Mineral elements and their relation to anthocyanin content in pigmented rice plants using definitive screening design and self-organizing maps.

BACKGROUND: Mineral elements are nutrients required by an organism to perform functions necessary for survival. Stress-induced metabolism following nutritional stress has been reported to increase levels of anthocyanin. However, the role of mineral elements commonly found in soil and their contribution to the accumulation of anthocyanin content in rice plants is uncertain. RESULT: Amongst the ten mineral elements investigated, the cultivation of rice plants in clean sand showed that the Mg-, Se-, and Cu-treated plants had the highest accumulated anthocyanin content in the leaves, whereas B, Cr, and Se had the greatest effect on grains. Yield component data showed major positive effects from Mg, Cr, and B. The interaction of Zn*Se and Mg*Cu positively affected the anthocyanin content in grains. The self-organizing map indicated that the total anthocyanin content was relatively proportional to the concentration of Mn, B, and Cr, but disproportional to that of Se. However, rice plants with added Fe produced the smallest amount of total anthocyanin content, less than the control, in the four stages of rice growth. CONCLUSION: The appropriate concentrations of mineral elements in soil could promote the proliferation of anthocyanin content in rice plants and grains. © 2023 Society of Chemical Industry.

Influence of black rice (Oryza sativa L.) bran derived anthocyanin-extract on growth rate, immunological response, and immune-antioxidant gene expression in Nile tilapia (Oreochromis niloticus) cultivated in a biofloc system.

This study investigated the effects of dietary supplementation with anthocyanin extracted from black rice bran (AR) on the growth rate, immunological response, and expression of immune and antioxidant genes in Nile tilapia raised in an indoor biofloc system. A total of 300 Nile tilapia fingerlings (15.14 ± 0.032 g) were maintained in 150 L tanks and acclimatized for two weeks. Five experimental AR diets (0, 1, 2, 4, and 8 g kg-1) with various anthocyanin doses were used to feed the fish. We observed that the growth and feed utilization of fish fed with different dietary AR levels increased significantly after eight weeks (p < 0.05). In addition, the serum immunity of fish fed AR diets was much greater than that of those fed non-AR diets (p < 0.05). However, there were little or no difference in between fish fed AR enriched diets and the control AR-free diet (p > 0.05). After eight weeks, fish fed AR-supplemented diets had significantly higher mRNA transcript levels in immune (interleukin [IL]-1, IL-8, and liposaccharide-binding protein [LBP]) and antioxidant (glutathione transferase-alpha [GST-α] and glutathione reductase [GSR]) genes compared to control fish fed the AR-free diet, with the greatest enhancement of mRNA transcript levels (in the case of IL-8 by up to about 5.8-fold) in the 4 g kg-1 AR diet. These findings suggest that dietary inclusion of AR extract from black rice bran at 4-8 g kg-1 could function as a herbal immunostimulant to enhance growth performance, feed consumption, and immunity in Nile tilapia.

Bioactive Constituents in Cold-Pressed Plant Oils: Their Structure, Bioactivity and Chromatographic Analysis.

Cold-pressed oils are oils prepared from pressing plant materials with a screw or hydraulic press, yielding oils with little contamination of harmful chemicals and high content of nutrients and functional constituents. Cold-pressed oils have gained increasing recognition as food supplements for preventing and ameliorating body deterioration due to ageing and the progression of lifestyle diseases or non-communicable diseases. This article aimed to review their structure, bioactivity, and chromatographic analysis of the mostly found functional compounds in cold-pressed oils, including phytosterols, carotenoids, tocols (tocopherols and tocotrienols), phenolic compounds (flavonoids, phenolic acids, tannins, stilbenes, and lignans), and squalene.

Development of a Simultaneous Normal-Phase HPLC Analysis of Lignans, Tocopherols, Phytosterols, and Squalene in Sesame Oil Samples.

The objective of this study was to develop a simultaneous analytical method for the determination of lignans, tocols, phytosterols, and squalene using high-performance liquid chromatography coupled with a diode array and fluorescence detector (HPLC-DAD-FLD). The method employed a VertisepTM UPS silica HPLC column (4.6 × 250 mm, 5 µm) with a mobile phase mixture of n-hexane/tetrahydrofuran/2-propanol. This approach enabled the simultaneous analysis of ten compounds within 22 min. The linear correlation (R2) exceeded 0.9901. The limit of detection (LOD) and limit of quantitation (LOQ) were up to 0.43 µg mL-1 for lignans and tocopherols and up to 326.23 µg mL-1 for phytosterol and squalene. The precision and accuracy of the intra-day and inter-day variation were less than 1.09 and 3.32% relative standard deviations (RSDs). Furthermore, the developed method was applied for the analysis of targeted compounds in twenty-eight sesame oil samples (1775-8965 µg g-1 total lignans, 29.7-687.9 µg g-1 total tocopherols, 2640-9500 µg g-1 phytosterol, and 245-4030 µg g-1 squalene). The HPLC method that has been developed was proven to be a reliable and effective tool for the determination of those functional compounds among sesame oil samples.

Enhancing Functional Compounds in Sesame Oil through Acid-Soaking and Microwave-Heating of Sesame Seeds.

This study investigated whether pre-treating sesame (Sesamum indicum L.) seeds with a combination of acid-soaking and microwave-heating could significantly enhance the quality of the resulting sesame oil, particularly by increasing its content of functional compounds such as lignans, tocopherol, phytosterol, and squalene. The study revealed that soaking the sesame seeds in a solution of HCl and citric acid, along with microwave-heating, significantly increased the content of these compounds. The detected ranges were sesamin (1365-6927 µg g-1), sesamolin (605-3493 µg g-1), tocopherol (69.31-282.76 µg g-1), asarinin (ND-383.52 µg g-1), sesamol (ND-49.59 µg g-1), phytosterol (3690-6201 µg g-1), and squalene (532-1628 µg g-1). Additionally, the study found that the pre-treatment of sesame seeds had a minimal effect on the fatty acid composition, antioxidant activity (92.94-95.08% DPPH scavenging activity), and oxidative stability (2.13-2.90 mg MDA kg-1 oil). This is the first study to demonstrate that using acid-soaking and microwave-heating to prepare sesame seeds can produce sesame oil enriched with functional compounds, potentially benefiting cosmetic, pharmaceutical, and health applications.

Phosphatidylcholine hydroperoxide promotes VEGF-induced angiogenesis in endothelial cells and rat aorta ring cultures.

BACKGROUND: Phosphatidylcholine hydroperoxide (PCOOH) is a primary oxidation product of PC, and is markedly accumulated in blood plasma and arterial walls in atherosclerotic animals and humans. The role of PCOOH in the induction of angiogenesis is unknown. METHODS: In this study, we investigated whether PCOOH stimulated angiogenic responses (e.g., vascular endothelial growth factor (VEGF)-induced cell proliferation, migration, and tube formation, and angiogenesis-related gene/protein expression) in human umbilical vein endothelial cells (HUVEC) and in an ex vivo rat aorta model. RESULTS: VEGF induced proliferation, migration, and tube formation of HUVEC, and these angiogenic responses were all enhanced by PCOOH but not by native (nonoxidized) PC. The angiogenic effects of PCOOH are considered to be mediated via generation of reactive oxygen species and activation of both PI3K/AKT and MAPK pathways. The angiogenic activities of PCOOH were also confirmed by the rat aortic ring assay. CONCLUSIONS: These results indicate that PCOOH can elicit several angiogenic responses. GENERAL SIGNIFICANCE: The present study implies an important role of PCOOH in atherosclerosis progression and plaque instability.

Microwave treatment of rice bran and its effect on phytochemical content and antioxidant activity.

An alternative approach for rice bran stabilization is microwave treatment. However, the effects of the microwave treatment on the contents of bioactive compounds and antioxidant activities of the rice bran have rarely been reported in detail. In this study, microwave pretreatment (130-880 W for 0.5-5.0 min) of rice bran was proposed where the antioxidant activity, total flavonoids, and total phenolic contents were determined using UV-Vis spectrometry. Tocols, γ-oryzanols, squalene, phytosterols and phenolic compounds were quantified using high-performance liquid chromatography. The results showed an increase in the antioxidant activity (0.5 folds), total phenolic contents (1.3 folds), total flavonoid contents (0.9 folds), total tocols (2.6 folds), total γ-oryzanols (1.6 folds), and total phytosterols (1.4 folds). Phytochemicals were enhanced, especially trans-p-coumaric acid (10.3 folds) and kaempferol (8.6 folds). The microwave treatment at 440 W for 2.5 min provided the best contents of the bioactive compounds and antioxidant activity. This work revealed the microwave treatment as a potential tool for stabilizing rice bran and increasing the usability of its phytochemicals, which applies to several industries concerning the use of rice bran as an ingredient.

Smartphone digital image colorimetric determination of the total monomeric anthocyanin content in black rice via the pH differential method.

In this research, the pH differential method was explored for the first time using a mobile phone as a detector, replacing UV-Vis spectrophotometry. A smartphone digital image colorimetric (SDIC) system was developed for the determination of the total monomeric anthocyanin content in black rice grains using colour values. The change in colour of anthocyanin cyanidin-3-glucoside (C3G) at pH 1.0 and pH 4.5 was investigated and described through colour model systems (RGB and CIELAB). Under optimum conditions, the quantitative method based on the chroma difference, was able to quantify the total amount of monomeric anthocyanins with a linear correlation (R2 = 0.989) to that of UV-Vis spectrophotometry and high performance liquid chromatography (HPLC) (reference method). The SDIC system displayed good precision (≤1.88% RSD) and a low detection limit (2.2 ± 0.1 µg g-1). The validated results demonstrated that the developed method was a cost-effective alternative for the quantitation of anthocyanins. In addition, the effect of six mineral elements on black rice cultivation was investigated. The results showed that the addition of the Ca fertiliser resulted in black rice grains with an anthocyanin content 2-fold higher than that of the control with a significant difference (p < 0.05).

Comparative Studies on the Hepatoprotective Effect of White and Coloured Rice Bran Oil against Acetaminophen-Induced Oxidative Stress in Mice through Antioxidant- and Xenobiotic-Metabolizing Systems.

Rice bran oil (RBO) comprises various nutrients and phytochemicals which exhibit several health benefits. There are no studies regarding the functional effects of different colours of RBO. This study was aimed to compare the constituents and antioxidant activities of white rice bran oil (WRBO) and coloured rice bran oil (CRBO). Each RBO showed similar free fatty acid profiles. However, greater amounts of vitamin E, phytosterols, carotenoids, and chlorophylls were found in CRBO, which had lower γ-oryzanol content than WRBO. Oxidative stress was induced in male mice by an overdose of acetaminophen (APAP) at 300 mg/kg body weight. The mice were then fed with RBO at the equivalent dose to 100 mg/kg body weight of γ-oryzanol three hours later and sacrificed six hours after APAP treatment. The administration of 100 mg γ-oryzanol equivalent in CRBO ameliorated APAP-induced hepatotoxicity in mice more strongly than 100 mg γ-oryzanol equivalent in WRBO, as evidenced by the significant reduction of serum ALT, hepatocellular necrosis, and hepatic lipid peroxidation. CRBO could improve xenobiotic-metabolizing and antioxidant enzyme activities, including glutathione S-transferase, superoxide dismutase, glutathione peroxidase, and glutathione reductase, and also increase mRNA expression of various antioxidant-responsive genes. Vitamin E, phytosterols, carotenoids, and chlorophyll might be the protective compounds in CRBO that alleviate APAP-induced hepatotoxicity through the interruption of APAP metabolism and the activation of antioxidant systems at both transcriptional and enzymatic levels. These findings might provide a protective role of CRBO on oxidative stress associated with several degenerative diseases.

GC-MS and HPLC-DAD analysis of fatty acid profile and functional phytochemicals in fifty cold-pressed plant oils in Thailand.

Cold-pressed oil is one of the healthiest plant extracts, but its use is limited only in some kinds of plants. Therefore, we aimed to investigate some potential cold-pressed oils with attractive fatty acid profiles and high amounts of functional phytochemicals. Fifty cold-pressed plant oils were prepared from various plant materials in Thailand, in which some of them were from uncommon or unattended plant materials. The oils included were nut oils (n = 9), pseudo-cereal oils (n = 9), legume oils (n = 3), amaranth oils (n = 3), marrow seed oils (n = 8), cruciferous seed oils (n = 7), and leafy green seed oils (n = 11). Gas-chromatography mass-spectrometry (GC-MS) and high-performance liquid chromatography coupled with a diode array detector (HPLC-DAD) were employed to analyze fatty acid profile and five functional phytochemicals (e.g., phytosterols, cholecalciferol, and squalene). Saturated fatty acids were detected around 7.87-36.04%, monounsaturated fatty acids 10.17-80.25%, and polyunsaturated fatty acids nondetectable (ND)-78.25%, phytosterols 663-15123 µg g-1, squalene 265-5979 µg g-1, and cholecalciferol ND-1287.75 µg g-1. The study showed chemical characteristic of the analyzed oils: some contained good fatty acid composition and some were rich in functional phytochemical content. Among the obtained oils, marrow seed oils are a good source of phytosterol, cholecalciferol, and linoleic acid. Pseudo-cereal oils are rich in squalene and linolenic acid. Legume oils are rich in phytosterols and oleic acid. Besides, principal component analysis (PCA) was applied to identify the significance of oils that share compositional similarity (e.g., the samples from pseudo-cereal oil were found on the lower side of the PCA space, which separated them from marrow and leafy green seed oils distributed on the upper part of the plot). In summary, the qualitative and quantitative data would provide a good foundation for further application or selection of those plant oils for health purposes.

alpha-Tocopherol attenuates the cytotoxic effect of delta-tocotrienol in human colorectal adenocarcinoma cells.

Recent studies have demonstrated that tocotrienol (T3) is superior to tocopherol (Toc) for cancer chemoprevention. However, there is little information on whether Toc influences the anticancer property of T3. In this study, we investigated the influence of Toc on the cytotoxic effects of delta-T3 in DLD-1 human colorectal adenocarcinoma cells. Toc, especially alpha-Toc, attenuated delta-T3-induced cytotoxicity and apoptosis in DLD-1 cells, whereas Toc alone did not exhibit any cytotoxic effect. delta-T3-induced cell cycle arrest and proapoptotic gene/protein expression (e.g., p21, p27, and caspases) were abrogated by alpha-Toc. Furthermore, coadministration of alpha-Toc decreased delta-T3 uptake into DLD-1 cells in a dose-dependent manner. These results indicate that alpha-Toc is not only less cytotoxic to cancer cells, but it also reduces the cytotoxicity of delta-T3 by inhibiting its cellular uptake.

Antiangiogenic and anticancer potential of unsaturated vitamin E (tocotrienol).

Several lines of evidence support the beneficial effect of tocotrienol (T3; an unsaturated vitamin E) on inhibition of tumor development. Many factors, including decrease in oxidative stress and modulation of cell signaling pathways in tumor and endothelial cells, have been implicated in such anticancer action of T3, while the in vivo potency and exact intracellular mechanisms for the anticancer properties of T3 remain not fully understood. We have hypothesized that the inhibitory effect of T3 on cancer may be attributable to the antiangiogenic activity of T3, and we found that T3 acts as a potent regulator of growth-factor-dependent signaling in endothelial cells and as an antiangiogenic agent minimizing tumor growth. In this work, we review the history and biological action (i.e., anticancer) of vitamin E and describe current research on the antiangiogenic effects of T3 and its mechanisms.

Antioxidant effect of lutein towards phospholipid hydroperoxidation in human erythrocytes.

Peroxidised phospholipid-mediated cytotoxity is involved in the pathophysiology of many diseases; for example, phospholipid hydroperoxides (PLOOH) are abnormally increased in erythrocytes of dementia patients. Dietary carotenoids (especially xanthophylls, polar carotenoids such as lutein) have gained attention as potent inhibitors against erythrocyte phospholipid hydroperoxidation, thereby making them plausible candidates for preventing diseases (i.e. dementia). To evaluate these points, we investigated whether orally administered lutein is distributed to human erythrocytes, and inhibits erythrocyte PLOOH formation. Six healthy subjects took one capsule of food-grade lutein (9.67 mg lutein per capsule) once per d for 4 weeks. Before and during the supplementation period, carotenoids and PLOOH in erythrocytes and plasma were determined by our developed HPLC technique. The administered lutein was incorporated into human erythrocytes, and erythrocyte PLOOH level decreased after the ingestion for 2 and 4 weeks. The antioxidative effect of lutein was confirmed on erythrocyte membranes, but not in plasma. These results suggest that lutein has the potential to act as an important antioxidant molecule in erythrocytes, and it thereby may contribute to the prevention of dementia. Therefore future biological and clinical studies will be required to evaluate the efficacy as well as safety of lutein in models of dementia with a realistic prospect of its use in human therapy.

Optimization of 1-deoxynojirimycin extraction from mulberry leaves by using response surface methodology.

Mulberry 1-deoxynojirimycin (DNJ, a potent alpha-glycosidase inhibitor) has therapeutic potency against diabetes mellitus. However, the amount of DNJ in mulberry leaves is low (about 0.1%), and a more effective extraction method is needed. Ultrasound-assisted extraction (UAE) was applied in this study for mulberry DNJ extraction, and five factors, the percentage of ethanol in the extraction solvent (x(1)), ratio of the extraction solvent to mulberry sample (x(2)), ultrasonic power (x(3)), extraction temperature (x(4)) and extraction time (x(5)), were investigated by fractional factorial 2((5-1)) design (FFD) to obtain the optimum extraction efficiency (DNJ yield, Y(1)) and extraction productivity (total yield, Y(2)). The results showed that x(2), x(3) and x(5) had significant impact on Y(1) and Y(2), and were further optimized by response surface methodology (RSM). Under the optimized conditions (x(2), x(3) and x(5) of 7 ml/g, 180 W and 260 s, respectively), DNJ-enriched powder (0.8%) was produced with high extraction efficiency (98%) and productivity (20%), enabling this product to be used for nutraceutical purposes.

Effects of co-administration of tea epigallocatechin-3-gallate (EGCG) and caffeine on absorption and metabolism of EGCG in humans.

Based on the ratios of (-)-epigallocatechin-3-gallate (EGCG) and caffeine (CAF) levels found in commercial tea drinks, EGCG and CAF were co-administered to human volunteers at various EGCG/CAF ratios, and plasma EGCG was determined by high performance liquid chromatography with chemiluminescence detection. As for the results, in plasma taken after ingestion of a beverage containing 95 mg of EGCG alone, the area under the plasma EGCG concentration-time curve (AUC) was 857 ngxh/ml. A higher AUC (1,370 ngxh/ml) was observed when subjects ingested a beverage containing EGCG (95 mg) and a low amount of CAF (40 mg). In the case of ingestion of a beverage containing EGCG (95 mg) and a high amount of CAF (180 mg), the AUC tended to be somewhat higher (1,165 ngxh/ml), but not significantly so, compared with the beverage with EGCG alone. These findings (modulation of plasma EGCG level by CAF) provide ideas for modulating the bioavailability of tea catechins, which can be applied to tea-related drinks and foods.

Simultaneous determination of tocols, γ-oryzanols, phytosterols, squalene, cholecalciferol and phylloquinone in rice bran and vegetable oil samples.

In this study, a simultaneous analytical method of tocols, γ-oryzanols, phytosterols, squalene, cholecalciferol and phylloquinone were developed using HPLC-DAD-FLD. The developed method allowed the quantification of 18 compounds in 30 min. Method validation showed linearity of calibration curves (α = 0.05). RSD of intra-day, inter-day and inter-laboratory precision were less than 4.88%. The limit of detections (LODs) and limit of quantifications (LOQs) were low (0.009-2.166 µg g-1) with recoveries around 96.0-102.9%. Results derived from the established method demonstrated a wide variation of detected compounds in rice bran and vegetable oil samples (22.4-1774.6 µg g-1 tocols, ND-26484 µg g-1 γ-oryzanols, ND-12655 µg g-1 phytosterols, ND-3189 µg g-1 squalene, ND-105.3 µg g-1 cholecalciferol, and ND-54.4 µg g-1 phylloquinone). Thus, the developed HPLC-DAD-FLD method is a powerful analytical tool for the above mentioned compounds useful in food and pharmaceutical application.

Effect of wall materials on some physicochemical properties and release characteristics of encapsulated black rice anthocyanin microcapsules.

Anthocyanin-rich extracts derived from the bran of a select Thai black rice cultivar, Leum Pua, were encapsulated by spray drying. In this microencapsulation method, twelve wall materials comprising maltodextrin (M), gum arabic (G), whey protein isolate (W), and their combinations were employed. For comparison, an ionic gelation method was employed using alginate-whey protein isolate (AL-W) as a wall material. Thirteen microcapsules were analysed by scanning electron microscopy, and their encapsulation efficiencies were almost 100%, except for that of the W (83%). M (88%) and its combination with W exhibited the best anthocyanin retention, while pure W contained the highest amount of total phenolic compounds and antioxidant activity. An in vitro digestion study using simulated gastrointestinal fluids revealed the highest released total phenolic content and antioxidant activity of the intestinal fluids when the microcapsule was spray-dried with pure W.

Cold plasma treatment to improve germination and enhance the bioactive phytochemical content of germinated brown rice.

The changes in the bioactive phytochemicals of six cultivars of Thai germinated brown rice (GBR) were monitored in parallel to those of cold plasma-treated GBR (PGBR). After treatment with the optimal plasma conditions, the germination percentage, root length, and seedling height measurements of the most sensitive rice cultivar increased by 84%, 57%, and 69%, respectively. For all of the rice cultivars, there were no significant differences in the antioxidant activities of the GBRs and PGBRs. Conversely, higher contents of γ-oryzanols were observed in the PGBR group than in the GBR group during the 2-day germination period. Certain cultivars in the PGBR group reached their maximum values for total phenolic compounds, total vitamin E, certain simple phenolics, phytosterols, triterpenoids, and anthocyanins one day earlier than the same values for GBR. In contrast, the concentrations of 2-acetyl-1-pyrroline in both the GBR and PGBR samples were reduced significantly with increased germination time.

Tocotrienol inhibits secretion of angiogenic factors from human colorectal adenocarcinoma cells by suppressing hypoxia-inducible factor-1alpha.

Tocotrienol (T3), unsaturated vitamin E, has recently gained considerable attention as a potent antiangiogenic agent minimizing tumor growth, the exact intracellular mechanisms of which remain poorly understood. Because hypoxia-inducible factor-1alpha (HIF-1alpha), its downstream target vascular endothelial growth factor (VEGF), and other angiogenic factors such as interleukin-8 (IL-8) and cyclooxygenase 2 (COX-2) play critical roles in neovascularization, we tested the hypothesis that the inhibitory effect of T3 on tumor angiogenesis is via regulation of these angiogenic factors. We used 2 cancer cell lines, human colorectal adenocarcinoma cells (DLD-1) and human hepatoma cells (HepG2). T3 isomers (2 micromol/L) inhibited hypoxia-induced VEGF secretion from DLD-1, with delta-T3 showing potent inhibition. Delta-T3 suppressed hypoxia-induced VEGF and IL-8 expression in DLD-1 at both mRNA and protein levels, and we found the inhibitory mechanism of delta-T3 by reducing HIF-1alpha protein expression or increasing HIF-1alpha degradation. Also, delta-T3 (2 micromol/L) did not affect hypoxia-induced COX-2 mRNA expression; however, delta-T3 tended to suppress (P = 0.044) hypoxia-induced COX-2 protein expression, implying a possible post-transcriptional mechanism by delta-T3. Overall, our results confirmed that T3 has an inhibitory effect on angiogenic factor secretion from cancer cells and revealed the possible mechanisms, providing new information about the antiangiogenic effects of T3.

Development of a high-performance liquid chromatography-based assay for carotenoids in human red blood cells: application to clinical studies.

Peroxidized phospholipid-mediated cytotoxicity is involved in the pathophysiology of many diseases; for example, there is an abnormal increase of phospholipid hydroperoxides in red blood cells (RBCs) of dementia patients. Dietary carotenoids have gained attention as potent inhibitors of RBC phospholipid hydroperoxidation, thereby making them plausible candidates for preventing disease. However, the occurrence of carotenoids in human RBCs is still unclear. This is in contradistinction to plasma carotenoids, which have been investigated thoroughly for analytical methods as well as biological significance. In this study, we developed a method to analyze RBC carotenoids using high-performance liquid chromatography (HPLC) coupled with ultraviolet (UV) diode array detection (DAD) and atmospheric pressure chemical ionization (APCI) mass spectrometry (MS). Under optimized conditions that included extraction, separation, and detection procedures, six carotenoids (lutein, zeaxanthin, beta-cryptoxanthin, alpha-carotene, beta-carotene, and lycopene) were separated, detected by DAD, and concurrently identified based on APCI/MS and UV spectra profiles when an extract from human RBCs was subjected to HPLC-DAD-APCI/MS. The amounts of carotenoids varied markedly (1.3-70.2 nmol/L packed cells), and polar oxygenated carotenoids (xanthophylls) were predominant in RBCs. The HPLC-DAD-APCI/MS method would be a useful tool for clinical studies for evaluating the bioavailability of RBC carotenoids.