defenses. It results in the oxidation of all cellular elements and, to a large extent, proteins, causing inter alia the formation of carbonyl groups in their structures. The study focused on assessment of changes in the plasma protein-bound carbonyls in police horses after combat training and after rest and the applicability of infrared spectroscopy with a Fourier transform, utilizing the attenuated total reflectance (FTIR-ATR) in detecting plasma protein oxidation. Methods We evaluated the influence of both the different concentrations of hydrogen peroxide and combat training on protein carbonylation in horse blood plasma. The oxidation of plasma proteins was assessed using a spectrophotometric method based on the carbonyl groups derivatization with 2,4-dinitrophenylhydrazine (DNPH). The measured values were correlated with the carbonyl groups concentrations determined by means of the FTIR-ATR method. Results The linear correlation between the DNPH and FTIR-ATR methods was shown. The concentration of plasma protein-bound carbonyls significantly deceased in police horses after one-day rest when compared to the values measured directly after the combat training (a drop by 23%, p<0.05 and 29%, p<0.01 measured by DNPH and FTIR-ATR methods, respectively). These results were consistent with the proteins phosphorylation analysis. ConclusionThe FTIR-ATR method may be applied to measure the level of plasma proteins peroxidation.

Quantification of malondialdehyde in exhaled breath condensate using pseudo two-dimensional ultra-performance liquid chromatography coupled with single quadrupole mass spectrometry

We developed a robust analytical method for quantification of malondialdehyde (MDA) in exhaled breath condensate (EBC) via derivatization with 2,4-dinitrophenylhydrazine (DNPH). The target MDA-DNPH hydrazone was separated by ultra-performance liquid chromatography using two reversed-phase analytical columns (C18 and phenyl-hexyl) inter-connected via a two-position, six-port switching valve to a single-quadrupole mass spectrometer. The target derivative was analyzed under positive electrospray ionization using single ion monitoring mode (m/z = 235 for the target derivative, and m/z = 237 for its labeled isotopic analog). This pseudo two-dimensional chromatographic separation provided optimum separation conditions for the target derivative resulting in the limit of detection of 0.58 nM in EBC sample (or 36.2 pmol on-column amount), which is comparable to those reported previously using different techniques, including tandem mass spectrometry. Based on the calibration solutions, the method had a linear quantification range of 1.0–200 nM (r2 = 0.998). The method showed good relative recoveries (92.2–102.0%) and acceptable precisions (3.6–12.2% for inter-day precision, and 4.3–12.4% for intra-day precision for two quality control levels, prepared from 5 nM and 25 nM solutions). The derivative was found to be stable at room temperature for 48 h or during analysis. The method was used to analyze 205 exhaled breath condensate samples collected from individuals from a healthy population of student athletes. MDA was detected in approximately 95% of these samples, with concentrations ranging from 1.16 to 149.63 nM. The median concentration was 6.82 nM, (IQR 4.08–9.88). These data demonstrate that our method can be successfully used to measure MDA in population studies.

Protein Carbonylation Pattern Is Altered in Myelodisplastic Syndromes

* A.R. and M.L. have contributed equally. INTRODUCTIONMyelodysplastic syndrome (MDS) is a clonal disease that arises from the expansion of mutated hematopoietic stem cells. Oxidative stress could play an important role in the pathology of MDS since a correlation has been observed between high ROS levels and decreased survival in MDS patients. In addition, the oxidation of proteins leads to their aggregation, change and conformational changes that can cause the loss of their functions. Due to alteration of the proteome has been described in patients with MDS, in this study we considered the discovery of protein carbonylation to analyze its oxidation pattern. In addition, as the oxidative stress signaling pathways can regulate the cell cycle through p53, we analyzed p21, an important p53 target during the stress response. Finally, the possible benefit of treatment with Deferasirox as antioxidant therapy was studied. METHODSCarbonylation pattern was analyzed in the different cell lines (n = 14, 6 MDS, 4 reactive controls and 4 MDS + DFX) by immunohistochemistry by derivatization with 2,4-dinitrophenylhydrazine (DNPH) and detection with anti-DNP antibodies. The degree of protein carbonylation was studied, also by derivatization and detection with anti-DNP, in primary cells of erythroblasts expanded for 11 days, by one-dimensional and two-dimensional electrophoresis (n = 16; 7 MDS, 6 controls, 2 reactive controls and 1 MDS + DFX). Expression levels of p21 were analyzed by qRT-PCR (n=19; 8C, 8 MDS, 3DFX). RESULTSImmunohistochemical assays revealed a higher level of carbonylation in patients with MDS (Fig. 1a). Interestingly, Deferasirox-treated MDS patients had lower levels than non-treated patients (Fig. 1b). In addition, to evaluate the carbonylation in the erythroid precursors, the pattern of carbonylated proteins in the expanded erythroblasts was analyzed. Again, a higher level of MDS was observed compared to the control group, which was reversed after treatment with Deferasirox. Two-dimensional electrophoresis revealed that differences in carbonylation are due to a limited number of proteins or aggregates, rather than to a large spectrum of proteins. Finally, an increased p21 expression was observed in patients with myelodysplasia and, surprisingly, this effect was reversed with Deferasirox treatment. CONCLUSIONPatients with MDS showed a differential protein pattern respect to control patients, both in the myeloid and erythroid series. The increase in oxidative stress in myelodysplastic patients seems to activate signaling pathways involving p21. Treatment with Deferasirox can reverse the level of oxidation and the increase of p21 in patients with MDS. DisclosuresMartinez Lopez:Celgene: Research Funding, Speakers Bureau; Jansen: Research Funding, Speakers Bureau; BMS: Research Funding, Speakers Bureau; Novartis: Research Funding, Speakers Bureau.

Effects of Charcoal on Carbonyl Delivery from Commercial, Research, and Make-Your-Own Cigarettes.

Previous literature has shown that adding charcoal to cigarette filters can have varying effects on the delivery of toxic carbonyls depending on filter design, amount of charcoal, and puffing profiles. However, these studies have relied on either comparisons between commercially available charcoal and noncharcoal filtered cigarettes or experimental modification of filters to insert a charcoal plug into existing cellulose acetate filters. Make-your-own (MYO) cigarettes can help obviate many of the potential pitfalls of previous studies; thus, we conducted studies using commercial charcoal cigarettes and MYO cigarettes to determine the effects of charcoal on carbonyl delivery. To do this, we analyzed carbonyls in mainstream smoke by HPLC-UV after derivatization with 2,4-dinitrophenylhydrazine (DNPH). Charcoal was added in-line after the cigarettes or through the use of MYO charcoal cigarette tubes. MYO cigarettes had carbonyl deliveries similar to that of 3R4F research cigarette, regardless of tobacco type. The greatest effect on carbonyl delivery was observed with 200 mg of charcoal, significantly reducing all carbonyls under both methods tested. However, "on-tow" design charcoal filters, available on many commercially available charcoal brands, appeared to have a minimal effect on carbonyl delivery under intense smoking methods. Overall, we found that charcoal, when added in sufficient quantity (200 mg) as a plug, can substantially reduce carbonyl delivery for both MYO and conventional cigarettes. As carbonyls are related to negative health outcomes, such reductions may be associated with reductions in carbonyl-related harm in smokers.

Frying influence on virgin olive oils by monitoring change of aliphatic monoaldehydes

When virgin olive oil (VOO) have been fried long time at high temperature, some aliphatic monoaldehydes including hexanal, heptanal, octaldehyde, nonanal and decanal were found in fried VOO and food samples. The HPLC method has been developed for determination of aliphatic monoaldehydes in VOO and food samples after different fried conditions. These five aliphatic monoaldehydes could be separated and analyzed in 60 min on reversed phase ODS column with methanol/0.1% acetic acid water solution mobile phase at 1.0 mL min-1 flow rate, after being derivatized with 2,4-dinitrophenylhydrazine (DNPH). The method was validated with recovery results ranging from 86% to 114%. Calibration plots of aliphatic aldehydes were linear (r≥0.9991) in the concentration range from 1.0×10-6 to 1.5×10-4 mol L-1. The LODs were between 6.5×10-8 and 3.98×10-7 mol L-1. The proposed method provides a reliable and sensitive quantitative evaluation for aliphatic monoaldehydes in fried oils and fried potatoes samples. The experiments results verify that the VOOs are not suitable to be fried long time at high temperature. The frying time should be less than 3.0 min, and the frying temperature should be below 150 °C for the healthy diet.

Vortex-assisted liquid-liquid microextraction based on hydrophobic deep eutectic solvent for determination of malondialdehyde and formaldehyde by HPLC-UV approach

A simple, and cost-effective vortex-assisted liquid-liquid microextraction (VALLME) based on hydrophobic deep eutectic solvent (DES) has been developed to extract malondialdehyde (MDA) and formaldehyde (FA) after derivatization with 2,4-dinitrophenylhydrazine (DNPH). Following the extraction process, a high performance liquid chromatography equipped with UV–Vis detector (HPLC-UV) was used for separation and quantification of the target analytes. The key variables affecting the derivatization process (concentration of DNPH, derivatization time, and derivatization temperature) were optimized by a central composite design (CCD). The effective parameters of VALLME including pH of the extraction medium, ionic strength, extracting agent (DES) volume, dispersive solvent volume, centrifugation rate, centrifugation time, and time of vortex were studied by a fractional factorial design to identify the significant factors effective in the extraction process. In the next step, a CCD was used to obtain the optimum conditions of the effective parameters. Under optimal conditions, good linearities (R2 > 0.99) in the concentration ranges of 5–3000 and 20–2000 ng mL−1 were obtained for MDA and FA, respectively. The detection limits of MDA and FA were found as 2.0 and 10.0 ng mL−1, respectively. The method was successfully applied to the quantification of both analytes in three different real samples (human urine, apple juice and rain water) and satisfactory results in terms of recoveries (91–112%) were achieved.

A METHOD FOR DETERMINATION OF SAXITOXINS USING HPLC-MS WITH 2,4-DINITROPHENYLHYDRAZINE PRECOLUMN DERIVATIZATION

A method for saxitoxin (STX) determination is proposed. It consists of target compound extraction with 0,05 M acetic acid, derivatization with 2,4-dinitrophenylhydrazine (DNPH), and subsequent HPLC electrospray ionization MS analysis. The reaction was performed in acetonitrile-trifluoroacetic acid (98,5:1,5 v/v) at 65 °C; the half-life of STX was ca. 3,6 hours. The method was applied to analysis of natural cyanobacteria samples and a cultivated Nostoc Pruniforme strain. Detection was performed by measuring total ion current, and chromatograms for STX were obtained using an extracted ion procedure at m/z 462,15 (STX hydrazone). A frequent complication of the derivatization with DNPH is the formation of two stereoisomers, E and Z, giving two separate peaks under HPLC. Quantum-chemical calculation predicted that, due to steric hindrance, only one isomer (Z) of STX adduct is formed, and hence only one peak of the analyte is present in the chromatogram. The following metrological parameters were achieved: limits of determination (20 ng/g of dry cyanobacterial biomass), reproducibility (4%), and medium-term precision (± 7%).

Electrochemical impedance spectroscopy measurements for determination of derivatized aldehydes in several matrices.

A simple, selective and sensitive electrochemical method is described for the determination of different aldehydes at glassy carbon electrode using electrochemical impedance spectroscopy (EIS). The measurements were performed after their derivatization with 2,4-dinitrophenylhydrazine (DNPH) in acidic medium. The impedance measurements were investigated in the frequency range from 100 mHz to 100 kHz at a potential of 1.0 V versus Ag/AgCl. The Nyquist plots were modeled with a Randle’s equivalent circuit. The charge transfer resistance was identified as the dependent parameter on relevant concentration of aldehydes (determined as their hydrazones). Under the optimized conditions, the linearity was established over the concentration range of 1000–0.05 μmol L−1. The limits of detection (LODs) obtained were from 0.097 to 0.0109 μmol L−1. Finally, the developed method has been applied to the determination of aldehydes in drinking water, orange juice and apple vinegar samples with relative standard deviations (RSDs) < 3.1% and acceptable recovery rate (around of 80%).

Sensitive and selective quantification of free and total malondialdehyde in plasma using UHPLC-HRMS

Quantification of malondialdehyde (MDA) as a marker of lipid peroxidation is relevant for many research fields. We describe a new sensitive and selective method to measure free and total plasmatic MDA using derivatization with 2,4-dinitrophenylhydrazine (DNPH) and ultra-HPLC-high-resolution MS. Free and total MDA were extracted from minute sample amounts (10 μl) using acidic precipitation and alkaline hydrolysis followed by acidic precipitation, respectively. Derivatization was completed within 10 min at room temperature, and the excess DNPH discarded by liquid-liquid extraction. Quantification was achieved by internal standardization using dideuterated MDA as internal standard. The method's lowest limit of quantification was 100 nM and linearity spanned greater than three orders of magnitude. Intra- and inter-day precisions for total MDA were 2.9% and 3.0%, respectively, and those for free MDA were 12.8% and 24.9%, respectively. Accuracy was 101% and 107% at low and high concentrations, respectively. In human plasma, free MDA levels were 120 nM (SD 36.26) and total MDA levels were 6.7 μM (SD 0.46). In addition, we show the applicability of this method to measure MDA plasma levels from a variety of animal species, making it invaluable to scientists in various fields.

Effects of Topography-Related Puff Parameters on Carbonyl Delivery in Mainstream Cigarette Smoke.

Smoking topography parameters differ substantially between individual smokers and may lead to significant variation in tobacco smoke exposure and risk for tobacco-caused diseases. However, to date, little is known regarding the impact of individual puff parameters on the delivery of many harmful smoke constituents including carbonyls. To examine this, we determined the effect of altering individual puff parameters on mainstream smoke carbonyl levels in machine-smoked reference cigarettes. Carbonyls including formaldehyde, acetaldehyde, crotonaldehyde, propionaldehyde, methyl ethyl ketone (MEK), acrolein, and acetone were determined in cigarette smoke by HPLC after derivatization with 2,4-dinitrophenylhydrazine (DNPH). Deliveries of all carbonyls were nearly two-fold greater when cigarettes were smoked according to the more intense Health Canada Intense (HCI) protocol compared to the International Organization of Standardization (ISO) method, consistent with the two-fold difference in total puff volume between methods (ISO: 280-315 mL; CI: 495-605 mL). When individual topography parameters were assessed, changes in puff volume alone had the greatest effect on carbonyl delivery as predicted with total carbonyls being strongly correlated with overall puff volume (r2: 0.52-0.99) regardless of how the differences in volume were achieved. All seven of the carbonyls examined showed a similar relationship with puff volume. Minor effects on carbonyl levels were observed from vent blocking and changing the interpuff interval, while effects of changing puff duration and peak flow rate were minimal. Overall, these results highlight the importance of considering topography, especially puff volume, when the toxicant delivery and potential exposure smokers receive are assessed. The lack of an impact of other behaviors, including puff intensity and duration independent of volume, indicate that factors such as temperature and peak flow rate may have minimal overall effects on carbonyl production and delivery.

Electrochemical Study of 2,4‐Dinitrophenylhydrazine as Derivatization Reagent and Aldehydes at Carbon Glassy Electrode

AbstractA selective method based on derivatization with 2,4‐dinitrophenylhydrazine (DNPH) is described for the determination of several carbonyl compounds. The factors affecting the derivatization reaction of aldehyde and DNPH were investigated. The product of the derivatization reaction has been characterized by UV/Vis spectrophotometry, NMR, infrared spectroscopy and cyclic voltammetry. Then, an electrochemical study for the determination of aldehyde‐2.4‐dinitrophenylhydrazone was performed at glassy carbon electrode (GCE) using square wave voltammetry (SWV). After the optimization of experimental parameters, the limits of detection (at 3σ) obtained for all aldehyde‐2,4‐DNPH were varied from 15.82 to 78.39 μmol L−1 and relative standard deviations were between 1.8 and 4.5%. Finally, the proposed method was applied to determine the aldehydes concentration in drinking water and orange juice samples with satisfactory results.

Novel DNPH-based method for determination of protein carbonylation in muscle and meat

Protein oxidation is considered an ongoing deteriorative process during storage of fresh and processed meat. Carbonyl compounds have traditionally been detected spectrophotometrically after derivatization with 2,4-dinitrophenylhydrazine (DNPH) to form protein-bound hydrazones with absorbance at 370nm. Here we describe a novel DNPH-based method to quantify protein carbonylation in muscle and meat. The additional steps of the novel method aimed at increasing the protein solubility and inducing protein unfolding before labeling with DNPH. Compared to the traditional method, the new procedure reflected an increased protein carbonylation level measuring overall two to fourfold more carbonyls in muscles from different species as well as in soluble, salt-soluble and insoluble protein fractions. The study suggested that protein unfolding is a more important phenomenon than solubilization for increased DNPH labeling. The novel method resulted in three to fourfold larger carbonyl content determined in chicken, pork and beef (2.8, 3.6 and 3.1nmol/mg of protein, respectively).

Development of a Novel Derivate Assay for Formaldehyde Determination by HPLC in Beer Samples

In this paper, a novel derivative reagent, ethoxyamine hydrochloride (EAHC), was proposed for formaldehyde detection with a simple and highly sensitive pre-column derivation–high-performance liquid chromatography. The effects such as the concentration of derivative reagent, buffer solution pH, reaction temperature, and reaction time on the derivate reaction were also investigated. Under the optimized experimental conditions, the peak area showed a linear relationship with the concentration of formaldehyde in a range of 0.046 to 4.6 mg L−1, with a correlation coefficient of 0.9998. The limit of detection (LOD) and the limit of quantification (LOQ) were 0.016 and 0.042 mg L−1, respectively. The proposed method was applied to the assay of different beer samples, and the recovery of the developed method was in the range of 91.3–104.3 %. To further evaluate the validity of the developed method, formaldehyde was also determined in different beer samples by the referenced HPLC method based on pre-column derivatization with 2, 4-dinitrophenylhydrazine (DNPH). Through the t test method, no obvious difference between the two methods was found at a 95 % confidence level. The results showed that the proposed method was a rapid, convenient, and feasible method for the determination of formaldehyde in beer samples.