C50 CAROTENOIDS FROM THE CHEESE‐RIPENING BACTERIA ARTHROBACTER ARILAITENSIS
Résumé
Cheeses harbor a complex microbiota, usually a succession of different microorganisms during milk coagulation and ripening. Surface-ripened cheeses are characterized by a ripening from the cheese surface to the interior due to the activity of both yeasts and bacteria. Many such cheeses are produced all over the world with Europe as a leader: Taleggio in Italy; Munster, Epoisses, Livarot in France; Limburger, Tilsit in Germany; Herve in Belgium [1]. The cheese microflora contributes to a large extent to the development of the typical flavor, texture, and color of the final product. Color is one of the major attributes which affects the consumer perception of quality. Pigments generated by the smear bacteria at the surface of cheeses were poorly studied, exception made for those of the bacteria Brevibacterium linens (aromatic carotenoids, isorenieratene and hydroxyl derivatives). Indeed, for a long time this bacterium has been considered the single source of the color of red-smear ripened soft cheeses. High-throughput sequencing revolutionized the field of microbial ecology, allowing for a more accurate identification of microbial taxa, including those which are difficult to culture and/or are present in low abundance, now revealing the highly diverse bacterial populations of cheese rinds, i.e. with up to 19 bacterial genera described in some cheeses.
Arthrobacter bacteria have been isolated on surface-ripened cheeses and their pigments were found in cheese rinds [1]. The genus Arthrobacter is very complex in Nature with 80 species isolated from various environments and listed in the taxonomy, producing a great variety of pigments: orange and yellow (carotenoids –pioneering works in the early seventies [2]), blue and green (indigoidine, indochrome), red (porphyrins, carotenoids). The present study focuses on the chemical characterization of pigments biosynthesized by the cheese-ripening bacteria Arthrobacter arilaitensis.
Lyophilized biomass was extracted with 3 x MeOH and 2 x MTBE, with sonication and centrifugation at each step. Samples were analyzed using the same solvent system (Solvent A: MeOH; Solvent B: MTBE) with the gradient conditions described in [1]. A fused core column Ascentis Express Supelco C18, (150 x 4.6 mm, particle size 2.7 m) and a Shimadzu Nexera HPLC-PDA-IT TOF system were used for the HPLC analyses.
The UV-vis carotenoid profiles were similar for both strains investigated. Considering the UV-vis spectra (including fine structure), the APCI (-) and (+) mass spectra, and the elution order, 8 different carotenoids were tentatively identified. Four of them (sarcinaxanthin mono-glucoside pentaacetate, decaprenoxanthin di-glucoside, decaprenoxanthin mono-glucoside and decaprenoxanthin-C16:0) are possibly present in very low amount, and were only detected in the APCI (-) ionization mode. The other four free main carotenoids, decaprenoxanthin, sarcinaxanthin, 9-Z- decaprenoxanthin and 15-Z- decaprenoxanthin have quite clearly been characterized by both their UV-vis spectra and MS spectra in APCI (-) and (+) ionization mode. Arthrobacter arilaitensis species does indeed produce C50 carotenoids, and in particular, it mainly produces free all-E- decaprenoxanthin, and in minor amount two of its cis isomers, the 9-Z- decaprenoxanthin and 15-Z- decaprenoxanthin.
This is in complete accordance with genomic data published about A. arilaitensis (whole genome sequencing [3]) and its Idi, CrtE, CrtB, CrtI, CrtEb, CrtYe and CrtYf carotenogenesis cluster, similar to the cluster of the decaprenoxanthin producing Corynebacterium glutamicum.
[1]P. Galaup, A. Gautier, Y. Piriou, A. de Villeblanche, A. Valla, and L. Dufossé. Innov. Food Sci. Emerg. Technol., 8, 373-378 (2007).
[2]N. Arpin, S. Liaaen-Jensen, and M. Trouilloud. Acta Chem. Scand., 26, 2524-2526 (1972).
[3]C. Monnet et al. PLoS ONE, 5, e15489 (2010).
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