A liquid chromatography-electrospray ionization-mass spectrometry method for quantification of cyclotides in plants avoiding sorption during sample preparation

• Cyclotides are plant-produced, bioactive, cyclic mini-proteins with interesting pharmaceutical and agricultural applications. A reverse phase liquid chromatography electrospray ionization mass spectrometry (RP-LC-ESI-MS) method for analysis of cyclotides in plant materials with a minimum of sample pretreatment is presented. Three exemplary cyclotides (kalata B1, kalata B2 and cycloviolacin 02) were used as reference substances for the method development. Linearity (r(2) > 0.99) was achieved in the concentration range 0.05-10 mg/L and the limit of detection was 1.7-4.0 mu g/L. The present study is the first to demonstrate that cyclotides dissolved in water sorb to glass vials, but the addition of 15% of acetonitrile or 40 mg/L of bovine serum albumin is sufficient to keep the cyclotides in solution. Cyclotides were extracted from candied violets, violet tea, and the plants Oldenlandia affinis and Viola odorata using 70% methanol containing 0.1% formic acid (v/v). The plant content was determined to be 23.5-14,200 mu g/g (dry weight). The highest content of cyclotide was found in wild Danish V. odorata, and it is the highest content of cyclotide in a plant reported hitherto. Candied violets contained 0.00-8.66 mu g/g (dry weight), while no cyclotides were detected in commercial violet tea.

Ämnesord

NATURVETENSKAP  -- Kemi (hsv//swe)

NATURAL SCIENCES  -- Chemical Sciences (hsv//eng)

Nyckelord

Cyclotides

LC-MS

Quantification

Sorption

Adducts

Proteins

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