C50 Carotenoids Research Articles

Genomewide characterisation of the genetic diversity of carotenogenesis in bacteria of the order Sphingomonadales.

The order Sphingomonadales is a taxon of bacteria with a variety of physiological features and carotenoid pigments. Some of the coloured strains within this order are known to be aerobic anoxygenic phototrophs that contain characteristic photosynthesis gene clusters (PGCs). Previous work has shown that majority of the ORFs putatively involved in the biosynthesis of C40 carotenoids are located outside the PGCs in these strains. The main purpose of this study was to understand the genetic basis for the various colour/carotenoid phenotypes of the strains of Sphingomonadales. Comparative analyses of the genomes of 41 strains of this order revealed that there were different patterns of clustering of carotenoid biosynthesis (crt) ORFs, with four ORF clusters being the most common. The analyses also revealed that co-occurrence of crtY and crtI is an evolutionarily conserved feature in Sphingomonadales and other carotenogenic bacteria. The comparisons facilitated the categorisation of bacteria of this order into four groups based on the presence of different crt ORFs. Yellow coloured strains most likely accumulate nostoxanthin, and contain six ORFs (group I: crtE, crtB, crtI, crtY, crtZ, crtG). Orange coloured strains may produce adonixanthin, astaxanthin, canthaxanthin and erythroxanthin, and contain seven ORFs (group II: crtE, crtB, crtI, crtY, crtZ, crtG, crtW). Red coloured strains may accumulate astaxanthin, and contain six ORFs (group III: crtE, crtB, crtI, crtY, crtZ, crtW). Non-pigmented strains may contain a smaller subset of crt ORFs, and thus fail to produce any carotenoids (group IV). The functions of many of these ORFs remain to be characterised.

Zechstein Main Dolomite oil characteristics in the Southern Permian Basin: I. Polish and German sectors

Geochemical analyses were used to classify 39 Zechstein (Late Permian, Lopingian) Main Dolomite (Ca2) crude oil samples from fields in the eastern and southern sector of the Southern Permian Basin (SPB) of Europe and to provide new insights into the origin of the oil. Geochemical data indicate that Ca2 oils were generated in the early-to-late oil window and are mostly non-waxy oils. Various biomarker and stable carbon isotopic ratios were used to identify source and depositional settings for source rocks of Ca2 oils arranged within 10 distinct oil groups. Specifically, the geochemical analyses and oil-oil correlations revealed a set of characteristic biomarkers including an even-over-odd predominance (EOP) for the C20-30 n-alkanes, C40 carotenoid occurrence (isorenieratane, chlorobactane, β-isorenieratane), bisnorhopane/hopane (BNH/H) ratios >0.1, high abundances of C35 homohopanes and elevated concentrations of C32 and C34 homohopanes, a predominance of C29 homologues among 4-desmethyl steranes in the majority of oil samples, and a high abundance of diasteranes. Stable carbon isotopes and biomarkers provided ample evidence that Ca2 oils were generated from predominantly algal-rich marly carbonate/evaporite source rocks located in the lower slope/shallow-basin and lagoonal facies of the Ca2 basin, all deposited under suboxic-anoxic (euxinic) conditions. In the case of all higher maturity oils, the source rocks could not be reliably identified but high (>2) C24Tet/C23 values suggest a carbonate-evaporite depositional setting.

Carotenoid Production by Recombinant Corynebacterium glutamicum: Strain Construction, Cultivation, Extraction, and Quantification of Carotenoids and Terpenes.

Corynebacterium glutamicum is a workhorse of industrial amino acid production employed for more than five decades for the million-ton-scale production of L-glutamate and L-lysine. This bacterium is pigmented due to the biosynthesis of the carotenoid decaprenoxanthin. Decaprenoxanthin is a carotenoid with 50 carbon atoms, and, thus, C. glutamicum belongs to the rare group of bacteria that produce long-chain C50 carotenoids. C50 carotenoids have been mainly isolated from extremely halophilic archaea (Kelly and Jensen, Acta Chem Scand 21:2578, 1967; Pfander, Pure Appl Chem 66:2369-2374, 1994) and from Gram-positive bacteria of the order Actinomycetales (Netzer et al., J Bacteriol 192:5688-5699, 2010). The characteristic yellow phenotype of C. glutamicum is due to the cyclic C50 carotenoid decaprenoxanthin and its glycosides. Decaprenoxanthin production has been improved by plasmid-borne overexpression of endogenous genes of carotenogenesis. Gene deletion resulted in the production of the C40 carotenoid lycopene, an intermediate of decaprenoxanthin biosynthesis. Heterologous gene expression was required to develop strains overproducing nonnative carotenoids and terpenes, such as astaxanthin (Henke et al., Mar Drugs 14:E124, 2016) and (+)-valencene (Frohwitter et al., J Biotechnol 191:205-213, 2014). Integration of additional copies of endogenous genes expressed from strong promoters improved isoprenoid biosynthesis. Here, we describe C. glutamicum strains, plasmids, and methods for overexpression of endogenous and heterologous genes, gene deletion, replacement, and genomic integration. Moreover, strain cultivation as well as extraction, identification, and quantitative determination of terpenes and carotenoids produced by C. glutamicum is detailed.

Complete genome sequence of Planococcus faecalis AJ003T, the type species of the genus Planococcus and a microbial C30 carotenoid producer

A novel type strain, Planococcus faecalis AJ003T, isolated from the feces of Antarctic penguins, synthesizes a rare C30 carotenoid, glycosyl-4,4ʹ-diaponeurosporen-4ʹ-ol-4-oic acid. The complete genome of P. faecalis AJ003T comprises a single circular chromosome (3,495,892 bp; 40.9% G + C content). Annotation analysis has revealed 3511 coding DNA sequences and 99 RNAs; seven genes associated with the MEP pathway and five genes involved in the carotenoid pathway have been identified. The functionality and complementation of 4,4ʹ-diapophytoene synthase (CrtM) and two copies of heterologous 4,4ʹ-diapophytoene desaturase (CrtN) involved in carotenoid biosynthesis were analyzed in Escherichia coli.

Complete genome sequence of Flavobacterium kingsejongi WV39, a type species of the genus Flavobacterium and a microbial C40 carotenoid zeaxanthin producer

A novel species, Flavobacterium kingsejongi WV39, isolated from feces of Antarctic penguins and a type species of the genus Flavobacterium, is yellow because it synthesizes a C40 carotenoid zeaxanthin. The complete genome of F. kingsejongi WV39 is made up of a single circular chromosome (4,224,053bp, 39.8% G+C content). Annotation analysis revealed 3,955 coding sequences, 72 RNAs (18 rRNA+54 tRNA), and five genes involved in zeaxanthin biosynthesis. The key gene encoding β-carotenoid hydroxylase (CrtZ), which is the last enzyme in the zeaxanthin biosynthetic pathway, was cloned and subjected to complementary analysis in a heterologous E. coli strain. The CrtZ of F. kingsejongi WV39 showed a higher activity than other reported CrtZs.

Ultrafast Excited-State Dynamics of all-trans-Capsanthin in Organic Solvents.

The C40 carotenoid capsanthin is of photophysical interest because it belongs to the family of terminally carbonyl-substituted apocarotenes. These have the potential to exhibit intramolecular charge transfer (ICT) character in the excited state. We studied its ultrafast dynamics in different solvents using broadband transient absorption spectroscopy in the 260-1600 nm range. Photoexcitation initially populated the S2(11Bu+) state which decayed to the S1(21Ag-) state in 120-150 fs. The lifetime of the S1 state decreased from 10.3 ps (isooctane) to 9.1 ps (methanol). Together with the absence of stimulated emission this suggested a weak ICT character of S1. A steric influence of the five-membered ring in capsanthin was identified based on a comparison with the sister apocarotenone citranaxanthin which features methyl substitution at the keto group. While both apocarotenones exhibit the same S1 lifetime in isooctane, the decrease in lifetime in polar solvents is weaker for capsanthin because presumably the five-membered ring sterically perturbs stabilization of ICT character by the solvent. For the S1 state of capsanthin we further observed intramolecular vibrational redistribution and collisional energy transfer to the solvent with time constants of 340-420 fs and 8.5-9.1 ps, respectively. No evidence for excited-state photoisomerization of capsanthin was found.

Synthesizing value-added products from methane by a new Methylomonas.

Methane and methanol are potential carbon sources of industrial micro-organisms in addition to crop-derived bio-carbon sources. Methanotrophs that can utilize these simple, stable and large amounts chemicals are expected to be developed into 'cell factories' for the production of specific chemicals. In this study, a methanotroph that can synthesize lycopene, C30 carotenoid and exopolysaccharides (EPS) with relative better performances from C1 substrates was isolated, and its performances were evaluated. The isolated strain was identified as Methylomonas sp. ZR1 based on 16S rRNA sequence analysis. Its maximum specific growth rate achieved 0·200h-1 under flask culture conditions, and 0·386h-1 in bubble column reactors. ZR1 was able to utilize 35gl-1 of methanol and even exhibited slight growth in the presence of 40gl-1 of methanol. Furthermore, ZR1 was proved to synthesize lycopene (C40 carotenoids) besides the C30 carotenoids through HPLC-DAD and HPLC-MS/MS analysis methods. Its carotenoid extracts exhibited excellent antioxidative activities measured by the ABTS+ method. Plenty of polysaccharides were also synthesized by ZR1, the components of the polysaccharides were identified as glucose, mannose and galactose with a proportion of 1:2:1 by GC-MS, and its yield achieved 0·13gg-1 cell dry weight. The isolated strain has great potential for the production of value-added bioproducts from C1 compounds because of its excellent C1 substrate utilizing abilities and its abilities to naturally synthesize lycopene, C30 carotenoids and EPS. In this study, we isolated a fast-growing methanotroph, its C1 carbon substrate utilizing ability is excellent in comparison with reported methanotrophs. Furthermore, besides polysaccharides and C30 carotenoids which were commonly synthesized by methanotrophs, our findings suggested that C40 lycopene could also be naturally synthesized from methane by methanotrophs.

In Vitro Antioxidant, Antihemolytic, and Anticancer Activity of the Carotenoids from Halophilic Archaea.

Halophilic archaea represent a promising natural source of carotenoids. However, little information is available about the biological effects of carotenoids from halophilic archaea. In this study, the carotenoids produced by seven halophilic archaeal strains Halogeometricum rufum, Halogeometricum limi, Haladaptatus litoreus, Haloplanus vescus, Halopelagius inordinatus, Halogranum rubrum, and Haloferax volcanii were identified by ultraviolet/visible spectroscopy, thin-layer chromatography, and high-performance liquid chromatography-tandem mass spectrometry. The C50 carotenoids bacterioruberin and its derivatives monoanhydrobacterioruberin and bisanhydrobacterioruberin were found to be the predominant carotenoids. The antioxidant capacities of the carotenoids from these strains were significantly higher than β-carotene as determined by 1,1-diphenyl-2-picrylhydrazyl radical scavenging assay. The antihemolytic activities of these carotenoid extracts against H2O2-induced hemolysis in mouse erythrocytes were 3.9-6.3 times higher than β-carotene. A dose-dependent in vitro antiproliferative activity against HepG2 cells was observed for the extract from Hgm. limi, while that from Hpn. vescus exhibited a relatively high activity in a dose-independent manner. These results suggested that halophilic archaea could be considered as an alternative source of natural carotenoids with high antioxidant, antihemolytic, and anticancer activity.

Juice volatile composition differences between Valencia orange and its mutant Rohde Red Valencia are associated with carotenoid profile differences

A targeted approach using HS-SPME-GC–MS was performed to investigate volatile compounds of ordinary Valencia (VAL) and its more deeply colored mutant Rohde Red Valencia orange (RRV) at different developmental stages. Fifty-six volatile components classified into six chemical groups were quantified. The individual volatile compounds in each group were significantly influenced by the genotype and harvest date. Cluster analysis suggested two principal clusters. Cluster I included fruits of VAL and RRV harvested in July, September, and November and was characterized by the 2-ethyl-1-hexanol acetate and linalool chemotype; cluster II included fruits harvested in January and March, with the valencene/β-myrcene chemotype. Principal component analysis confirmed that fruits of both cultivars harvested at different dates possessed a unique aroma active profile, especially, the odor-active volatile-norisoprenoids degraded from C40 carotenoid were significantly higher in RRV than in VAL at full maturity. These results could provide information for further study of the flavor metabolism pathways in the two closely related cultivars.

Coproduction of cell-bound and secreted value-added compounds: Simultaneous production of carotenoids and amino acids by Corynebacterium glutamicum

Corynebacterium glutamicum is used for production of the food and feed amino acids l-glutamate and l-lysine at the million-ton-scale. One feed formulation of l-lysine simply involves spray-drying of the fermentation broth, thus, including secreted l-lysine and C. glutamicum cells which are pigmented by the C50 carotenoid decaprenoxanthin. C. glutamicum has been engineered for overproduction of various compounds including carotenoids. In this study, C. glutamicum was engineered for coproduction of a secreted amino acid with a cell-bound carotenoid. Asa proof of principle, coproduction of l-glutamate with the industrially relevant astaxanthin was shown. This strategy was applied to engineer l-lysine overproducing strains for combined overproduction of secreted l-lysine with the cell-bound carotenoids decaprenoxanthin, lycopene, β-carotene, zeaxanthin, canthaxanthin and astaxanthin. By fed-batch fermentation 48g/Ll-lysine and 10mg/L astaxanthin were coproduced. Moreover, C. glutamicum was engineered for coproduction of l-lysine and β-carotene from xylose and arabinose as alternative feedstocks.

Carotenoids from the extreme halophilic archaeon Haloterrigena turkmenica: identification and antioxidant activity.

Haloterrigena turkmenica was able to synthesize carotenoids when grown in halobacteria medium. These molecules have antioxidant properties and find application in food, cosmetic, and pharmaceutical fields. The carotenoids were extracted with methanol, separated by RP-HPLC, and identified by mass spectrometry and UV/Vis spectra analyses. The C50 carotenoids were the main pigments, and C30, C40, and C51 carotenoids were also detected. Seven geometric isomers were distinguished for bacterioruberin, monoanhydrobacterioruberin, and bisanhydrobacterioruberin. The assignment to a specific isomer was tentatively attempted through the analysis of the corresponding UV/Vis spectrum, the intensity of the cis peak, and its spectral fine structure. Lycopene, phytoene, and lycopersene were among the minor carotenoids further identified. The extract displayed antioxidant power higher than alpha-tocopherol, butylhydroxytoluene, and ascorbic acid used as reference compounds. Our studies identified for the first time seven geometric isomers of bacterioruberin derivatives and 30 carotenoids in a haloarchaeon.

Isoprenoid Pyrophosphate-Dependent Transcriptional Regulation of Carotenogenesis in Corynebacterium glutamicum.

Corynebacterium glutamicum is a natural producer of the C50 carotenoid decaprenoxanthin. The crtEcg0722crtBIYEb operon comprises most of its genes for terpenoid biosynthesis. The MarR-type regulator encoded upstream and in divergent orientation of the carotenoid biosynthesis operon has not yet been characterized. This regulator, named CrtR in this study, is encoded in many actinobacterial genomes co-occurring with terpenoid biosynthesis genes. CrtR was shown to repress the crt operon of C. glutamicum since DNA microarray experiments revealed that transcript levels of crt operon genes were increased 10 to 70-fold in its absence. Transcriptional fusions of a promoter-less gfp gene with the crt operon and crtR promoters confirmed that CrtR represses its own gene and the crt operon. Gel mobility shift assays with purified His-tagged CrtR showed that CrtR binds to a region overlapping with the −10 and −35 promoter sequences of the crt operon. Isoprenoid pyrophosphates interfered with binding of CrtR to its target DNA, a so far unknown mechanism for regulation of carotenogenesis. The molecular details of protein-ligand interactions remain to be studied. Decaprenoxanthin synthesis by C. glutamicum wild type was enhanced 10 to 30-fold upon deletion of crtR and was decreased 5 to 6-fold as result of crtR overexpression. Moreover, deletion of crtR was shown as metabolic engineering strategy to improve production of native and non-native carotenoids including lycopene, β-carotene, C.p. 450 and sarcinaxanthin.

Overexpression of the primary sigma factor gene sigA improved carotenoid production by Corynebacterium glutamicum: Application to production of β-carotene and the non-native linear C50 carotenoid bisanhydrobacterioruberin

Corynebacterium glutamicum shows yellow pigmentation due to biosynthesis of the C50 carotenoid decaprenoxanthin and its glycosides. This bacterium has been engineered for production of various non-native cyclic C40 and C50 carotenoids such as β-carotene, astaxanthin or sarcinaxanthin. In this study, the effect of modulating gene expression more broadly by overexpression of sigma factor genes on carotenoid production by C. glutamicum was characterized. Overexpression of the primary sigma factor gene sigA improved lycopene production by recombinant C. glutamicum up to 8-fold. In C. glutamicum wild type, overexpression of sigA led to 2-fold increased accumulation of the native carotenoid decaprenoxanthin in the stationary growth phase. Under these conditions, genes related to thiamine synthesis and aromatic compound degradation showed increased RNA levels and addition of thiamine and the aromatic iron chelator protocatechuic acid to the culture medium enhanced carotenoid production when sigA was overexpressed. Deletion of the gene for the alternative sigma factor SigB, which is expected to replace SigA in RNA polymerase holoenzymes during transition to the stationary growth phase, also increased carotenoid production. The strategy of sigA overexpression could be successfully transferred to production of the non-native carotenoids β-carotene and bisanhydrobacterioruberin (BABR). Production of the latter is the first demonstration that C. glutamicum may accumulate a non-native linear C50 carotenoid instead of the native cyclic C50 carotenoid decaprenoxanthin.

Carotenoids Database: structures, chemical fingerprints and distribution among organisms.

To promote understanding of how organisms are related via carotenoids, either evolutionarily or symbiotically, or in food chains through natural histories, we built the Carotenoids Database. This provides chemical information on 1117 natural carotenoids with 683 source organisms. For extracting organisms closely related through the biosynthesis of carotenoids, we offer a new similarity search system ‘Search similar carotenoids’ using our original chemical fingerprint ‘Carotenoid DB Chemical Fingerprints’. These Carotenoid DB Chemical Fingerprints describe the chemical substructure and the modification details based upon International Union of Pure and Applied Chemistry (IUPAC) semi-systematic names of the carotenoids. The fingerprints also allow (i) easier prediction of six biological functions of carotenoids: provitamin A, membrane stabilizers, odorous substances, allelochemicals, antiproliferative activity and reverse MDR activity against cancer cells, (ii) easier classification of carotenoid structures, (iii) partial and exact structure searching and (iv) easier extraction of structural isomers and stereoisomers. We believe this to be the first attempt to establish fingerprints using the IUPAC semi-systematic names. For extracting close profiled organisms, we provide a new tool ‘Search similar profiled organisms’. Our current statistics show some insights into natural history: carotenoids seem to have been spread largely by bacteria, as they produce C30, C40, C45 and C50 carotenoids, with the widest range of end groups, and they share a small portion of C40 carotenoids with eukaryotes. Archaea share an even smaller portion with eukaryotes. Eukaryotes then have evolved a considerable variety of C40 carotenoids. Considering carotenoids, eukaryotes seem more closely related to bacteria than to archaea aside from 16S rRNA lineage analysis. Database URL: http://carotenoiddb.jp

Production of the Marine Carotenoid Astaxanthin by Metabolically Engineered Corynebacterium glutamicum.

Astaxanthin, a red C40 carotenoid, is one of the most abundant marine carotenoids. It is currently used as a food and feed additive in a hundred-ton scale and is furthermore an attractive component for pharmaceutical and cosmetic applications with antioxidant activities. Corynebacterium glutamicum, which naturally synthesizes the yellow C50 carotenoid decaprenoxanthin, is an industrially relevant microorganism used in the million-ton amino acid production. In this work, engineering of a genome-reduced C. glutamicum with optimized precursor supply for astaxanthin production is described. This involved expression of heterologous genes encoding for lycopene cyclase CrtY, β-carotene ketolase CrtW, and hydroxylase CrtZ. For balanced expression of crtW and crtZ their translation initiation rates were varied in a systematic approach using different ribosome binding sites, spacing, and translational start codons. Furthermore, β-carotene ketolases and hydroxylases from different marine bacteria were tested with regard to efficient astaxanthin production in C. glutamicum. In shaking flasks, the C. glutamicum strains developed here overproduced astaxanthin with volumetric productivities up to 0.4 mg·L−1·h−1 which are competitive with current algae-based production. Since C. glutamicum can grow to high cell densities of up to 100 g cell dry weight (CDW)·L−1, the recombinant strains developed here are a starting point for astaxanthin production by C. glutamicum.

4,4'-diaponeurosporene, a C30 carotenoid, effectively activates dendritic cells via CD36 and NF-κB signaling in a ROS independent manner.

Carotenoids could be divided into C30 carotenoids and C40 carotenoids. The immune functions of C40 carotenoids had been extensively researched, while those of C30 carotenoids still remain unclear. In this study, the immune functions of a biosynthetic C30 carotenoid, 4,4′-diaponeurosporene (Dia), were identified on dendritic cells (DCs). DCs treated with 1 μM Dia for 24 h showed morphologic and phenotypic characteristics of mature state and had an increased production of IL-6, IL-10, IL-12p70 and TNFα, while β-carotene had a suppressive effect on DCs maturation. Moreover, Dia promoted antigen uptake of DCs in vitro and increased the quantity of antigen loaded DCs in mesenteric lymph nodes (MLN). Dia-treated DCs also had an enhanced ability to stimulate T cell proliferation and Th1 polarization. Further researches showed that Dia activated DCs via CD36 as well as ERK, JNK, and NF-κB signals in a reactive oxygen species (ROS) independent manner.

Complete genome sequence of carotenoid-producing Microbacterium sp. strain PAMC28756 isolated from an Antarctic lichen

Microbacterium sp. strain PAMC28756, of the family Microbacteriaceae, was isolated from Stereocaulon sp., an Antarctic lichen. Complete genome sequencing of Microbacterium sp. PAMC28756 revealed, for the first time in the genus Microbacterium, a series of key genes involved in C50 carotenoid biosynthesis. An analysis of the Microbacterium sp. PAMC28756 genome will lead to a better understanding of the carotenoid biosynthesis pathway. Furthermore, the sequence data will provide novel insight into UV radiation resistance in extremely cold environments.

Generation of structurally novel short carotenoids and study of their biological activity

Recent research interest in phytochemicals has consistently driven the efforts in the metabolic engineering field toward microbial production of various carotenoids. In spite of systematic studies, the possibility of using C30 carotenoids as biologically functional compounds has not been explored thus far. Here, we generated 13 novel structures of C30 carotenoids and one C35 carotenoid, including acyclic, monocyclic, and bicyclic structures, through directed evolution and combinatorial biosynthesis, in Escherichia coli. Measurement of radical scavenging activity of various C30 carotenoid structures revealed that acyclic C30 carotenoids showed higher radical scavenging activity than did DL-α-tocopherol. We could assume high potential biological activity of the novel structures of C30 carotenoids as well, based on the neuronal differentiation activity observed for the monocyclic C30 carotenoid 4,4′-diapotorulene on rat bone marrow mesenchymal stem cells. Our results demonstrate that a series of structurally novel carotenoids possessing biologically beneficial properties can be synthesized in E. coli.

In vitro study of the carotenoid-cleavage enzyme from Staphylococcus pasteuri TS-82 revealed substrate specificities and generation of norisoprenoid flavors.

Reactions of a crude enzyme extracted from S. pasteuri TS-82 to cleave carbon-carbon bonds in bicyclic and monocyclic carotenoid substrates were investigated. Dependencies of enzyme activities on processing temperature and pH were investigated and non-volatile and volatile breakdown products were characterized. The crude enzyme showed a maximum activity with zeaxanthin, followed in decreasing order by β-carotene, canthaxanthin, astaxanthin, and β-apo-8'-carotenal. The optimum pH value of the enzyme was 3.0 for both bicyclic and monocyclic substrates, whereas the optimum temperature of the enzyme was substrate specific at 60°C for C40 carotenoids and 50°C for β-apo-8'-carotenal. Liquid Chromatography-Mass Spectra (LC-MS) and Gas Chromatography- Mass Spectra (GC-MS) indicated that the crude enzyme was able to catalyze substrates with cleavage at 9-10 and 9'-10' double bonds with C13 norisoprenoids being the main volatile reaction products in each case. Astaxanthin is a major source for α,β-dihydro-β-ionone.

Characterisation of the C50 carotenoids produced by strains of the cheese-ripening bacterium Arthrobacter arilaitensis

Pigments were extracted from cheese-ripening bacteria belonging to the species Arthrobacter arilaitensis, which plays a role in the colour of cheese. A chromatographic method was applied to characterise the pigment profile. Based on the UV–visible (UV–vis) spectra, the mass spectra, and the elution order, eight different carotenoids were identified from the strains. Four of the pigments, namely, all-E-decaprenoxanthin, all-E-sarcinaxanthin, 9-Z-decaprenoxanthin and 15-Z-decaprenoxanthin, which were present as free-form main carotenoids (i.e., non-glycosylated), have been characterised by their UV–vis and mass spectra, both in atmospheric-pressure chemical ionisation negative [APCI (−)] and positive modes. The other minor carotenoids, namely, sarcinaxanthin monoglucoside pentaacetate, decaprenoxanthin monoglucoside, decaprenoxanthin diglucoside, decaprenoxanthin-C16:0 (decaprenoxanthin-palmitate), present in very low amounts, were only detected in the APCI (−) ionisation mode. Our results provide the first chemical characterisation of the pigments produced by cheese-ripening A. arilaitensis strains, which has not previously been reported.