Human oral bioavailability and pharmacokinetics of tocotrienols from tocotrienol-rich (tocopherol-low) barley oil and palm oil formulations
Introduction
The natural vitamin E family comprises eight chemically distinct molecules (Fig. 1): α-, β-, γ-, and δ-tocopherol (α-, β-, γ-, δ-T); and α-, β-, γ-, and δ-tocotrienol (α-, β-, γ-, δ-T3). These tocochromanols contain a polar chromanol head group with a long isoprenoid side chain. Depending on the nature of the isoprenoid chain, a distinction is made between tocopherols (Ts, containing a saturated phytyl chain) or tocotrienols (T3s, unsaturated geranylgeranyl chain) (Dörmann, 2007). The α-, β-, γ-, and δ-vitamers are determined by the number and position of methyl substituents in the chromanol nucleus. Results from human studies suggest that tocotrienols have biological and health effects (Aggarwal et al., 2010, Chen et al., 2011); for example, a blood cholesterol lowering effect, anticancer and tumour suppressive activities, and antioxidant properties. Furthermore, numerous animal studies indicate that α-T3 (not γ-T3) exhibits neuroprotective effects at the nanomolar level (Aggarwal et al., 2010). The uptake of T3s into membranes is faster and higher than that of Ts (Viola et al., 2012), and intramembrane mobility and collision rates with free radicals are higher; these results from in vitro studies may partly explain the special biological activities of T3s (Serbinova, Kagan, Han, & Packer, 1991). Moreover, the plasma pyruvate kinase activity of rats fed α-T3 was similar to that of rats fed α-T. These data may indicate that both α-T3 and α-T act as potent antioxidants in vivo (Ikeda et al., 2003). Furthermore, studies recently reviewed by Nakamura and Omaye (2009) also indicate that T and T3 function as non-antioxidants, which may help control reactive oxygen species (ROS) by the inhibition of ROS generating enzymes. Therefore, there is great interest in the utilisation and commercialisation of these multiple, disease-preventing functional properties in T3 dietary supplements or functional foods.
Current commercial sources of T3s are palm, rice, and annatto, and the most common source is palm oil from large-scale oil palm plantations. Crude palm (Elaeis guineensis) oil (total T3: 364 mg/kg) is particularly rich in γ-T3 (39% of the average total tocochromanol contents of 587 mg/kg) (McLaughlin & Weihrauch, 1979). Rice bran oil, a by-product of the rice milling industry, is a major source of γ-T3 but is low in α-T3 (total T3: 466 mg/kg, total T + T3: 860 mg/kg); and annatto seeds, which are essentially tocopherol-free, naturally contain only δ-T3 (90% of total T3, total T3: 1400 mg/kg) and γ-T3 (10% of total T3) (Aggarwal et al., 2010, Frega et al., 1998). T3-rich sources endemic to Europe are certain cereals like barley and rye. Barley (Hordeum vulgare L.) is unique because it contains all eight vitamers, with T3s contributing about 76% to the total tocochromanols and α-T3 comprising the largest proportion (47%) of the total tocochromanols (Andersson et al., 2008). Recently, dried brewer’s spent grain, a barley by-product of the brewing industry available in large volumes, has been shown to be a feasible feedstuff for the production of an α-T3-rich barley oil with more than 700 mg/kg tocochromanol (Bohnsack, Ternes, Büsing, & Drotleff, 2011). Tocotrienols from barley oil can easily be isolated by molecular distillation (Liu, Shi, Posada, Kakuda, & Xue, 2008) for application in highly concentrated food supplements. In this respect, barley oil appears to be an interesting alternative to the market leader, palm oil. Barley oil may thus represent a potentially profitable innovation in the growing European functional foods market (Annunziata & Vecchio, 2011).
The bioavailability of bioactive compounds is a key determinant for their health effects. The different T3-vitamers have been shown to differ considerably in their bioavailabilities. Yap, Yuen, and Lim (2003) gave mixed T3s to rats and found that the absolute oral bioavailability of α-T3 was 28%, while that of both γ-T3 and δ-T3 was 9%. These values were estimated by dividing the total area under the plasma concentration–time curve (AUC0–∞) obtained from intra-gastric administration by that from the AUC0–∞ for intravenous administration. In humans, T3s were detected in postprandial plasma (Fairus et al., 2006, Fairus et al., 2012), where they were high mainly in triacylglycerol-rich particles, high-density lipoproteins (HDL), and low-density lipoprotein (LDL) after intervention with the palm T3-rich fraction (with single doses of 526 mg and 1011 mg of the palm T3-rich fraction containing about 32% α-T), although at concentrations significantly lower than α-T. The AUC0–8h for plasma α-T3 was the largest of all T3s: about 60% larger than for γ-T3. T3s were not detected in plasma in the fasted state. The authors of those studies found that bio-discrimination between vitamin E forms influences the rate of T3 absorption, mainly because the affinity of α-T (100%) for the α-T transfer protein (α-TTP), which mediates secretion of vitamers from the liver into the circulating system, is much higher than that of α-T3 (12%) or other T3s. There is presumably interference of dietary α-T on T3 bioavailability, as α-T may out-compete T3s for binding at the α-TTP because of its preferential selectivity for α-T. Moreover, α-T has been reported to attenuate the cholesterol-lowering effect of T3s through activation of the HMG-CoA-reductase activity (whereas T3s have a desirable inhibiting effect) (Khor & Ng, 2000). Therefore, application of T3 preparations with minimal concentrations of T in the mixture has been recommended (Gee, 2011, Qureshi et al., 2002), and T3 formulations of palm oil origin which are low in T have accordingly now been introduced into the market.
Vitamers which are not transferred from liver to blood via α-TTP are metabolised. Following vitamin E supplementation, the major metabolites found in human urine are short-chain degradation products with an intact chromanol ring, in particular carboxyethyl-hydroxychromans (CEHCs, with a three-carbon side chain) (Fig. 1). While Ts and T3s lead to similar metabolite profiles (Zhao et al., 2010), T3s are thought to be more extensively metabolised than Ts, and this has recently been shown in vivo by comparing γ-T3 and γ-T (Freiser & Jiang, 2009). CEHCs have been shown to be produced in humans from α-, γ-, and δ-tocopherol (Clarke et al., 2006, Schultz et al., 1995, Swanson et al., 1999), as well as from α- and γ-tocotrienol (Lodge, Ridlington, Leonard, Vaule, & Traber, 2001). The process of vitamin E metabolism is initiated following hydroxylation of the terminal (ω-) methyl group by cytochrome P450 and the subsequent removal of carbon units from its side chain by β-oxidation (Birringer, Pfluger, Kluth, Landes, & Brigelius-Flohé, 2002). This results in shortening of the side chain and increasing water solubility of the metabolites, with CEHCs representing the final metabolites in this pathway. CEHCs are subsequently primarily sulfated or glucuronidated and excreted in urine (Lodge et al., 2001). Vitamin E metabolites in this degradation pathway with a longer side chain (for example. the carboxymethylbutyl-hydroxy-chroman, CMBHC, with a five-carbon side chain, is three carbon atoms longer than CEHC) (Fig. 1) have also been detected in human urine, although generally at only 3–6% of the amount of CEHC (Schuelke et al., 2000). As the side chain degradation metabolites still maintain the intact chromanol moiety, they can act as antioxidants or may have other disease preventing properties (Yang, Lee, Sang, & Yang, 2013). Urinary metabolites could be useful markers in nutritional assessment studies on T3, as their levels reflect the metabolised vitamin E. It has been proposed to use α-CEHC (Schultz et al., 1995) and other metabolites as biomarkers to assess vitamin E status in addition to the blood levels of the respective parent tocochromanols (Zhao et al., 2010).
There is very little literature on human (single-dose oral administration) T3 bioavailability (Fairus et al., 2012, Gee, 2011). Most research has focused on the bioavailability of the palm T3-rich fraction which is composed in the characteristic, γ-T3-dominated pattern. However, to the best of our knowledge, there are no studies comparing the bioavailability of T3 preparations with different vitamer patterns that are also low in T. In order to meet the current recommendations mentioned above, we sought to compare the bioavailability of T3s from naturally α-T3-rich barley oil extract with that of T3s from γ-T3-dominated palm oil extract, both of which were low in T. We therefore orally administered a single dose of vitamin E-rich extract from barley oil known for its high α-T3 content (Bohnsack et al., 2011) to healthy volunteers, and then determined the tocochromanol concentrations in plasma and the main metabolites of T3 in urine by liquid chromatography–mass spectrometry (LC–MS) using data-dependent experiments. This experiment was repeated with the same dose of T3s from palm oil extract, which was the only commercially available α-T3-containing extract low in Ts. We examined in particular whether the composition of the T3 vitamers affected the bioavailability of the single vitamers. We expected the bioavailability of total T3 from barley oil to be superior because it is rich in α-T3, the vitamer with the highest bioavailabilty. In this respect, T3s from annatto were not tested, as annatto contains only γ- and δ-T3, which were found not to differ in their bioavailability (Yap et al., 2003), as mentioned above. The results of this examination should be useful for the development of T3-rich formulations from barley oil for nutraceutical or biomedical applications for the prevention of civilisation-related diseases.
Section snippets
Study design and procedure
The study was a single-centre, randomised, two-arm crossover intervention conducted by trained professionals according to standardised methods at the Institute of Food Science of the Leibniz University of Hannover, Germany. The study protocol was approved by the Freiburg Ethics Commission International (FEKI), in Freiburg, Germany. Written informed consent was obtained from all subjects prior to participating in the study in accordance with the principles of the Helsinki Declaration. Seven
Plasma analysis and pharmacokinetics
The results of plasma analysis after administration of T3-rich formulations (low in T) from barley oil and palm oil to healthy volunteers are presented in Table 3. Following administration of the T3-rich formulations (450 mg total T3s, Table 2) from barley oil and palm oil to subjects, α-, β-, γ- and δ-T3 plasma levels increased in both groups. We observed that in all cases plasma concentrations of T3 vitamers reached their maximum between 2 and 2.4 h after supplementation, decreased within 12 h,
Conclusion
Results of plasma and urine analysis indicate that the composition of the mixed-T3 vitamers affects the bioavailability of the single vitamers. In particular, low-level (or absent) α-T is a key determinant for enhanced absorption of ingested T3s. As hypothesised, our data prove that the absorption of total T3 from the barley oil formulation was significantly higher than that from palm oil, due to the higher percentage of α-T3, which is known for the highest oral bioavailability among T3
Acknowledgments
This work is part of the “Food Network” project funded by the German Ministry for Science and Culture of Lower Saxony through the Research Association of Agricultural and Nutritional Science of Lower Saxony (Forschungsverbund Agrar- und Ernährungswissenschaften Niedersachsen, FAEN). The authors also wish to gratefully acknowledge Leiber GmbH (Bramsche, Germany) for their support. Most of all, we would like to thank the participants who contributed their time toward the success of this project.
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2019, Food Research InternationalCitation Excerpt :However, studies have demonstrated the inhibitory effect of γ- and δ-tocopherol through in vitro assays on human lung cancer cells and in vivo on tumors induced in rats (Li et al., 2011; Lu et al., 2010). Another human study suggested that a low level or absence of α-tocopherol enhance the absorption of ingested tocotrienols, mainly α-tocotrienol, after oral supplementation with barley oil and encapsulated palm oil containing 450 mg of total tocotrienols (Drotleff, Bohnsack, Schneider, Hahn, & Ternes, 2014). Oils such as those from buriti and pracaxi could be considered functional foods and are good ingredients for the production of functional foods due to the presence of natural antioxidants (methyl tocols and carotenes) that can improve human health.