Summary
An effective, simple and rapid analytical method using HPLC was developed for the analysis of monosodium glutamate (MSG) in various food samples obtained from local market in Turkey. The ...determination of MSG was performed by its derivatisation with o‐phthaldialdehyde (OPA). A high‐performance liquid chromatography‐ultraviolet/diode array detection method was performed by using C18 (150 mm × 4.6 mm, 2.7 μm) column with the mobile phase consisting of 10 mm phosphate buffer solution (pH = 5.90) and methanol (75:25, v/v). The applied method was optimised and the validated. The method was linear from 1 to 50 μg mL−1 of MSG. The correlation coefficient value of the developed method was obtained as R2 = 0.9999. The limit of detection and limit of quantification limits were 0.015 and 0.050 μg mL−1, respectively. MSG contents of the food samples range from 0.09 g kg−1 to 120.80 g kg−1. The validated method was successfully applied for the analysis of MSG in several food samples.
HPLC analysis of monosodium glutamate in various food samples.
•Hydrolysis of proteins in foods is performed by acid, alkaline and enzymatic methods.•o-Phthaldialdehyde (OPA) reacts with amino acids to form fluorescent derivatives.•OPA-amino acids adducts are ...efficiently separated by HPLC.•Sample preparation for HPLC analysis is simple, easy, convenient, and automated.•The method can be used to determine amino acids in proteins of animal tissues/foods.
Studies of protein nutrition and biochemistry require reliable methods for analysis of amino acid (AA) composition in polypeptides of animal tissues and foods. Proteins are hydrolyzed by 6M HCl (110°C for 24h), 4.2M NaOH (105°C for 20h), or proteases. Analytical techniques that require high-performance liquid chromatography (HPLC) include pre-column derivatization with 4-chloro-7-nitrobenzofurazan, 9-fluorenyl methylchloroformate, phenylisothiocyanate, naphthalene-2,3-dicarboxaldehyde, 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate, and o-phthaldialdehyde (OPA). OPA reacts with primary AA (except cysteine or cystine) in the presence of 2-mercaptoethanol or 3-mercaptopropionic acid to form a highly fluorescent adduct. OPA also reacts with 4-amino-1-butanol and 4-aminobutane-1,3-diol produced from oxidation of proline and 4-hydroxyproline, respectively, in the presence of chloramine-T plus sodium borohydride at 60°C, or with S-carboxymethyl-cysteine formed from cysteine and iodoacetic acid at 25°C. Fluorescence of OPA derivatives is monitored at excitation and emission wavelengths of 340 and 455nm, respectively. Detection limits are 50fmol for AA. This technique offers the following advantages: simple procedures for preparation of samples, reagents, and mobile-phase solutions; rapid pre-column formation of OPA-AA derivatives and their efficient separation at room temperature (e.g., 20–25°C); high sensitivity of detection; easy automation on the HPLC apparatus; few interfering side reactions; a stable chromatography baseline for accurate integration of peak areas; and rapid regeneration of guard and analytical columns. Thus, the OPA method provides a useful tool to determine AA composition in proteins of animal tissues (e.g., skeletal muscle, liver, intestine, placenta, brain, and body homogenates) and foods (e.g., milk, corn grain, meat, and soybean meal).
One approach for the synthesis of isoindolinones, a privileged bioactive heterocyclic core structure, involves a condensation reaction of o-phthaldialdehydes with a suitable nitrogen-containing ...nucleophile. This fascinating reaction is revisited here in the context of the use of o-phthaldialdehydes that contain additional substituents in the aromatic ring leading to a detailed analysis of the regioselectivity of the reaction. Eleven monosubstituted o-phthaldialdehydes were synthesised and reacted with alanine. The regioselectivity observed across the eleven substrates led to the design of a disubstituted substrate that reacted with very high control. A gram-scale reaction followed by esterification gave one major regioisomer in high yield. In addition, the regioselectivity observed on reaction of two novel monodeuterated substrates led to an increased mechanistic understanding.
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•PT altered gluten features and morphology in a dose-dependent manner.•PT increased the hardness, viscoelasticity and thermal stability of gluten.•PT interfered gluten cross-linking ...and gluten conformation.
The effects of persimmon tannin (PT) on the texture, viscoelasticity, thermal stability, and morphology of gluten were studied and the underlying mechanisms were also explored. The results showed that PT increased the hardness and viscoelasticity but lowered the cohesiveness and extensibility of gluten in a dose-dependent manner. Additionally, PT increased the denaturation temperature and enthalpy of gluten, and induced the formation of gluten with compact structure. High concentration of PT (8%) significantly increased the hardness and viscoelasticity of gluten, and induced the formation of compact structure of gluten by disturbing the conformation of gluten, and interfering gluten cross-linking through decreasing disulfide bonds, free sulfydryl groups, and free amino groups. In contrast, low concentration (0.25%) of PT slightly altered the gluten properties and morphology. Our work extended the study on the supplementation of phenolic compounds in wheat flour-based products.
A sensitive chiral high performance liquid chromatography (HPLC) method for the determination of aliphatic primary amino alcohol isomers with o‐phthaldialdehyde/mercaptoethanol precolumn ...derivatization has been developed and validated. Seven chiral columns were tested in a reversed phase mode. Excellent enantioseparation with the resolution more than 2.0 was achieved on Chiralcel OJ‐3R. The effect of various chromatographic conditions including column temperature, acetonitrile content in the mobile phase, buffer pH, buffer concentration, and buffer type in the mobile phase on the retention and the selectivity was investigated. The final mobile phase consisted of binary mixture of 20mM ammonium formate solution with acetonitrile (75:25; v/v). The analyses were performed at mobile phase flow rate of 1.0 mL/min and the column temperature of 40°C. The fluorescence detection was performed at excitation wavelength of 345 nm and emission wavelength of 450 nm. The developed method was fully validated in terms of linearity, sensitivity, accuracy, precision, intermediate precision, and selectivity according to International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use guidelines using internal normalization procedure. The proposed chiral method was proved to be highly sensitive, simple, and rapid and was successfully applied to the determination of D‐Valinol content in commercially available samples of L‐Valinol.
The purpose of this ex vivo study was to compare the cleaning performance of three commercially available orthodontic cleaners on polymethyl methacrylate (PMMA) test specimens covered with biofilm.
...Twenty subjects wore an individually manufactured vacuum-formed maxillary splint with four integrated PMMA test specimens for 7 days. The four test specimens were located on the buccal surfaces of the maxillary molars. After a 7-day wearing period, the PMMA test specimens colonized by biofilm were divided into two halves. One half was placed in 150 ml of tap water or in 150 ml of cleaning solution of the cleaners Retainer Brite® (Dentsply International Raintree Essix, Sarasota, FL, USA), Kukis® Xpress (Reckitt Benckiser, Heidelberg, Germany) or Dontodent (Propack, Heidelberg, Germany) while the other half remained uncleaned. The modified o‑phthaldialdehyde (OPA) method was used to determine the amount of protein on both halves of the test specimens. The difference was tested for significance as a measure of the cleaning effect using a paired sample t‑test.
The cleaning performance of the three orthodontic cleaners was higher than the cleaning performance of tap water (mean 25.9 ± 6.5%). While Retainer Brite® (mean 54.5 ± 7.1%) removed significantly more biofilm than Dontodent (mean 41.5 ± 9.2%, p < 0.001) and Kukis® Xpress (mean 39.9 ± 11.5%, p < 0.001), there was no significant difference in the cleaning performance between Kukis® Xpress and Dontodent (p = 1).
Seven-day-old biofilm is only removed partially by the investigated orthodontic cleaners, so that they are not suitable as the only measure for removing established biofilms.
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•A novel HILIC method for the determination of pharmaceutically active thiols was presented.•The effect of chromatographic conditions on the retention was investigated.•The postcolumn ...reaction conditions such as reaction temperature and reagent flow rate were studied.•The proposed HILIC method was validated in accordance with ICH guidelines.
A rapid, precise and specific hydrophilic interaction chromatography (HILIC) combined with postcolumn derivatization using o-phthaldialdehyde and fluorescence detection was developed and validated for the determination of selected pharmaceutically active thiols. The analysis was carried out on a Diol HILIC column using a mobile phase consisting of acetonitrile and solution of 10 mmol/L citric acid adjusted with 1-propylamine to pH 5.5 in ratio 75:25 (v/v) for separation of cysteine and homocysteine and in ratio 85:15 (v/v) for separation of N-acetyl-l-cysteine and captopril. The postcolumn derivatization reaction was performed at room temperature using reagent (5 mmol/L OPA in 0.05 mol/L 4- (2-hydroxyethyl) piperazine-1-ethanesulfonic acid at pH 7) delivered at the flow rate of 0.3 mL/min. Fluorescence detection was carried out at excitation and emission wavelength of 345 nm and 450 nm, respectively. The effect of chromatographic conditions including acetonitrile content, salt concentration in the mobile phase and mobile phase pH on the retention of tested thiols was investigated. The postcolumn reaction conditions such as reaction temperature, derivatization reagent flow rate, o-phthaldialdehyde concentration and derivatization reagent pH were deeply studied. The developed method was validated in terms of linearity, accuracy, precision and selectivity according to the International Conference on Harmonisation guidelines. The HILIC method was successfully applied for the analysis of commercially available samples of pharmaceutically active thiols such as captopril, N-acetyl-l-cysteine (NAC) and cysteine.
To help to clarify therapeutic functions of lipoic acid (LA) in biochemical and clinical practice we have elaborated a fast, simple and accurate HPLC method enabling determination of LA in human ...urine. The proposed analytical approach includes reduction of LA with tris(2‐carboxyethyl)phosphine and simultaneous separation and derivatization of the analyte with butylamine and o‐phthaldialdehyde followed by spectrofluorimetric detection at λex = 340 nm and λem = 440 nm. The assay was performed using gradient elution and the mobile phase containing 0.0025 mol L−1 o‐phthaldialdehyde in 0.0025 mol L−1 NaOH and acetonitrile. Linearity of the detector response for LA was observed in the range of 0.3–8 μmol L−1. Limits of detection and quantification for LA in urine samples were 0.02 and 0.03 μmol L−1, respectively. The total analysis time, including sample work‐up, was <20 min. The analytical procedure was successfully applied to analysis of real urine samples delivered from six healthy volunteers who received a single 100 mg dose of LA.
Abstract
Plasma amino acids are generally analyzed through ion exchange chromatography, a reproducible but time‐consuming method. Here, we report the optimization of a reverse‐phase‐high‐performance ...liquid chromatography with fluorescence detector (RP‐HPLC‐FLD) assay for the detection and quantification of plasma amino acids for potential applications in metabolic disorders (e.g., aminoacidopathies, a rare group of Inborn Errors of Metabolism). For assay development, initially standard amino acids were derivatized with ortho‐phthalaldehyde‐3‐mercaptopropionic acid (OPA‐3‐MPA) and filtered through a 0.20 μm syringe filter. The excitation and emission wavelengths of 240–450 nm (λex—λem) were used for the detection of amino acids. Chromatographic separation was achieved by gradient RP‐HPLC‐FLD through C18 symmetry column (150 × 4.6 mm, particle size 3.5 μm). HPLC assay was successfully optimized and was able to detect amino acids in the range of 10–400 ng/mL and good linearity (R
2
> 0.98) was achieved in the mixture for each standard amino acid. Moreover, the current assay showed great efficiency with two additional advantages: the use of low‐cost mobile phases, and the detection and quantification of amino acids at low level (ng/mL) concentration in biofluids. This assay could be applied for the analysis of human plasma to identify aminoacidopathies in newborn screening programs, and other metabolic disorders.
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•A novel chiral HPLC method for the enantioseparation of N-acetyl-dl-cysteine was presented.•N-acetyl-dl-cysteine enantiomers were derivatized using o-phthaldialdehyde and primary ...aliphatic amine.•The impact of primary aliphatic amine nature on retention of L-NAC derivatives was investigated.•The chiral HPLC method was validated using internal normalization.
A sensitive and rapid high-performance liquid chromatography (HPLC) method was developed to enantioseparation of N-acetyl-dl-cysteine after precolumn derivatization using o-phthaldialdehyde and primary aliphatic amines. Seven polysaccharide-based chiral columns were tested in a reversed phase mode. Under the optimal chromatographic conditions, N-acetyl-dl-cysteine derivatives were completely enantioseparated on Chiralcel OZ-3R column with the resolution more than 2.5. The impact of various primary aliphatic amine additives as co-reagents (ethyl-, 1-propyl-, 1-butyl-, 1-pentylamine, (R)-sec-butylamine, tert-butylamine, isobutylamine, cyclopropyl-, cyclobutyl-, cyclopentyl and cyclohexylamine) used in precolumn derivatization step on the retention behavior (retention factor, selectivity and column efficiency) of N-acetyl-dl-cysteine derivatives was investigated. The effect of chromatographic conditions including acetonitrile content in the mobile phase, mobile phase pH, salt concentration in the mobile phase and column temperature on the retention and selectivity was investigated. The developed method was properly validated in terms of linearity, sensitivity (limit of detection and limit of quantification), accuracy, precision, intermediate precision and selectivity according to International Council for Harmonisation (ICH) of Technical Requirements for Pharmaceuticals for Human Use guidelines using internal normalization procedure. Proposed HPLC method was successfully applied to the determination of optical purity in commercially available N-acetyl-L-cysteine samples.
