For the purpose of determinations, analytical purity reagents (analytical standards) designed for liquid chromatography were used: n-propanol and ethanol from Sigma Aldrich (Steinheim, Germany) and methanol from J.T. Baker (Phillipsburg, KS, USA). All chemicals used for the Kjeldahl method were of p.a. purity grade, manufactured by Merck (Darmstadt, Germany). Buffers, ninhydrin, and a mixture of standards for amino acid identification were obtained from Sykam (Eresing, Germany), standardised for amino acid analyses in the physiological range. Deionised water from a deioniser from Hydrolab Polska HLP 5P was used.
Ninhydrin
Manufactured by Sykam
Sourced in Germany
Ninhydrin is a chemical reagent used in analytical and forensic chemistry. Its core function is to detect the presence of amino acids, proteins, and other compounds containing primary amines. When applied to a surface, Ninhydrin reacts with these compounds to produce a distinctive purple-blue color, allowing for the identification and visualization of trace evidence.
Automatically generated - may contain errors
Lab products found in correlation
Methanol, by Avantor (1 mentions)
Sample dilution buffer, by Sykam (1 mentions)
Milli q purifier, by Merck Group (1 mentions)
Lc ms grade methanol, by Thermo Fisher Scientific (1 mentions)
Sulfuric acid, by Merck Group (1 mentions)
Acetonitrile, by Thermo Fisher Scientific (1 mentions)
Pepsin, by Merck Group (1 mentions)
Pronase e, by Merck Group (1 mentions)
Creatinine, by Merck Group (1 mentions)
N heptane, by Merck Group (1 mentions)
Leucine aminopeptidase, by Merck Group (1 mentions)
Prolidase, by Merck Group (1 mentions)
Boric acid, by Carl Roth (1 mentions)
D glucose, by Merck Group (1 mentions)
Carrez solution 1, by Merck Group (1 mentions)
Hydrochloric acid (hcl), by Merck Group (1 mentions)
Ethanol, by Merck Group (1 mentions)
Acetonitrile, by Merck Group (1 mentions)
D fructose, by Merck Group (1 mentions)
Carrez 2 solution, by Merck Group (1 mentions)
4 protocols using ninhydrin
1
Analytical Standards for LC and Kjeldahl
2
Amino Acid Quantification by HPLC
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Ultrapure water used for the preparation of all solutions were obtained from a Milli-Q purifier (Millipore, Eschborn, Germany). Eluent buffer A (0.12 M citric acid–sodium citrate buffer at pH 3.45) and B (0.20 M citric acid–sodium citrate buffer at pH 10.85), regeneration buffer D (0.50 M sodium hydroxide), sample dilution buffer (0.12 M citric acid–sodium citrate buffer at pH 2.20) and ninhydrin (triketohydrindene hydrate) were purchased from Sykam GmbH Company (Germany). Unless otherwise indicated, reagents used in this work were of analytical reagent grade quality.
A working standard solution was diluted from the Amino Acid Standard Hydrolysate (Sykam, Germany), which contains 2.5 mM of each amino acid including aspartic acid (Asp), threonine (Thr), serine (Ser), glutamic acid (Glu), proline (Pro), glycine (Gly), alanine (Ala), cysteine (Cys), valine (Val), methionine (Met), isoleucine (Ile), leucine (Leu), tyrosine (Tyr), phenylalanine (Phe), histidine (His), lysine (Lys) and arginine (Arg).
A working standard solution was diluted from the Amino Acid Standard Hydrolysate (Sykam, Germany), which contains 2.5 mM of each amino acid including aspartic acid (Asp), threonine (Thr), serine (Ser), glutamic acid (Glu), proline (Pro), glycine (Gly), alanine (Ala), cysteine (Cys), valine (Val), methionine (Met), isoleucine (Ile), leucine (Leu), tyrosine (Tyr), phenylalanine (Phe), histidine (His), lysine (Lys) and arginine (Arg).
3
Quantification of Glycation Adducts
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LC-MS grade methanol and acetonitrile were obtained from Fisher Chemical (Loughborough, UK). Pepsin, pronase E, sulfuric acid and creatinine were from Merck (Darmstadt, Germany), and CML and [ 2 H 2 ]CML were obtained from PolyPeptide Group (Strasbourg, France). 2-Amino-2-(hydroxymethyl)propane-1, 1-diole-hydrochloride (TRIS-HCl) was purchased from Serva Feinbiochemica (Heidelberg, Germany). Nonafluoropentanoic acid (NFPA), hydrochloric acid, prolidase, leucine aminopeptidase and n-heptane were from Sigma-Aldrich (Seelze, Germany). Lithium citrate buffer, lithium citrate/borate buffer, lithium hydroxide and ninhydrin were purchased from Sykam (Fürstenfeldbruck, Germany). Wieninger's catalyst was obtained from Honeywell (Seelze, Germany). Boric acid was purchased from Carl Roth (Karlsruhe, Germany) and sodium hydroxide from Grüssing (Filsum, Germany). Water was distilled (twice) before analysis.
Pyrraline [11] and MG-H1 [12] were synthesized according to the specified literature. The synthesis of [ 13 C 6 , 15 N 2 ] pyrraline and [ 13 C 6 ]MG-H1 was performed in the same manner but using [ 13 C 6 , 15 N 2 ]lysine (pyrraline) and [ 13 C 6 ]arginine (MG-H1) instead of the unlabeled amino acids.
Pyrraline [11] and MG-H1 [12] were synthesized according to the specified literature. The synthesis of [ 13 C 6 , 15 N 2 ] pyrraline and [ 13 C 6 ]MG-H1 was performed in the same manner but using [ 13 C 6 , 15 N 2 ]lysine (pyrraline) and [ 13 C 6 ]arginine (MG-H1) instead of the unlabeled amino acids.
4
Analytical Standards for HPLC Analysis
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For the purpose of determinations, analytical purity reagents (analytical standards) designed for liquid chromatography were used: hydrochloric acid, formic acid, ethanol, and acetonitrile from Sigma Aldrich (Steinheim, Germany), and methanol from J.T. Baker Mallinckrodt Baker B.V. Holland. Buffers, ninhydrin, and a mixture of standards for amino acid identification were obtained from Sykam (Eresing, Germany), standardized for amino acid analyses in the physiological (native) range. Analytical standards of chlorogenic, caffeic, and ferulic acid for HPLC were obtained from Extrasynthese (Genay, France). Analytical standards D-(+)-glucose, D-(−)-fructose, D-(+)- sucrose, D-(+)- melezitose, D-(+)-turanose, D-(+)-trehalose, D-(+)-raffinose BioXtra, Ca, K, Mg, Fe, Cu, Mn, Zn, Al, Cd, and Pb were obtained from Sigma- Aldrich (Steinheim, Germany), and D-(+)-maltose standard was obtained from Toronto Research Chemicals. Carrez I solution, Carrez II solution (Sigma Aldrich, Steinheim, Germany), and deionised water from a deioniser from Hydrolab Polska HLP 5P were used.
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