Comprehensive Profiling of Free Proteinogenic and Non-Proteinogenic Amino Acids in Common Legumes Using LC-QToF: Targeted and Non-Targeted Approaches

Overview

Journal Foods

Specialty Biotechnology

Date 2025 Feb 26

PMID 40002056

Authors

Bharathi Avula

Kumar Katragunta

Iffat Parveen

Kiran Kumar Tatapudi

Amar G Chittiboyina

Yan-Hong Wang

Ikhlas A Khan

Affiliations

Soon will be listed here.

Abstract

Legumes, a dietary staple for centuries, have seen an influx of conventional and unconventional varieties to cater to human care conscious consumers. These legumes often undergo pretreatments like baking, soaking, or boiling to mitigate the presence of non-proteinogenic amino acids (NPAAs) and reduce associated health risks. The recent tara flour health scare, linked to the NPAA baikiain, emphasizes the need for robust analytical methods to ensure the safety and quality of both traditional and novel plant-based protein alternatives. While traditional techniques provide insights into protein and non-proteinogenic amino acid profiles, modern liquid chromatography-mass spectrometry (LC-MS) offers superior sensitivity and specificity for NPAA detection. This study employed an LC-QToF method with MS/MS analysis to comprehensively map the distribution of free NPAAs and proteinogenic amino acids (PAAs) in various legume samples. A total of 47 NPAAs and 20 PAAs were identified across the legume samples, with at least 7-14 NPAAs detected in each sample. Sulfur-containing NPAAs, such as -methyl--cysteine, γ-glutamyl--methyl cysteine, and -methyl homoglutathione, were predominantly found in and species. Cysteine and methionine were the sulfur-containing PAAs identified. Gel electrophoresis and soluble protein quantification were also conducted to understand legume protein composition holistically. This orthogonal approach provides a valuable tool for ensuring the overall quality of plant-based proteins and may aid in investigating food poisoning or outbreaks related to such products.

References

Joshi J, Saboori-Robat E, Solouki M, Mohsenpour M, Marsolais F . Distribution and possible biosynthetic pathway of non-protein sulfur amino acids in legumes. J Exp Bot. 2019; 70(16):4115-4121. DOI: 10.1093/jxb/erz291. View

Laemmli U . Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970; 227(5259):680-5. DOI: 10.1038/227680a0. View

Lane E . L-DOPA for Parkinson's disease-a bittersweet pill. Eur J Neurosci. 2018; 49(3):384-398. DOI: 10.1111/ejn.14119. View

Baldi G, Salamini F . Variability of essential amino acid content in seeds of 22 Phaseolus species. Theor Appl Genet. 2014; 43(2):75-8. DOI: 10.1007/BF00274960. View

Glover W, Baker T, Murch S, Brown P . Determination of β-N-methylamino-L-alanine, N-(2-aminoethyl)glycine, and 2,4-diaminobutyric acid in Food Products Containing Cyanobacteria by Ultra-Performance Liquid Chromatography and Tandem Mass Spectrometry: Single-Laboratory Validation. J AOAC Int. 2015; 98(6):1559-65. DOI: 10.5740/jaoacint.15-084. View

Krauss G . A selective staining test for the sensitive detection of some rare amino acids. J Chromatogr. 1972; 74(1):152-4. DOI: 10.1016/s0021-9673(01)94987-8. View

Gardner C, Hartle J, Garrett R, Offringa L, Wasserman A . Maximizing the intersection of human health and the health of the environment with regard to the amount and type of protein produced and consumed in the United States. Nutr Rev. 2019; 77(4):197-215. PMC: 6394758. DOI: 10.1093/nutrit/nuy073. View

Guba A, Baba O, Tozser J, Csosz E, Kallo G . Fast and Sensitive Quantification of AccQ-Tag Derivatized Amino Acids and Biogenic Amines by UHPLC-UV Analysis from Complex Biological Samples. Metabolites. 2022; 12(3). PMC: 8949038. DOI: 10.3390/metabo12030272. View

Bell E . Associations of ninhydrin-reacting compounds in the seeds of 49 species of Lathyrus. Biochem J. 1962; 83:225-9. PMC: 1243536. DOI: 10.1042/bj0830225. View

10.

Filee R, Schoos R, Boemer F . Evaluation of physiological amino acids profiling by tandem mass spectrometry. JIMD Rep. 2013; 13:119-28. PMC: 4110342. DOI: 10.1007/8904_2013_265. View

11.

Amico V, Oriente G, Tringali C . Quantitative gas-liquid chromatography of non-protein amino acids. J Chromatogr. 1976; 116(2):439-441. DOI: 10.1016/s0021-9673(00)89914-8. View

12.

Rozan P, Kuo Y, Lambein F . Amino acids in seeds and seedlings of the genus Lens. Phytochemistry. 2001; 58(2):281-9. DOI: 10.1016/s0031-9422(01)00200-x. View

13.

Li C, Brownson D, Mabry T, Perera C, Bell E . Nonprotein amino acids from seeds of Cycas circinalis and Phaseolus vulgaris. Phytochemistry. 1996; 42(2):443-5. DOI: 10.1016/0031-9422(95)00851-9. View

14.

Oh C, Kim J, Kim K, Mabry T . Rapid gas chromatographic screening of edible seeds, nuts and beans for non-protein and protein amino acids. J Chromatogr A. 1995; 708(1):131-41. DOI: 10.1016/0021-9673(95)00381-v. View

15.

Rao S, ADIGA P, Sarma P . THE ISOLATION AND CHARACTERIZATION OF BETA-N-OXALYL-L-ALPHA,BETA-DIAMINOPROPIONIC ACID: A NEUROTOXIN FROM THE SEEDS OF LATHYRUS SATIVUS. Biochemistry. 1964; 3:432-6. DOI: 10.1021/bi00891a022. View

16.

SULSER H, Sager F . Identification of uncommon amino acids in the lentil seed (Lens culinaris Med.). Experientia. 1976; 32(4):422-3. DOI: 10.1007/BF01920772. View

17.

Liao D, Cram D, Sharpe A, Marsolais F . Transcriptome Profiling Identifies Candidate Genes Associated with the Accumulation of Distinct Sulfur γ-Glutamyl Dipeptides in Phaseolus vulgaris and Vigna mungo Seeds. Front Plant Sci. 2013; 4:60. PMC: 3606967. DOI: 10.3389/fpls.2013.00060. View

18.

Bruckner H, Bosch I, Graser T, Furst P . Determination of alpha-alkyl-alpha-amino acids and alpha-amino alcohols by chiral-phase capillary gas chromatography and reverse-phase high-performance liquid chromatography. J Chromatogr. 1987; 395:569-90. DOI: 10.1016/s0021-9673(01)94145-7. View

19.

Choi H, Mun J, Kim D, Zhu H, Baek J, Mudge J . Estimating genome conservation between crop and model legume species. Proc Natl Acad Sci U S A. 2004; 101(43):15289-94. PMC: 524433. DOI: 10.1073/pnas.0402251101. View

20.

Bradford M . A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976; 72:248-54. DOI: 10.1016/0003-2697(76)90527-3. View