Analysis of Plasma Amino Acids Using RP-HPLC and Pre-Column Derivatization with OPA/3-MPA

Abstract

Background and Objective: Measurement of amino acids is an important tool for metabolic studies and evaluation of patients' clinical condition. The aim of this study was to analyze the plasma amino acids using reverse phase high performance liquid chromatography techniques (RP-HPLC) with pre-column derivatization by o-phthalaldehyde (OPA) in combination with 3-mercaptopropionic acid (3-MPA). Methods: Overall, 107 neonates and babies suspected of having metabolic disorder were enrolled in this study. The level of amino acids in plasma samples was analyzed within 65 minutes by HPLC with pre-column derivatization by OPA/3-MPA. This was a gradient RP-HPLC method that was performed using two solvents with a ratio of methanol and sodium acetate. L-norvaline internal standard was used as the reference peak for amino acids. Standard mixture of amino acids was used to determine the concentrations of amino acids. Results: According to the values of coefficient of variation obtained for each amino acid, the results indicated a good chromatographic separation of amino acids by this method. The use of OPA/3-MPA derivative reagent increased the efficiency and resolution of amino acids chromatographic separation. Conclusion: Due to simple preparation and accurate assessment, determination of plasma amino acids using OPA/3-MPA derivatives and RP-HPLC is a suitable method in many clinical samples.

Metabolites and Semen Characteristics in Different Bulls Fertility

“…for total antioxidant capacity، malondialdehyde concentration، and amino، fatty، and carboxylic acid determinations. Determination of amino، fatty، and carboxylic acids concentrations: Some amino acids concentration (alanine، glycine، leucine، serine، proline، asparagine، aspartic، methionine، glutamic، tryptophan، and lysine) in the seminal plasma were estimated using high-performance liquid chromatography (1).===== The fat seminal plasma extraction was calculated using the AOAC method (13). Some fatty acid and carboxylic acid concentrations (acetic، butyric، propanoic، palmitic، oleic، linoleic، stearic، linolenic، arachidic، myristic، rvonic، cisdocosadienoic، tricosanoic، erucic، undecanoic) in seminal plasma were estimated using gas chromography (GC-2010، 17)..…”

Effect of Sperms Freezability on Some Semen Attributes and Amino Acid Concentrations in Seminal Plasma of Holstein Bulls

“…Determination of amino acids concentrations in seminal plasma: The concentration of amino acids in the seminal plasma was estimated using high-performance liquid chromatography HPLC). Concurrently, the concentration of essential amino acids (tryptophan, lysine, threonine, valine, methionine) and non-essential amino acids (serine, glutamine, glycine, arginine, and alanine and cysteine and aspartic) were calculated based on the method of Mohammad et al (25).…”

Effect of N- carbamylglutamate on amino acids and blood biochemistry in young male goats

“…A high-performance liquid chromatography method with pre-column derivatization for separation and quantification of amino acids in plasma samples were conducted using an amino acid analyzer, model YL9100 Korea, the mobile phase was acetonitrile: buffer: DW (60: 10: 30), a Flow rate (1ml/min) and column separation was C18-NH2 (25 cm * 4.6 mm), the Detector = Florescence Ex =365 nm, Em = 445 nm (Mohammad et al, 2016).…”