High-Throughput Quantitation of Proline Betaine in Foods and Suitability as a Valid Biomarker for Citrus Consumption

Abstract

Proline betaine has been proposed as a candidate dietary biomarker for citrus intake. To validate its suitability as a dietary biomarker and to gain insight into the range of this per-methylated amino acid in foods and beverages, a quick and accurate stable isotope dilution assay was developed for quantitative high-throughput HILIC-MS/MS screening of proline betaine in foods and urine after solvent-mediated matrix precipitation. Quantitative analysis of a variety of foods confirmed substantial amounts of proline betaine in citrus juices (140-1100 mg/L) and revealed high abundance in tubers of the vegetable Stachys affinis, also known as Chinese artichocke (∼700 mg/kg). Seafood including clams, shrimp, and lobster contained limited amounts (1-95 mg/kg), whereas only traces were detected in fish, cuttlefish, fresh meat, dairy products, fresh vegetable (<3 mg/kg), coffee, tea, beer, and wine (<7 mg/L). The human excretion profiles of proline betaine in urine were comparable when common portions of orange juice or fried Stachys tubers were consumed. Neither mussels nor beer provided enough proline betaine to detect significant differences between morning urine samples collected before and after consumption. As Stachys is a rather rare vegetable and not part of peoples' daily diet, the data reported here will help to monitor the subject's compliance in future nutritional human studies on citrus products or the exclusion of citrus products in the wash-out phase of an intervention study. Moreover, proline betaine measurement can contribute to the establishment of a toolbox of valid dietary biomarkers reflecting wider aspects of diet to assess metabolic profiles as measures of dietary exposure and indicators of dietary patterns, dietary changes, or effectiveness of dietary interventions.

Diet‐Related Metabolites Associated with Cognitive Decline Revealed by Untargeted Metabolomics in a Prospective Cohort

“…Proline betaine, a well‐validated biomarker of citrus intake, was another important food‐derived metabolite found associated with greater CD in this study. This observation is in agreement with a smaller metabolomics study that identified proline betaine among the increased serum metabolites in AD patients, Yet, it contrasts with the reported beneficial role of citrus fruit, which are good sources of antioxidant and vasculoprotective nutrients (e.g., vitamin C and flavanones) .…”

The biology of ergothioneine, an antioxidant nutraceutical

“…The first were normal cells, that, as with many transporters (119) , do not in fact express SLC22A4 at significant levels, while the second had been engineered to overexpress the transporter. They then simply looked for those molecules that were most differentially taken up, a molecule called stachydrine, also known as proline betaine, being the main one observed, Stachydrine is a constituent of citrus juices (165)(166)(167) . Some elementary cheminformatics based on structure similarity searches (57,168) indicated that ERG, as a betaine, was indeed similar to stachydrine.…”

Control of metabolite efflux in microbial cell factories: current advances and future prospects

“…[201][202][203][204][205]) be referred to as 'untargeted metabolomics' to determine the differential uptake of substances from pooled serum when it was incubated with HEK293 cells either lacking measurable amounts, or containing cloned-up levels, of SLC22A4. A specific mass of m/z 144.84 was detected as being particularly taken up in the transporter-containing cells, and this was identified as the dipeptide proline betaine (aka stachydrine), a characteristic constituent of citrus fruits and their juices [206][207][208]. Gründemann and colleagues [184] then performed some elementary cheminformatics (as in [209]) to look for other molecules that were chemically (structurally) similar to stachydrine, recognising that among them was ergothioneine (also known as 2-mercaptohistidine trimethylbetaine; IUPAC name (2S)-3-(2-Thioxo-2,3-dihydro-1H-imidazol-4-yl)-2-(trimethylammonio)propanoate) (Fig 4).…”