Characterisation, health-promoting properties and food applications of anthocyanin-rich pigments of flowers from the Geraniaceae and Lamiaceae plant families

Colour is an important characteristic of food and plays a significant role in enhancing the visual appeal of food products. The colourants mostly used in the food industry are synthetic azo-dyes due to their brightness and stability. However, scientific evidence has shown that these compounds may pose adverse health effects. This has created a demand for more natural food colourants by consumers who have become more health conscious. Plant species from the Geraniaceae (Pelargonium) and Lamiaceae (Salvia (sage) and Plectranthus (spurflower)) families produce flowers with a range of different colours. The flowers are non-toxic and could be a source of novel stable pigments for potential application as natural colourants in foods with added health benefits. Plant species from the Geraniaceae (Pelargonium grandiflorum Willd., P. × hortorum L.H. Bailey and Pelargonium zonale (L.) L’Hér.) and Lamiaceae (Salvia aurea × dolomitica, Salvia dolomitica Codd and Plectranthus zuluensis T. Cooke) families were used in this research. The petals of their flowers were collected, air-dried at 30ºC for 24 hours and ground into a powder. Pigmented extracts were prepared by extracting the flower powder with formic acid-acidified water for 24 hours. The pigmented flower extracts were evaluated for total phenolic content (Folin-Ciocalteu assay), anthocyanin and phenolic profiles and concentrations of phenolic compounds (Ultra Performance Liquid Chromatography-Quadropole Time-of-Flight Mass Spectrometry). Degradation kinetics experiments were conducted to determine the thermal and oxidative stability of the pigmented flower extracts. Health-promoting properties of the pigmented flower vi | P a g e extracts were determined using chemical radical scavenging assays (2,2-diphenyl-1-picrylhydrazyl [DPPH] radical scavenging, oxygen radical absorption capacity [ORAC] and nitric oxide [NO] radical scavenging), cellular assays (Caco-2 cellular antioxidant activity, NO radical scavenging in RAW 264.7 macrophages and anti-adipogenic activity in 3T3-L1 adipocytes) and anti-glycation activity (Advanced Glycation End Products assay). The pigmented flower extracts were applied as natural colourants in yogurt, fondant and gelatine candies while the flower petal powder was applied directly as colourant in yogurt. The food products with the natural colourants were evaluated for their colour stability (chromaticity analysis) during storage. Anthocyanins in the Geraniaceae pigments were mainly 3,5-diglucoside of delphinidin, petunidin, pelargonidin, peonidin and malvidin and acetyl-acylated malvidin, delphinidin and petunidin. Anthocyanins in the Lamiaceae pigments were rutinosides of pelargonidin, glucosides of petunidin, pelargonidin and p-coumaric acid- and malonyl-acylated delphinidin and malvidin. Geraniaceae pigments also contained rutinosides of kaempferol and quercetin, quercetin-3-O-glucoside (isoquercetin), luteolin-7-O-glucoside and hydrolysable tannins (β-glucogallin, penta and tetra-galloyl glucose). Lamiaceae pigments contained naringin and hesperidin. The only phenolic acid in the Geraniaceae pigments was p-coumaric acid and the Lamiaceae pigments contained rosmarinic acid and its derivatives such as sagerinic acid and salvianolic acid B. Lamiaceae pigments were more thermally and oxidatively stable (lower degradation rate constants and longer half-life) than Geraniaceae pigments. The greater stability of the Lamiaceae pigments could be due to hydrophobic intramolecular self-association interactions of their aromatic acylated anthocyanins in combination with stabilising interactions of these aromatic acylated anthocyanins with rosmarinic acid and its derivatives. The pigmented flower extracts from the Geraniaceae had higher total phenolic content and DPPH radical scavenging activity than the Lamiaceae. All pigmented flower extracts exhibited Caco-2 cellullar antioxidant activity (reduction of oxidative stress), inhibition of LPS-induced NO production in RAW 267.4 macrophage cells (anti-inflammatory effects) and ability to reduce lipid accumulation in 3T3-L1 adipocytes after 14 days treatment (anti-obesity effects). Despite their lower total phenolic content and DPPH radical scavenging activity, Lamiaceae pigments showed similar Caco-2 cellular antioxidant activity and inhibition of NO production in RAW 267.4 macrophages to Geraniaceae pigments. This may be related to the observed vii | P a g e greater thermal and oxidative stability of the Lamiaceae pigments. P. grandiflorum and P. × hortorum pigmented flower extracts showed significant anti-glycation activity (anti-diabetic effects) which was notably similar to that of the anti-diabetic drug aminoguanidine. The pigmented flower extracts were successfully applied as natural colourants in yogurt (low pH food), fondant (sugar-based food system) and gelatine (protein-based food system) in comparison with the commercial E163 anthocyanin colourant. The colour of the food products was stable over a 15-day storage period with only some indications of slight decreases in redness and total colour. Compared to the commercial E163 colourant which only gave a purple colour, the pigmented flower extracts imparted a greater range of colours to the food products (red, purple, pink and orange) and in different shades of intensity and brightness. The application of the powdered flower samples directly into yogurt as colourant led to development of brown colouration in the yogurt possibly due to residual activity of browning enzymes in the flowers. This indicates that direct incorporation of the powdered flower samples as colourants in foods may not always be desirable. This research shows the potential of flowers from the Geraniaceae and Lamiaceae plant families for application as novel natural food colourants. It represents important innovations in natural food colourants and provides novelty in terms of the stability of the pigments, the variety of colours they confer to food systems and their health-promoting properties. The findings of this study open up new opportunities for production of natural colourants for application in the food industry and also meets consumer demands.