Detection of betacyanin in red-tube spinach (Spinacia oleracea) and its biofortification by strategic hydroponics.

Betacyanins have been reported as water-soluble, nitrogenous pigments found in the order Caryophyllales, and they are known for powerful natural antioxidant. The biofortification of secondary metabolites such as anthocyanins and betacyanins has recently been performed in food crops by metabolic engineering through genetic modification. However, the distribution of genetically modified foods is strictly regulated. Therefore, we aimed to develop a new method for biofortifying natural antioxidants, betacyanins, without genetic modification. We first detected the presence of betacyanins in red-tube spinach (Spinacia oleracea) through ultraviolet-visible spectroscopy and mass spectrometry. We then hydroponically cultivated plants in the presence of three candidate compounds for betacyanin biofortification: dopamine, Ca2+, and sucrose. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) and antioxidant activity analyses showed that sucrose was most successful in biofortifying betacyanins, and reverse transcription polymerase chain reaction (RT-PCR) indicated that several genes involved in betacyanin biosynthesis were induced by sucrose. Therefore, strategic hydroponics represents a new approach for cultivating betacyanin-enriched vegetables.

Folate Biofortification in Hydroponically Cultivated Spinach by the Addition of Phenylalanine.

Folate is an important vitamin mainly ingested from vegetables, and folate deficiency causes various health problems. Recently, several studies demonstrated folate biofortification in plants or food crops by metabolic engineering through genetic modifications. However, the production and sales of genetically modified foods are under strict regulation. Here, we developed a new approach to achieve folate biofortification in spinach (Spinacia oleracea) without genetic modification. We hydroponically cultivated spinach with the addition of three candidate compounds expected to fortify folate. As a result of liquid chromatography tandem mass spectrometry analysis, we found that the addition of phenylalanine increased the folate content up to 2.0-fold (306 µg in 100 g of fresh spinach), representing 76.5% of the recommended daily allowance for adults. By measuring the intermediates of folate biosynthesis, we revealed that phenylalanine activated folate biosynthesis in spinach by increasing the levels of pteridine and p-aminobenzoic acid. Our approach is a promising and practical approach to cultivate nutrient-enriched vegetables.