Natural β-carotene and whole body irradiation in rats

β-Carotene and other carotenoids are reported to be potent free radical quenchers, singlet oxygen scavengers, and lipid antioxidants. Whole-body irradiation is known to cause an immunosuppression effect in mammals through the possible initiation and production of reactive oxygen species. We decided to test the possible antioxidative effect against whole-body irradiation of a natural β-carotene, composed of equal amounts of the all-trans and 9-cis isomers, obtained from the unicellular alga Dunaliella bardawil. Rats were fed on ground commercial food enriched with natural β-carotene (50 mg/kg diet). On completion of 1 week with β-carotene, the rats were exposed to a single dose of 4 Gy whole-body irradiation, after which their livers and blood were removed for β-carotene and retinol analysis in comparison with control livers of animals irradiated or not, or supplemented with β-carotene after irradiation. A normal increase in body weight with no ill effects was noted in the groups of rats whose diet was supplemented by β-carotene before and after irradiation, compared with the reduction in the specific growth rate in the group of rats irradiated without β-carotene. Liver β-carotene and retinol decreased significantly after irradiation compared with the rats which were not irradiated. This decrease was not shown in rats fed β-carotene prior to irradiation, and the effect of irradiation was partially cured by supplementation with β-carotene after irradiation. High-pressure liquid chromatography (HPLC) analysis of the irradiated animals showed a selective decline in 9-cis β-carotene and in retinol over all-trans β-carotene and retinyl esters. These results suggest that 9-cis β-carotene and retinol protect in vivo against the cellular damage by free radicals induced after whole-body irradiation. Received: 14 May 1996 / Accepted in revised form: 24 June 1996     

Tissue-specific accumulation of carotenoids in carrot roots

Raman spectroscopy can be used for sensitive detection of carotenoids in living tissue and Raman mapping provides further information about their spatial distribution in the measured plant sample. In this work, the relative content and distribution of the main carrot (Daucus carota L.) root carotenoids, α-, β-carotene, lutein and lycopene were assessed using near-infrared Fourier transform Raman spectroscopy. The pigments were measured simultaneously in situ in root sections without any preliminary sample preparation. The Raman spectra obtained from carrots of different origin and root colour had intensive bands of carotenoids that could be assigned to β-carotene (1,520 cm⁻¹), lycopene (1,510 cm⁻¹) and α-carotene/lutein (1,527 cm⁻¹). The Raman mapping technique revealed detailed information regarding the relative content and distribution of these carotenoids. The level of β-carotene was heterogeneous across root sections of orange, yellow, red and purple roots, and in the secondary phloem increased gradually from periderm towards the core, but declined fast in cells close to the vascular cambium. α-carotene/lutein were deposited in younger cells with a higher rate than β-carotene while lycopene in red carrots accumulated throughout the whole secondary phloem at the same level. The results indicate developmental regulation of carotenoid genes in carrot root and that Raman spectroscopy can supply essential information on carotenogenesis useful for molecular investigations on gene expression and regulation.     

Enhancement of antioxidant production in <Emphasis Type="Italic">Spirulina platensis</Emphasis> under oxidative stress

The present study examined the possibility of increasing the contents of some bioactive compounds of Spirulina platensis cultivated in medium containing various hydrogen peroxide concentrations (2, 4, 6 and 8 mM) as a model for environmental stress. A positive correlation was observed between the increase of H₂O₂ and increasing amounts of cellular lipophilic antioxidants (total carotenoids and α-tocopherol) and hydrophilic antioxidants [glutathione (GSH) and ascorbic acid (AsA)]. HPLC profile of carotenoids revealed that algae responded to the change of H₂O₂ exposure by the accumulation of higher amounts of β-carotene, astaxanthine, luteine, zeaxanthin and cryptoxanthin. S. platensis showed significant linear increase in activities of antioxidant enzymes, i.e., catalase (CAT), peroxidase (PX), ascorbate peroxidase (APX) and superoxide dismutase (SOD), with increasing H₂O₂ concentrations. A pronounced increase of oxidative lesions’ indexes [thiobarbituric acid reactive substances (TBARS) and paramagnetic radical-EPR signal] was found in algal grown at 8 mM H₂O₂. These data revealed that S. platensis behaved with different strategies against H₂O₂ exposure which is dose dependent and their response strongly correlated with the scavenging enzymes (SOD, CAT, PX and APX) and antioxidant compounds (GSH, AsA, β-carotene, astaxanthine and α-tocopherol) in the antioxidant defense systems. Therefore, S. platensis could be considered as good candidates for successful cultivation in artificial open ponds under different environmental conditions, as high value health foods, functional foods and as source of wide spectrum of nutrients.     

Biochemical characterization of carotenoids in two species of <Emphasis Type="Italic">Trentepohlia</Emphasis> (Trentepohliales,Chlorophyta)

Two species of Trentepohlia, i.e., Trentepohlia aurea and Trentepohlia cucullata were collected from walls and tree bark, respectively, at two different seasons in a year. The total carotenoid content in both the species is very high during winter but decreases significantly during summer. By spectroscopic analysis, it was found that. T. aurea and T. cucullata growing in natural habitats are rich sources of carotenoids. The individual carotenoids were separated, identified, and estimated by HPLC, and identified as β-carotene along with some other carotenoids, i.e., neoxanthin, lutein, β-cryptoxanthin, β,γ-carotene, β,ε-carotene (absent during summer).     

Lycopene cyclization inBlakeslea trispora

Fungi produce and accumulate various carotenoids. Mycelia of the ZygomyceteBlakeslea trispora contained β-carotene and its precursors γ-carotene and lycopene. When strains of opposite sex grew together, the β-carotene concentration increased fourfold, that of γ-carotene remained unchanged, and other intermediates practically disappeared. The inhibitors nicotine, 2-(4-chlorophenylthio)-triethylamine, α-picoline, and imidazole increased the concentrations of lycopene and γ-carotene and decreased those of β-carotene. From our quantitative results, we conclude thatBlakeslea has two pathways for lycopene metabolism, of which other fungi have only one or the other. The main one, two cyclizations from lycopene to β-carotene, is carried out by an enzyme dimer, is stimulated by sexual interaction, and is sensitive to the inhibitors. The other pathway, a cyclization to γ-carotene is not affected by mating or the inhibitors.     

Effects of vitamins on hepatic nuclear binding of L-tryptophan

Summary. This study investigated the in vitro effects of selected vitamins on nuclear L-tryptophan receptor binding of rat liver. Our results revealed that some fat-soluble vitamins, β-carotene, retinyl acetate, calciferol, α-tocopherol, and Trolox, as well as some water-soluble vitamins, thiamine and riboflavin, acted to inhibit in vitro ³H-tryptophan binding to hepatic nuclei. On the other hand, pyridoxine had little or no effect. The addition of dithiothreitol, a protective agent for sulfhydryl groups, along with each vitamin decreased the vitamin's inhibitory effect on in vitro ³H-tryptophan binding to nuclei, with the exception of riboflavin and calciferol. The addition of L-leucine, which alone had no inhibitory effect on in vitro ³H-tryptophan binding to hepatic nuclei but when added with unlabeled L-tryptophan negated the effect of unlabeled L-tryptophan, caused a markedly diminished inhibitory binding effect due to each of the following vitamins, thiamine, β-carotene, retinyl acetate, and α-tocopherol and Trolox, but no effect on riboflavin and calciferol. Received December 29, 1999 Accepted March 8, 2000     

The Roles of Melatonin and Vitamin E Plus Selenium in Prevention of Oxidative Stress Induced by Naloxone-Precipitated Withdrawal in Heroin-Addicted Rats

The therapeutic effects of melatonin or vitamin E plus Se (vE + Se) on the restrain of the heroin withdrawal-induced oxidative stress were studied. For this, rats were divided into ten groups. The rats were injected by fixed or variable doses of heroin for 16 consecutive days, and naloxone was given 1 h after the last heroin injection. One hour after naloxone administration, some groups were treated with melatonin or vE + Se. After 1 h this, blood samples were taken, and the levels of malondialdehyde (MDA) and reduced glutathione (GSH) in whole blood, ascorbic acid, α-tocopherol, retinol, β-carotene, nitrite, nitrate, and ceruloplasmin levels in the serum were measured. Our findings showed that, naloxone administration precipitated the heroin withdrawal. This also increased the level of MDA and decreased the levels of GSH in blood. Melatonin or vE + Se administration prevented the rise in MDA levels and increased the GSH levels. On the other hand, there were some significant differences between α-tocopherol, retinol, β-carotene, nitrite, nitrate, and ceruloplasmin levels of experimental groups. Results of present study showed that heroin withdrawal increased the lipid peroxidation and depressed endogenous antioxidative systems. Additionally, melatonin or vE + Se administrations prevented lipid peroxidation and augmented endogenous antioxidant defense systems.     

Substrate contribution on free radical scavenging capacity of carotenoid extracts produced from <Emphasis Type="Italic">Blakeslea trispora</Emphasis> cultures

Blakeslea trispora produces carotenoids mixtures consisting mainly of lycopene, γ-carotene and β-carotene, together with trace amounts of other carotenoid precursors. The yield of these carotenoids and their composition are greatly affected by culture substrate. The scavenging capacity of carotenoids extract from cultures of B. trispora growing in various substrates was estimated using the 2,2-diphenyl-1-picrylhydrazyl method. Fractions enriched in β-carotene, γ-carotene and lycopene, obtained after column chromatography in alumina basic II, were also examined. Substrates containing starch and oils mixture, Ni²⁺, and that with pantothenic acid presented higher antioxidant activity. An increase in the antioxidant activity of the crude carotenoid extract compared to that of the isolated fractions enriched in β-carotene, γ-carotene and lycopene respectively, observed in most samples, indicated a possible synergistic effect. The results are of interest and by expanding this study to more substrates and other microorganisms- producing antioxidants, a formulation of extract with high free radical scavenging potential could be produced.     

Dynamic Changes of Photosynthetic Pigments in Soybean Callus under High Irradiance

Dynamic changes of neoxanthin (NEO), violaxanthin (VIO), anteraxanthin (ANT), zeaxanthin (ZEA), chlorophyll (Chl) a, Chl b, α-carotene, β-carotene, and their behaviour under increasing duration of high irradiance (HI) were investigated in the soybean hypocotyl callus culture. The calli were induced on solid (1.1 % agar) MS medium (pH 5.8) supplemented with 4.52 μM 2,4-D, 2.32 μM kinetin, and 3 % sucrose. After 30 d of culture, the green calli were irradiated with “white light” (133W m⁻²) for 0, 3.5, and 24 h. HPLC profiles were separated on a C₁₈ column. With increasing duration of HI, the content of total carotenoids (Cars) increased, but the ratio of Chl a+b/Cars decreased. With lengthening the duration of HI, there was induction of ZEA. Contents of ANT, α-carotene, and β-carotene remained nearly constant, but ratio of ZEA/Chl a+b increased with lengthening the HI duration. This revised version was published online in August 2006 with corrections to the Cover Date.     

Using molecular markers to identify two major loci controlling carotenoid contents in maize grain

Maize is an important source of pro-vitamin A; β-carotene, α-carotene and β-cryptoxanthin, and the non-pro-vitamin A carotenoids including lutein and zeaxanthin. In the present study, a recombinant inbred (RI) population with 233 RI lines derived from a cross between By804 and B73 was employed to detect QTL for these nutritionally important components in maize grain. High Performance Liquid Chromatography was used to measure amounts of individual carotenoids over 2 years. A genetic linkage map was constructed with 201 molecular markers. In all, 31 putative QTL including 23 for individual and 8 for total carotenoids were detected on chromosome(s) 1, 3, 5, 6, 7, 8 and 10. The notable aspect of this study was that much of the phenotypic variation in contents of carotenoids could be explained by two loci (y1 and y9), and the QTL for carotenoids elucidated the interrelationships among these compounds at the molecular level. A gene targeted marker (Y1ssr) in the candidate gene phytoene synthase 1 (psy1) tightly linked to a major QTL explaining 6.6–27.2% phenotypic variation for levels of carotenoids was identified, which may prove useful to expedite breeding for higher level of carotenoids in maize grain. This functionally characterized gene (psy1) could also be exploited for further development of functional marker for carotenoids in maize. The QTL cluster located at y9 locus may also be used for pyramiding favorable alleles controlling contents of carotenoids from diverse maize germplasm.     

Isocratic rapid liquid chromatographic method for simultaneous determination of carotenoids,retinol, and tocopherols in human serum

An improved isocratic and rapid HPLC method was developed for the measurement of carotenoids, retinol and tocopherols in human serum. Vitamins were extracted with hexane. Mobile phase consisted of a mixture acetonitrile:methylene chloride:methanol with 20 mM ammonium acetate. This method used a small bead size (3 μm) Spherisorb ODS2 column with titane frits. Diode array and fluorescence detectors were used respectively for the detection of carotenoids and retinol/tocopherols. Chromatographic separation was complete in 13 min for β-cryptoxanthin, cis–trans-lycopene, α-carotene, β-carotene, cis-β-carotene, retinol, δ-tocopherol, γ-tocopherol and α-tocopherol. Echinenone and tocol were employed as internal standards for diode array and fluorescence detection. Accuracy was validated using standard reference material (SRM) 968C. Intra-assay and inter-assay precision were respectively 0.2–7.3% and 3.6–12.6%. Sensitivity was verified using the ICH recommendations and the limit of detection (LOD) obtained was sufficient for routine clinical application.     

Unique Carotenoids in the Terrestrial Cyanobacterium <Emphasis Type="Italic">Nostoc commune</Emphasis> NIES-24: 2-Hydroxymyxol 2′-Fucoside,Nostoxanthin and Canthaxanthin

Cyanobacteria produce some carotenoids. We identified the molecular structures, including the stereochemistry, of all the carotenoids in the terrestrial cyanobacterium, Nostoc commune NIES-24 (IAM M-13). The major carotenoid was β-carotene. Its hydroxyl derivatives were (3R)-β-cryptoxanthin, (3R,3′R)-zeaxanthin, (2R,3R,3′R)-caloxanthin, and (2R,3R,2′R,3′R)-nostoxanthin, and its keto derivatives were echinenone and canthaxanthin. The unique myxol glycosides were (3R,2′S)-myxol 2′-fucoside and (2R,3R,2′S)-2-hydroxymyxol 2′-fucoside. This is only the second species found to contain 2-hydroxymyxol. We propose possible carotenogenesis pathways based on our identification of the carotenoids: the hydroxyl pathway produced nostoxanthin via zeaxanthin from β-carotene, the keto pathway produced canthaxanthin from β-carotene, and the myxol pathway produced 2-hydroxymyxol 2′-fucoside via myxol 2′-fucoside. This cyanobacterium was found to contain many kinds of carotenoids and also displayed many carotenogenesis pathways, while other cyanobacteria lack some carotenoids and a part of carotenogenesis pathways compared with this cyanobacterium.     

Production and optimization of carotenoid-enriched dried distiller’s grains with solubles by <Emphasis Type="Italic">Phaffia rhodozyma</Emphasis> and <Emphasis Type="Italic">Sporobolomyces roseus</Emphasis> fermentation of whole stillage

Whole stillage—a co-product of grain-based ethanol—is used as an animal feed in the form of dried distiller’s grain with solubles (DDGS). Since animals cannot synthesize carotenoids and animal feed is generally poor in carotenoids, about 30–120 ppm of total carotenoids are added to animal feed to improve animal health, enhance meat color and quality, and increase vitamin A levels in milk and meat. The main objective of this study was to produce carotenoid (astaxanthin and β-carotene)-enriched DDGS by submerged fermentation of whole stillage. Mono- and mixed cultures of red yeasts, Phaffia rhodozyma (ATCC 24202) and Sporobolomyces roseus (ATCC 28988), were used to produce astaxanthin and β-carotene. Media optimization was carried out in shake flasks using response surface methodology (RSM). Macro ingredients, namely whole stillage, corn steep liquor and glycerol, were fitted to a second-degree polynomial in RSM. Under optimized conditions, astaxanthin and β-carotene yields in mixed culture and P. rhodozyma monoculture were 5 and 278, 97, and 275 μg/g, respectively, while S. roseus produced 278 μg/g of β-carotene. Since the carotenoid yields are almost twice the quantity used in animal feed, the carotenoid-enriched DDGS has potential application as “value-added animal feed or feed blends.”     

Changes in carotenoids, tocopherols and diterpenes during drought and recovery, and the biological significance of chlorophyll loss in Rosmarinus officinalis plants

Two-year-old rosemary (Rosmarinus officinalis L.) plants were subjected to severe stress by exposure to prolonged drought during a Mediterranean summer. Severely stressed plants recovered completely after the autumn rainfalls although the relative water content remained below 35% for 3 months and the chlorophyll content of leaves was reduced by up to 85% during the drought. In severe stress: (i) α-tocopherol increased 9-fold per g dry weight and 20-fold per unit of chlorophyll; (ii) lutein and β-carotene contents decreased on a dry-weight basis, but an 80% increase in lutein and constant levels of β-carotene were observed on a chlorophyll basis; (iii) there were transient and sustained increases in the de-epoxidation state of the xanthophyll cycle; and (iv) the highly oxidised abietane diterpene isorosmanol increased 8-fold as a result of the oxidation of carnosic acid. With the autumn rainfalls, water status, α-tocopherol and violaxanthin recovered first and the levels of photosynthetic pigments and abietane diterpenes increased later. The photoprotection conferred by the xanthophyll cycle and the antioxidant function of tocopherols, lutein and diterpenes may help to avoid irreversible damage in severe drought, making possible the recovery of functional membranes after the autumn rainfalls. Besides, chlorophyll loss reduces the amount of photons absorbed by leaves, which enhances the photoprotective and antioxidant capacity of leaves per amount of photons absorbed, since the ratios of xanthophylls, α-tocopherol and abietane diterpenes to chlorophyll increase. Received: 12 July 1999 / Accepted: 25 November 1999     

Plasma micronutrient concentrations in infants undergoing therapy for phenylketonuria

Twenty-seven infants with classical phenylketonuria were evaluated longitudinally for 6 mo while ingesting Phenex^TM -1 Amino Acid Modified Medical Food With Iron as their primary protein source. Intake of selected nutrients and biochemical indices of trace and ultratrace mineral status and plasma retinol and α-tocopherol concentrations were evaluated. The means of iron status indices (complete blood count, plasma ferritin, iron, transferrin saturation, total iron binding capacity) and the plasma concentrations of trace and ultratrace minerals (copper, manganese, molybdenum, selenium, zinc) and plasma retinol and α-tocopherol were in the reference ranges. Vitamin A intakes (r = 0.49,p < 0.05) and plasma retinol-binding protein concentrations (r = 0.42,p < 0.05) were positively correlated with plasma retinol concentrations at 3 mo of study. At 6 mo, concentrations of plasma transthyretin (r = 0.72,p < 0.01) and retinolbinding protein (r = 0.48,p < 0.05) were positively correlated with plasma retinol concentrations. At 6 mo, concentrations of plasma transthyretin (r = 0.52,p < 0.05) were positively correlated with retinol-binding protein concentrations. Phenex-1 supports normal mean iron status indices and mean concentrations of trace and ultratrace minerals, retinol, and α-tocopherol when fed in adequate amounts.     

Serum α- and γ-tocopherols, retinol, retinyl palmitate, and carotenoid concentrations in captive and free-ranging bottlenose dolphins (Tursiops truncatus)

Concentrations of retinol, retinyl palmitate, β-carotene, α-carotene, cryptoxanthin, lutein, lycopene, α-tocopherol, and γ-tocopherol were measured in blood samples collected from 15 captive and 55 free-ranging bottlenose dolphins (Tursiops truncatus). From June 1991 to June 1994, blood samples were collected from captive animals residing at two locations; at Seven Seas (Brookfield Zoo, Brookfield, IL) and Hawk’s Cay (Marathon Key, FL). Blood samples were collected from free-ranging animals from June 1991 to June 1996. Retinol levels were not significantly different between captive dolphin groups. However, Seven Seas animals had higher (P<0.01) serum retinol concentrations compared to free-ranging animals (0.061 vs 0.041 μg/ml). Retinyl palmitate was not detected in the serum of captive or free-ranging dolphins. Alpha-tocopherol levels were significantly (P<0.05) higher for Seven Seas dolphins (16.4 μg/ml) than for Hawk’s Cay (13.0 μg/ml) and free-ranging dolphins (12.5 μg/ml). Gamma-tocopherol concentrations were similar among captive and free-ranging dolphins. Free-ranging dolphins showed levels of circulating carotenoids (lutein and β-carotene) while the captive animals did not. Additional carotenoids (lycopene, α-carotene and cryptoxanthin) were analyzed but not detected in any samples. Serum vitamin differences between captive and free-ranging dolphins may reflect the natural diet or indicate some potential biological or nutritional status significance.     

Construction of new <Emphasis Type="Italic">Pichia pastoris</Emphasis> X-33 strains for production of lycopene and β-carotene

In this study, we used the non-carotenogenic yeast Pichia pastoris X33 as a receptor for β-carotene-encoding genes, in order to obtain new recombinant strains capable of producing different carotenoidic compounds. We designed and constructed two plasmids, pGAPZA-EBI* and pGAPZA-EBI*L*, containing the genes encoding lycopene and β-carotene, respectively. Plasmid pGAPZA-EBI*, expresses three genes, crtE, crtB, and crtI*, that encode three carotenogenic enzymes, geranylgeranyl diphosphate synthase, phytoene synthase, and phytoene desaturase, respectively. The other plasmid, pGAPZA-EBI*L*, carried not only the three genes above mentioned, but also the crtL* gene, that encodes lycopene β-cyclase. The genes crtE, crtB, and crtI were obtained from Erwinia uredovora, whereas crtL* was cloned from Ficus carica (JF279547). The plasmids were integrated into P. pastoris genomic DNA, and the resulting clones Pp-EBI and Pp-EBIL were selected for either lycopene or β-carotene production and purification, respectively. Cells of these strains were investigated for their carotenoid contents in YPD media. These carotenoids produced by the recombinant P. pastoris clones were qualitatively and quantitatively analyzed by high-resolution liquid chromatography, coupled to photodiode array detector. These analyses confirmed that the recombinant P. pastoris clones indeed produced either lycopene or β-carotene, according to the integrated vector, and productions of 1.141 μg of lycopene and 339 μg of β-carotene per gram of cells (dry weight) were achieved. To the best of our knowledge, this is the first time that P. pastoris has been genetically manipulated to produce β-carotene, thus providing an alternative source for large-scale biosynthesis of carotenoids.     

Increased lipid peroxidation in pregnant women after iron and vitamin C supplementation

Iron overload could promote the generation of free radicals and result in deleterious cellular damages. A physiological increase of oxidative stress has been observed in pregnancy. A routine iron supplement, especially a combined iron and vitamin C supplementation, without biological justifications (low hemoglobin [Hb] and iron stores) could therefore aggravate this oxidative risk. We investigated the effect of a daily combined iron supplementation (100 mg/d as fumarate) and vitamin C (500 mg/d as ascorbate) for the third trimester of pregnancy on lipid peroxidation (plasma TBARS), antioxidant micronutriments (Zn, Se, retinol, vitaminE, (β-carotene) and antioxidant metalloenzymes (RBC Cu-Zn SOD and Se-GPX). The iron-supplemented group (n=27) was compared to a control group (n=27), age and number of pregnancies matched. At delivery, all the women exhibited normal Hb and ferritin values. In the supplemented group, plasma iron level was higher than in the control group (26.90±5.52 mmol/L) and TBARs plasma levels were significantly enhanced (p<0.05) (3.62±0.36 vs 3.01±0.37 mmol/L). No significant changes were observed in plasma trace elements and red blood cell antioxidant metalloenzymes. Furthermore, the α-tocopherol plasma level was lowered in the iron-supplemented groups, suggesting an increased utilization of vitamin E. These data show that pharmalogical doses of iron, associated with high vitamin C intakes, can result in uncontrolled lipid peroxidation. This is predictive of adverse effects for the mother and the fetus. This study illustrates the potential harmful effects of iron supplementation when prescribed only on the assumption of anemia and not on the bases of biological criteria.     

Torularhodin and torulene are the major contributors to the carotenoid pool of marine Rhodosporidium babjevae (Golubev)

A carotenoid-producing yeast strain, isolated from the sub-arctic, marine copepod Calanus finmarchicus, was identified as Rhodosporidium babjevae (Golubev) according to morphological and biochemical characteristics and phylogenetic inference from the small-subunit ribosomal RNA gene sequence. The total carotenoids content varied with cultivation conditions in the range 66–117 μg per g dry weight. The carotenoid pool, here determined for the first time, was dominated by torularhodin and torulene, which collectively constituted 75–91% of total carotenoids under various regimes of growth. β-Carotene varied in the range 5–23%. A high-peptone/low-yeast extract (weight ratio 38:1) marine growth medium favoured the production of torularhodin, the carotenoid at highest oxidation level, with an average of 63% of total carotenoids. In standard yeast medium (YM; ratio 1.7:1), torularhodin averaged 44%, with increased proportions of the carotenes, torulene and β-carotene. The anticipated metabolic precursor γ-carotene (β,ψ-carotene) constituted a minor fraction (≤8%) under all conditions of growth.