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The Bioavailability of Vitamin C from Kiwifruit
CHAPTER SEVEN
The Bioavailability of Vitamin C from Kiwifruit Margreet C.M. Vissers1, Anitra C. Carr, Juliet M. Pullar, Stephanie M. Bozonet Pathology Department, Centre for Free Radical Research, University of Otago, Christchurch, New Zealand 1 Corresponding author: e-mail address: [email protected]
Contents 1. Vitamin C and Scurvy 2. The Biological Activity of Vitamin C 3. The Recommended Dietary Intake 4. Vitamin C and the Prevention of Chronic Diseases 5. Vitamin C and Respiratory Diseases 6. Food Sources of Vitamin C 7. Vitamin C Content of Kiwifruit 8. Effect of Kiwifruit Supplementation on Vitamin C Intake 9. Effect of Kiwifruit Intake on Plasma Vitamin C 10. Effect of Kiwifruit Intake on Tissue Vitamin C Levels 11. Animal Studies with Kiwifruit 12. Effect of Other Plant Components on Uptake of Vitamin C 13. Natural versus Synthetic Vitamin C 14. Conclusion References
126 127 129 130 131 131 133 134 136 137 139 141 142 143 143
Abstract Vitamin C is an essential component of the diet for humans, and an adequate intake is important not only for the prevention of scurvy but also to limit the risk of developing chronic diseases such as heart disease and cancer. To achieve a regular and adequate intake, daily consumption of fresh fruit and vegetables is recommended. The vitamin C content of food varies widely, however, and plasma levels generally reflect the amount consumed, regardless of its origin. Kiwifruit are one of the premier dietary sources of vitamin C, with all commercially important varieties having high content, and with one serving delivering the bulk of the recommended dietary intake. Recent studies have shown that the addition of kiwifruit to a marginal vitamin C diet markedly improves plasma vitamin C levels and can increase them to both healthy and optimal levels.
Advances in Food and Nutrition Research, Volume 68 ISSN 1043-4526 http://dx.doi.org/10.1016/B978-0-12-394294-4.00007-9
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2013 Elsevier Inc. All rights reserved.
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Vitamin C (ascorbic acid (AA), ascorbate) is a small, water soluble molecule synthesized from simple sugars in plants and animals (Levine, 1986; Loewus, Wagner, & Yang, 1975; Tsao, 1997). It is found in all cells at relatively high (1–10 mM) concentrations (Tsao, 1997). The biosynthetic pathway differs for plants and animals, but the end product remains the same (Loewus et al., 1975; Tsao, 1997). Most animals make ascorbate in the liver and from there it is transported around the body via the circulation and taken up into other tissues (Banhegyi, Braun, Csala, Puskas, & Mandl, 1997; Chatterjee, Majumder, Nandi, & Subramanian, 1975). Humans, however, have lost this ability, due to mutation in the gene for L-gulonolactone oxidase, the enzyme that catalyzes the terminal step in the biosynthetic pathway (Levine, 1986; Sauberlich, 1997). We share this mutation with a number of other species, notably other primates, guinea pigs, and fruit bats (Burri & Jacob, 1997). Together with these animals, we must obtain ascorbate from the diet and thus it is a vitamin. In plants, ascorbate is associated with chloroplasts, together with an ascorbate peroxidase enzyme (Loewus et al., 1975). It is thought to play an important role in the synthesis of the plant cell wall and may also have an antioxidant role, but its functions in plants are poorly understood (Rautenkranz, Li, Machler, Martinoia, & Oertli, 1994; Smirnoff & Pallanca, 1996). In animals, ascorbate plays a many-faceted role as a result of its activity as an enzyme cofactor, with purported involvement in hormone, neurotransmitter, and carnitine synthesis, and collagen cross-linking (Burri & Jacob, 1997; Diliberto, Daniels, & Viveros, 1991; Eipper & Mains, 1991; Levine, 1986; Nelson, Pruitt, Henderson, Jenness, & Henderson, 1981). It is also a versatile one- and two-electron reducing agent, and as such, is thought to be a major antioxidant (Levine, 1986; Tsao, 1997). However, despite extensive documentation of its ability to act as an antioxidant in vitro, there is little direct evidence that demonstrates that this is a major activity in vivo (Halliwell & Gutteridge, 1989). Dietary vitamin C can come from any number of foods containing ascorbate, be they plant or animal in origin, and intake depends on the type of food eaten. This chapter will first briefly explain the human need for adequate dietary intake of vitamin C and will then examine the qualities of kiwifruit that make it an exceptional source of this vitamin.
1. VITAMIN C AND SCURVY Vitamin C became well known in the 1930s when it was identified as the single agent that could alleviate scurvy, an acute disease that had plagued humanity when fresh food was in limited supply, notably in times of war and
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famine (Sauberlich, 1997). Long sea voyages were feared because they were associated with the devastation of scurvy, which begins with depression, general malaise, and joint pain, but rapidly progresses to intestinal bleeding, difficulty with breathing, and ulceration in the gums and other parts of the body and ultimately results in death (Lind, 1753; Sauberlich, 1997). The original report written by James Lind states “It is not easy to conceive a more dismal and diversified scene of misery” (Lind, 1753). Although it is commonly thought that scurvy is a condition limited to seafarers of old, it should be noted that it was probably more easily recognized in this group because of its synchronous onset and devastating loss of life. Its occurrence in individuals in the general population is less easily diagnosed, but scurvy will occur in anyone whose intake of vitamin C from food is about 10 mg/day or less (Sauberlich, 1997). Case reports do appear in the modern literature and are a reminder that vitamin deficiencies still occur (Ho, Prinsloo, & Ombiga, 2007), thereby emphasizing the need for constant daily replenishment of the vitamin. A more common condition is likely to be hypovitaminosis C that occurs when plasma levels fall below about 25 mM for any extended period and this can be associated with poor wound healing, mood disorders, and general lassitude (Hodges, Hood, Canham, Sauberlich, & Baker, 1971; Kinsman & Hood, 1971; Schleicher, Carroll, Ford, & Lacher, 2009; Zhang, Robitaille, Eintracht, & Hoffer, 2011).
2. THE BIOLOGICAL ACTIVITY OF VITAMIN C The scurvy symptoms described above are associated with the function of ascorbate as a cofactor for a number of enzymes with critical functions throughout the body (summarized in Table 7.1). Prominent among these are the copper-containing hydroxylases present in the brain and adrenal glands that catalyze the synthesis of adrenalin and peptide hormones (Diliberto et al., 1991; Eipper & Mains, 1991) and the iron-containing proline and lysine hydroxylases that form collagen cross-links (Burri & Jacob, 1997; Levine, 1986) and generate carnitine (Nelson et al., 1981). The very high concentrations of ascorbate in the adrenals, brain, and other tissues are thought to reflect the requirement for ascorbate to maintain these vital functions. In recent years, a number of new enzymes have been identified that also require ascorbate as a cofactor (Knowles et al., 2003; Ozer & Bruick, 2007; Schofield & Ratcliffe, 2004). Most of these belong to the family of Fe-containing 2-oxoglutrate-dependent dioxygenases, and they have important roles to play in the maintenance of normal metabolism and cellular responses to environmental stresses (Table 7.1).
Table 7.1 Enzymes for which ascorbate acts as a cofactor and their biological functions Enzyme groups Activity/function Biological activities affected
References
Proline hydroxylase C-P4H-1, lysine hydroxylase
Cross-linking of collagen
Maintenance of interstitial tissue structures. Development of scar tissue, wound healing. Integrity of blood vessels and cartilage.
Burri and Jacob (1997)
g-Butyrobetaine hydroxylase, trimethyllysine hydroxylase
Synthesis of carnitine
Transport of long chain fatty acids across the mitochondrial inner membrane. Energy metabolism.
Nelson et al. (1981) and Levine (1986)
Dopamine b-hydroxylase
Generates norepinephrine from dopamine
Synthesis of adrenalin in the central nervous system and adrenal medulla. Effects on energy and vigor.
Diliberto et al. (1991)
Peptidyl-glycine alpha-amidating monooxygenase
Synthesis of neurotransmitters and neuroendocrine peptides
Essential functioning of the nervous and endocrine systems.
Eipper and Mains (1991)
HIF hydroxylases (a family of three proline hydroxylases and one asparagine hydroxylase)
Hydroxylation of the alpha chain of hypoxia-inducible factor 1
Regulation of the transcription of genes under the control of hypoxia-inducible factor. This affects cell responses to hypoxic and metabolic stress with particular modification of cell life and death pathways, angiogenesis, and energy metabolism. Highly implicated in cancer survival and development of chronic diseases.
Knowles, Raval, Harris, and Ratcliffe (2003) and Schofield and Ratcliffe (2004)
Jumonji histone demethylating dioxygenases (up to 12 identified)
Histone demethylation
Regulation of the interaction between histones and DNA, affecting chromatin organization and also gene expression. Wide-ranging consequences as yet uncharacterized.
Ozer and Bruick (2007)
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3. THE RECOMMENDED DIETARY INTAKE Since the identification of ascorbate deficiency, there has been debate as to what amount should be recommended for the daily diet (Carr & Frei, 1999; Levine, 1986; Levine, Dhariwal, Welch, Wang, & Park, 1995). The principle behind the original recommended dietary intake (RDI) recommendations has been that intake should be sufficient to prevent the development of scurvy and that excess is to be avoided. Hence, the recommended amount has been set at a level that ensures there is minimal excretion in the urine while still preventing the development of scurvy. However, a significant case has been made that this is insufficient for good health (Carr & Frei, 1999; Levine et al., 1995); and in 2000, the recommended level in the United States was increased from 60 mg/day to 90 mg for men and from 45 to 75 mg/day for women. Higher levels are recommended for smokers and pregnant and lactating women to offset additional demand. In Australasia, the RDI (Vitamin C, 2006) remains at 45 mg/day. Much of the discussion around the setting of the RDI has focused on the pharmacodynamics of ascorbate in other species. It is notable that those animals that synthesize vitamin C produce enormously high levels daily (Chatterjee, 1973). A goat of 70 kg body mass—equivalent to the average human adult—produces 13,000 mg of ascorbate daily, as well as eating a plant diet rich in vitamin C (Chatterjee, 1973). The other species that share gulonolactone oxidase deficiency consume 20- to 80-fold more vitamin C than do humans, on a body weight basis (Milton, 1999). While this is thought to indicate that the RDI is currently set too low, it has been argued that human utilization of vitamin C is much lower than for other species and that therefore a much lower daily amount should be recommended. Human studies from the United States have indicated that saturation of both plasma and tissues occurs with a daily intake of 200–400 mg/day (Carr & Frei, 1999; Levine, Wang, Padayatty, & Morrow, 2001; Levine et al., 1996). There is concern that excessive intake of vitamin C will result in toxicity. In support of this, there is some evidence that high intake is contraindicated for individuals with iron overload conditions such as hemochromatosis, or with kidney failure (Cook & Reddy, 2001; Naidu, 2003; Padayatty et al., 2010). However, with the exception of these relatively uncommon conditions, it appears that extraordinarily high doses of vitamin C are well tolerated and that any excess is efficiently cleared through the kidneys
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(Naidu, 2003; Padayatty et al., 2010). Basal plasma levels do not rise above 100 mM even when gram doses are consumed, and this has given rise to the idea that taking vitamin C supplements will not be effective in raising body stores (Mandl, Szarka, & Banhegyi, 2009; Padayatty et al., 2004). However, if plasma levels are less than saturating, supplementation with vitamin C will result in increased body status (Carr & Frei, 1999; Mandl et al., 2009).
4. VITAMIN C AND THE PREVENTION OF CHRONIC DISEASES Whereas a small amount of vitamin C daily will prevent the onset of scurvy, there is a growing body of evidence indicating that maintaining plasma levels at closer to saturation levels offers protection from some chronic diseases (Carr & Frei, 1999; Levine & Eck, 2009; Ye & Song, 2008). It is fair to say that there has been enormous confusion about the evidence to support this view and that many population-based studies are difficult to interpret (Kushi, Fee, Sellers, Zheng, & Folsom, 1996; Losonczy, Harris, & Havlik, 1996; Muntwyler, Hennekens, Manson, Buring, & Gaziano, 2002). Most of the claims come from studies in which the intake of vitamin C is monitored and compared with the disease incidence. These prospective studies have generally supported an association of higher vitamin C intake with decreased incidence of cancer, heart disease, and cataract formation (Knekt et al., 2004; Roberts, Traber, & Frei, 2009; Sesso et al., 2008). A meta-analysis of 14 cohort studies has concluded that low dietary vitamin C resulted in increased risk of coronary heart disease (Ye & Song, 2008). However, the results of intervention studies in which healthy individuals were given supplementary vitamin C over and above their normal intake, are more equivocal (Sesso et al., 2008). Only a few have shown any health benefits (Roberts et al., 2009). This is thought to be related to the fact that the study populations were already healthy, making the supplement redundant (Roberts et al., 2009). Many of the studies referred to above have shown that protection from cardiovascular disease requires an average daily intake of 400 mg vitamin C per day. This is equivalent to the amount required for fully saturated plasma and tissue levels and supports the case being made for an increased RDI (Frei, Birlouez-Aragon, & Lykkesfeldt, 2012; Levine & Eck, 2009). Similar data have been obtained for the prevention of stroke and many types of
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cancer, with studies showing a decrease in risk for these diseases generally requiring higher daily intake, often between 100 and 500 mg daily (Cho et al., 2006; Kromhout, 1987; Michels et al., 2001; Myint et al., 2008; Yokoyama et al., 2000; Zhang et al., 1999).
5. VITAMIN C AND RESPIRATORY DISEASES Possibly, the most discussed and the most investigated aspect of vitamin C has been its effect on the common cold. Although the conditions investigated have varied widely, there is a general consensus that supplemental vitamin C does not protect an already well-nourished population from contracting the cold virus, but may shorten the duration of symptoms (Douglas, Hemila, Chalker, & Treacy, 2007; Hemila, 2009). Supplementation with vitamin C has been found to decrease the susceptibility to a cold in people who are stressed by extreme exercise (marathon runners) or cold exposure (arctic explorers) (Douglas et al., 2007). A few studies have shown a more marked effect with severe respiratory illness, indicating that vitamin C may have a significant benefit in both preventing and treating pneumonia (Hemila & Louhiala, 2007). More research is needed in this area, but the link with acute respiratory illness is of interest.
6. FOOD SOURCES OF VITAMIN C All living things contain vitamin C, and therefore, it is available to some extent in most food sources (The Natural Food Hub, 2001; U.S. Department of Agriculture, 2011). Its availability is determined by both the content of the food and also the method of preparation. Ascorbate is readily oxidized and can be destroyed by exposure to oxygen, heat, and some metal ions. Thus, cooking can decrease the amount of available vitamin C and meat, which can contain substantial amounts of vitamin C, does not deliver significant vitamin C as this vitamin is destroyed in the preparation process (U.S. Department of Agriculture, 2011). In addition, organ meats such as liver and brain, which contain more vitamin C than muscle, are not often eaten in the Western diet. Fresh food ensures the best delivery and, as fruit is generally consumed fresh, this is by far the most effective way to achieve an adequate daily intake (The Natural Food Hub, 2001). It should be noted
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that many foods deliver some vitamin C, and the key to effective intake is to ensure a good variety of fruit and vegetables in the daily diet. The vitamin C content of some common foods is shown in Table 7.2. From this list, it can be seen that kiwifruit is rated as one of very few foods that are considered to be an exceptional source of vitamin C, with a single Table 7.2 The vitamin C content of some common foods, based on content and serving size Rating as a source Vitamin C content Vitamin C per of vitamin C Food (mg/100 g) servinga (mg)
Guava
183
165
Exceptional
Kiwifruit, gold
105–120
95–108
Exceptional
Capsicum
180
95
Exceptional
Acerola
1677
80
Exceptional
Kiwifruit, green
65–90
50–74
Exceptional
Orange
53
70
Excellent
Mango
28
57
Excellent
Broccoli
90
51
Excellent
Rosehip
1500
45
Excellent
Grapefruit
34
44
Excellent
Brussels sprouts
80
40
Excellent
Persimmon
40
40
Excellent
Watermelon
10
27
Very good
Mandarin
31
26
Very good
Tamarillo
31
22
Very good
Tomato
19
23
Very good
Cabbage greens
30
20
Very good
Feijoa
31
16
Very good
Potato
20
15
Very good
Pineapple
15
13
Good
Banana
9
11
Good
Strawberry
57
7
Good
Apple
6
8
Fairly good
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Table 7.2 The vitamin C content of some common foods, based on content and serving size—cont'd Vitamin C content Vitamin C per Rating as a source Food (mg/100 g) serving (mg) of vitamin C
Grape
11
6
Fairly good
Carrot
9
5
Good
Lettuce
4
4
–
Cucumber
3
1
–
Avocado
8
2
–
Onion
7
1
–
Cow’s milk (fresh)
2
–
–
Nuts
–
–
–
Grains
–
–
–
Seeds (including peas and beans)
–
–
–
Data in this table are taken from The Natural Food Hub, 2001 and U.S. Department of Agriculture, 2011 a One serving is defined as the amount normally eaten and varies for different food types, for example, one slice of watermelon or avocado, two mandarins, ½ cup cooked vegetables, ½ cup grapes.
serving providing almost the entire RDI as determined in the United States, and twice the Australasian RDI (Vitamin C, 2006). The exceptional label is a composite of the amount of vitamin C present in the food per unit weight and the likely serving size (The Natural Food Hub, 2001; U.S. Department of Agriculture, 2011). Therefore, some foods that have very high vitamin C content, such as blackcurrants, strawberries, or lemons, are not considered a highly effective dietary source because less is consumed at any one time. It should also be noted that most fruits generally have higher vitamin C content than vegetables. Legumes such as peas and beans, nuts, grains, and seeds do not contain measureable amounts of vitamin C. This, together with the destruction of vitamin C in meat upon cooking, means that delivery of vitamin C to the diet is mostly, if not entirely, from fruit and vegetables, with fruit being the major contributor.
7. VITAMIN C CONTENT OF KIWIFRUIT There are many varieties of kiwifruit and their vitamin C content differs markedly. An analysis of more than 20 varieties reported a range of 25–205 mg/ 100 g (Hunter, Greenwood, Zhang, & Skinner, 2011; Nishiyama et al., 2004).
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These levels reflect the total AA content of the fruit, which is a combination of both AA and its oxidized form, dehydroascorbic acid (DHAA) (Nishiyama et al., 2004). Because it is known to be readily reduced in the body, DHAA is generally considered as part of the available vitamin C. However, animal studies have indicated that, when given orally, DHAA has limited antiscorbutic properties, and may therefore not be biologically available (Ogiri et al., 2002). For this reason, it is probably more accurate to consider only the AA content when estimating the effective vitamin C content. There is a large degree of variability in the proportion of DHAA present in the different kiwifruit cultivars (Nishiyama et al., 2004). The reasons for this variation are unknown, but may reflect levels of plant stress or inherent differences in the amount of the enzyme ascorbate oxidase, which catalyzes the oxidation of AA to DHAA. In the most common commercial cultivar, Actinidia deliciosa cv. ‘Hayward’ (green kiwifruit), approximately 15% of the total AA is present as DHAA, and a higher level (up to 30%) is found in the Actinidia chinensis cv. ‘Hort 16A’ (gold kiwifruit) (Nishiyama et al., 2004). This means that the effective concentration of AA for these two commercial cultivars is likely to be around 55–65 mg/100 g for the green and 75–85 mg/100 g for the gold. In agreement with this, our laboratory has measured 89 mg AA/ 100 g of gold kiwifruit, which is equivalent to about 80 mg per fruit. The vitamin C content of fruit is often found to decrease upon storage. This was shown not to be a significant factor in the main kiwifruit varieties, with levels remaining stable during postharvest ripening (Nishiyama et al., 2004). However, we have measured a decline in AA levels of around 30% after extended long-term chilled storage of around 9 months postharvest (M. Vissers and L. Braithwaite, unpublished data).
8. EFFECT OF KIWIFRUIT SUPPLEMENTATION ON VITAMIN C INTAKE The information above suggests that the addition of kiwifruit to the diet would contribute significantly to overall vitamin C intake. We have recently been able to demonstrate just how effective this can be with a human supplementation study. The study population comprised 14 nonsmoking healthy male university students aged between 18 and 30 years who had an average baseline fasting plasma ascorbate level of 38 mM. The participants kept detailed food and beverage diaries, which indicated a mean baseline intake of 3.9 0.5 servings of fruit and vegetables per day (Fig. 7.1A). Based on the specifics of the food consumed, this was estimated
Vitamin C from Kiwifruit
135
Figure 7.1 The effect of kiwifruit consumption on dietary vitamin C intake in humans. Data from a human intervention study involving 15 males aged 18–35 are shown. (A) Daily servings of fruit and vegetables consumed by the participants were estimated from weekly food diaries. Addition of kiwifruit to the diet increased the baseline consumption from four to six serves with the addition of two or three kiwifruit (KF) daily. (B) Vitamin C intake was significantly increased with the addition of even one half a KF per day. After washout, levels returned to baseline. Data represent means SEM from a group of 15 participants. indicates statistical significance (P < 0.05) from baseline as determined by one-way repeated measures analysis of variance with Fisher LSD pairwise multiple comparison procedure.
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to contribute 44 5 mg ascorbate per day, which is the equivalent of the Australasian RDI and indicates an average of around 11 mg vitamin C per serving of fruit or vegetables. When kiwifruit was given daily, it appeared that some individuals consumed this in addition to their daily diet, whereas others reduced their intake of other fruit and vegetables to accommodate their increased kiwifruit consumption. Consequently, addition of one half or one kiwifruit per day did not alter the participants’ average number of fruit and vegetable servings. Addition of two and three kiwifruit to the daily diet resulted in only a small increase, to a maximum of six serves per day at the three kiwifruit dose. Despite the lack of an increase in fruit and vegetable servings at the lowest doses, supplementation with one half and one kiwifruit per day significantly increased the average daily intake of vitamin C to around 80 and 130 mg, respectively (Fig. 7.1B). The latter is equivalent to the amount recommended for prevention of chronic disease (Carr & Frei, 1999; Padayatty & Levine, 2008) and reflects the high ascorbate content in the fruit. Total intake reached 220 mg/day when two kiwifruit per day were consumed (Fig. 7.1B). This is comparable to the intake required to achieve plasma saturation and is now considered by some to be optimal (Frei et al., 2012; Levine & Eck, 2009; Padayatty & Levine, 2001). These results clearly demonstrate the value and effectiveness of adding a high vitamin C-containing food like kiwifruit to the diet.
9. EFFECT OF KIWIFRUIT INTAKE ON PLASMA VITAMIN C In recent years, several studies have monitored plasma vitamin C in individuals supplemented with kiwifruit. In most of these studies, the objective was to monitor specific biological effects of nutrition and in these studies, kiwifruit was found to improve digestive health (Rush, Patel, Plank, & Ferguson, 2002), enhance iron uptake (Beck, Conlon, Kruger, Coad, & Stonehouse, 2011), increase DNA repair activity (Brevik et al., 2011, see also Chapter 16), modulate lipid profiles (Chang & Liu, 2009), and reduce platelet aggregation (Duttaroy & Jorgensen, 2004). Most of these studies showed that consuming kiwifruit resulted in an increase in plasma vitamin C, with saturation levels being reached at a dose of two or three kiwifruit per day. Collins et al. reported a dose-dependent effect upon supplementation with one to three kiwifruit per day (Brevik et al., 2011; Collins, Harrington, Drew, & Melvin, 2003). The increase was rather modest, however, with plasma steady
Vitamin C from Kiwifruit
137
state levels increasing by 7–16 mM (Brevik et al., 2011). These small increases are likely to reflect the fact that the participants already had saturating plasma vitamin C levels of about 60 mM at baseline (Brevik et al., 2011). To more accurately measure the contribution of kiwifruit to dietary vitamin C intake, we recently monitored plasma vitamin C levels in a group of 14 male students with an average baseline plasma vitamin C level of 38 mM (Carr, Pullar, Moran & Vissers, 2012). We were able to measure a strong effect of kiwifruit supplementation on plasma vitamin C levels, with increases of around 25 mM (Fig. 7.2A). When a subpopulation with baseline plasma vitamin C levels of 28 mM was investigated, we measured an increase of 22–43 mM with supplementation of one half to three kiwifruit per day. The effect was dose-dependent and was seen after the first week of supplementation. At two kiwifruit per day, plasma levels approached saturation with no further increase with three per day, although some noncompliance was evidenced at this highest dose. Plasma saturation was confirmed by monitoring urinary excretion, which showed that there was no increase until a dose of two kiwifruit per day (Fig. 7.2B). This coincided with plasma levels reaching around 60 mM. During the washout period of the study, plasma and urine levels rapidly returned to baseline (Fig. 7.2A and B), which strongly suggests significant daily turnover of body stores.
10. EFFECT OF KIWIFRUIT INTAKE ON TISSUE VITAMIN C LEVELS Tissues throughout the body concentrate ascorbate from the plasma via active transport. The uptake is generally thought to reflect plasma concentrations and some tissues saturate at lower vitamin C intake than is required for plasma saturation (Levine et al., 1996; Levine et al., 2001). Because of difficulty in accessing human tissue, most information about tissues has been obtained from the measurement of white blood cell levels. Leukocytes preferentially take up ascorbate to millimolar intracellular concentrations with saturation levels being reached with a daily intake of around 100 mg/day (Carr & Frei, 1999; Levine et al., 1996; Levine et al., 2001). This corresponds to plasma levels of around 50 mM (Lee, Hamernyik, Hutchinson, Raisys, & Labbe, 1982; Levine et al., 1996; Levine et al., 2001). In agreement with this, we found that the supplementation of one kiwifruit per day to our study
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Figure 7.2 Increase in plasma vitamin C as a result of kiwifruit consumption. (A) Plasma vitamin C levels of a group of 14 young men (aged 18–30 years) with an average baseline plasma vitamin C level of 38 mM were measured after increased intake of kiwifruit in addition to baseline diet. Study participants consumed kiwifruit as indicated for a period of 4–6 weeks and fasting plasma vitamin C levels were monitored weekly. (B) Urinary excretion of vitamin C by the same study participants as a function of daily kiwifruit intake. Data represent mean SEM. indicates statistical significance (P < 0.05) from baseline as determined by one-way repeated measures analysis of variance with Fisher LSD pairwise multiple comparison procedure.
Vitamin C from Kiwifruit
139
population resulted in optimal leukocyte vitamin C, with no further increase seen with two kiwifruit per day. Addition of one kiwifruit corresponded to a total dietary vitamin C intake of around 130 mg/day. That we were able to detect a statistically significant increase in leukocyte vitamin C from baseline levels in our study population is likely due to their low initial levels.
11. ANIMAL STUDIES WITH KIWIFRUIT Monitoring vitamin C levels in tissues other than white blood cells is difficult in humans, and animal studies have provided most of this information. A recently developed transgenic mouse model that lacks gulonolactone oxidase and displays the same phenotype as the human deficiency condition has provided a useful tool with which to assess the pharmacodynamics of vitamin C uptake and metabolism (Maeda et al., 2000; Parsons, Maeda, Yamauchi, Banes, & Koller, 2006). We used this model, the Gulo / mouse, to monitor the effect of variable dietary ascorbate on uptake into the tissues and also to investigate the bioavailability of ascorbate from kiwifruit (Vissers, Bozonet, Pearson, & Braithwaite, 2011). We found that the ascorbate levels differed in the various mouse tissue compartments (Vissers et al., 2011) and were in agreement with the expected relative distributions in the same tissues from studies of other vitamin C-deficient animals, including humans (Tsao, 1997). We monitored plasma, brain, heart muscle, liver, kidney, and white cells in response to varied dietary intake administered through the drinking water. Uptake for all tissues followed a sigmoid-shaped dose–response curve, similar to that observed in human studies. The rate of uptake differed significantly, and in particular, liver, heart, and kidney required around five times more ascorbate than brain tissue to reach saturation levels. We also determined the effectiveness of vitamin C uptake from kiwifruit in the Gulo / mouse and compared this with delivery of reagent vitamin C delivered through the drinking water or embedded in a gel. When mice were fed a fresh kiwifruit gel preparation (either green or gold varieties) of known ascorbate content for 4 weeks, the ascorbate levels measured in blood, liver, kidney, and heart were markedly higher than those measured in animals taking an equivalent dose of purified ascorbate via the water supply or in a gel form (Vissers et al., 2011). A period of 4 weeks was required to reach tissue steady state, which is similar to the time needed for humans. The
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increased bioavailability of vitamin C from kiwifruit was clearly apparent when we compared the dose response. When kiwifruit was used, a significantly (p < 0.05) lower daily ascorbate intake was required to reach 50% of the maximum levels (ED50) in serum, heart, kidney, and liver. This difference was around fivefold for some tissues (e.g., liver). Both varieties of kiwifruit delivered significantly more ascorbate to most tissues when compared directly with an equivalent dose of ascorbate supplied in the drinking water (Table 7.3). These results were somewhat surprising and the suggestion that kiwifruit could be a much better source of dietary vitamin C than a purified supplement is of interest for human nutrition. Our present studies aim to address this question in humans, and the results with the university student population discussed above suggest that the addition of kiwifruit to the diet is certainly a highly effective way to increase plasma and white cell levels. Whether kiwifruit would be more effective in humans than a purified ascorbate supplement is yet to be determined. Table 7.3 Direct comparison of vitamin C levels after supplementation with either reagent ascorbate or green or gold kiwifruit in the Gulo / mouse Gulo / mice Daily vitamin C intake and source Supplement ascorbate 1.7 mg (n ¼ 5)
Green Supplement kiwifruit ascorbate 1.8 mg (n¼4) 5.0 mg (n ¼ 5)
Gold kiwifruit 3.0 mg (n ¼ 4)
Serum
15.0 4.9
34.1 7.9*
24.6 7.8
64.9 8.9*
Brain
51.5 6.2
37.5 1.1*
55.8 1.4
64.7 3.2*
White blood cells
2.3 0.6
3.5 0.7
5.8 0.7
5.0 0.4
Liver
2.5 0.8
9.6 1.7** 10.7 0.7
Heart muscle
1.2 0.4
1.4 0.5
1.6 0.4
1.4 0.5
Kidney
2.4 0.9
4.6 1.1*
3.3 0.2
9.4 2.0*
20.6 2.8**
Vitamin C was supplied in kiwifruit or as a purified compound (sodium ascorbate) through the water supply (supplement) for a period of 4 weeks (Vissers et al., 2011). Measured concentrations as given are mM for serum; nmol/108 cells for white blood cells; and mg/100 mg wet weight for tissues from brain, liver, kidney, and heart. Means SE are shown and significant differences between kiwifruit and ascorbate supplement, as determined using the student’s t-test for unpaired samples, are indicated (*p < 0.05 or **p < 0.01).
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12. EFFECT OF OTHER PLANT COMPONENTS ON UPTAKE OF VITAMIN C In the mouse study described above, we also found that eating kiwifruit could elevate serum and tissue vitamin C levels beyond those seen in a wild-type control mouse that maintains high and stable levels by synthesis in the liver. This suggests that there may be a difference in the efficiency of intestinal absorption and/or clearance through the kidneys in the kiwifruitfed animal. This observation is reminiscent of a speculation often made in the mid twentieth century when it was proposed that some foods contain an unknown factor that enhances the bioavailability of vitamin C. Reference was made to a hypothetical factor described as an “AA-economizing factor,” “vitamin P,” or “vitamin C2” which affected the efficiency of ascorbate uptake (Cotereau, Gabe, et al., 1948). Alternatively, there may be other factors present that could affect the delivery of vitamin C. Kiwifruit are a rich source of vitamin E (up to 10% of the RDI per serving) (Celik, Ercisli, & Turgut, 2007), as well as a range of flavonoids (mostly in the skin) and have a high content of dietary fiber (Hunter et al., 2011; Nishiyama, 2007). Some early studies with people or guinea pigs suggested that ascorbate uptake is enhanced by catechin and other flavonoids (Cotereau et al., 1948), but very little good evidence can be found that this actually occurs (Nishiyama, 2007). In contrast, vitamin E deficiency has been shown to accelerate vitamin C loss in the ODS rat (Tanaka, Hashimoto, Tokumaru, Iguchi, & Kojo, 1997) and, given the high content of vitamin E in kiwifruit (Nishiyama, 2007), it is possible that an additional source of vitamin E may affect vitamin C pharmacokinetics. Thus, it will be of interest to further investigate the effects of vitamin C uptake from kiwifruit in humans. (Note from editors: The ODS, or Osteogenic Disorder Shionogi rat is a wellestablished model animal for studying vitamin C metabolism.) The effect of bioflavonoids on the uptake of ascorbate has been directly investigated in a few studies. In one study, a mixture of synthetic ascorbate and a citrus extract containing a mixture of bioflavonoids, proteins, and carbohydrates was administered to a group of human volunteers (Vinson & Bose, 1988). The ascorbate from the food mix was found to be more slowly absorbed and was effectively 35% more bioavailable, determined by measuring plasma levels and urinary excretion over a 24-h period. Another study found no difference between a synthetic ascorbate
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dose of 500 mg with or without added bioflavonoids (Johnston & Luo, 1994). The main difference between these two apparently contradictory studies was the ratio of bioflavonoids to ascorbate (4:1 in the first study and 0.05:1 in the second). This highlights the importance of determining the optimum study conditions before embarking on human intervention trials, and the results suggest that further investigations may be warranted.
13. NATURAL VERSUS SYNTHETIC VITAMIN C The suggestion above that there may be differences in the uptake and metabolism of vitamin C from foods has been a topic of frequent discussion and some investigations have provided useful information. Chemically speaking, synthetic AA is indistinguishable from the same chemical that is present in plants and animals. There have been a small number of human studies that have directly compared the uptake of synthetic ascorbate and an equivalent amount in citrus fruit and these have found that both contributed equally to plasma vitamin C levels (Gregory, 1993; Mangels et al., 1993; Pelletier & Keith, 1974). Different formulations of synthetic vitamin C are often found in supplements, with the claim being made that some will have better bioavailability (Nyyssonen et al., 1997; Yung, Mayersohn, & Robinson, 1982). These include AA preparations presented as powders, in capsules or tablet form, or as slow-release formulations. It remains unclear whether any one form is more absorbable than another. The biggest difference between these forms may well be the stability of the product during storage, as ascorbate is readily oxidized by exposure to air, heat, and light. For this reason, it is better kept as a capsule or tablet than as a powder when stored over longer periods. When present in a liquid, ascorbate is often more labile and can be destroyed by heating in the commercial preparation of fruit juices. To ensure consistency of these products, synthetic ascorbate is often added back prior to sale. Because vitamin C supplements are often taken in higher bolus amounts than would be found in food, formulations are often made to mitigate the effect of the free acid and to attempt to ensure slower and more prolonged uptake. Mineral salts of AA, most commonly sodium or calcium ascorbate, are neutral and this is thought to lessen bowel irritation, but there is little actual evidence that this does occur. “Ester C” is a mixture of mostly calcium ascorbate, together with some metabolites (DHAA and calcium threonate) and differs from esterified ascorbate, which include mixtures of vitamin C and lipids that are claimed to increase bioavailability. However, in vivo data that would
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determine the relative effectiveness of these products compared with AA either in purified form or from a food source is currently lacking.
14. CONCLUSION Despite the many and varied formulations of vitamin C supplements, it should be noted that all clinical information to date suggests that the ascorbate present in fresh food is bioavailable and readily absorbed and is the preferred source, as it will be delivered together with other important nutrients and minerals. Hence, the best advice that can be given is to encourage the consumption of a wide range of fresh foods and to include in the mix one to two serves per day of a high vitamin C-containing food such as kiwifruit.
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Levine, M. (1986). New concepts in the biology and biochemistry of ascorbic acid. The New England Journal of Medicine, 314, 892–902. Levine, M., Conry-Cantilena, C., Wang, Y., Welch, R. W., Washko, P. W., Dhariwal, K. R., et al. (1996). Vitamin C pharmacokinetics in healthy volunteers: Evidence for a recommended dietary allowance. Proceedings of the National Academy of Sciences of the United States of America, 93, 3704–3709. Levine, M., Dhariwal, K. R., Welch, R. W., Wang, Y., & Park, J. B. (1995). Determination of optimal vitamin C requirements in humans. The American Journal of Clinical Nutrition, 62, 1347S–1356S. Levine, M., & Eck, P. (2009). Vitamin C: Working on the x-axis. The American Journal of Clinical Nutrition, 90, 1121–1123. Levine, M., Wang, Y., Padayatty, S. J., & Morrow, J. (2001). A new recommended dietary allowance of vitamin C for healthy young women. Proceedings of the National Academy of Sciences of the United States of America, 98, 9842–9846. Lind, J. (1753). A treatise of the scurvy in three Parts. Containing an inquiry into the nature, causes, and cure, of that disease, together with a critical and chronological view of what has been published on the subject. Edinburgh: Sands/Murray/Cochran. Loewus, F. A., Wagner, G., & Yang, J. C. (1975). Biosynthesis and metabolism of ascorbic acid in plants. Annals of the New York Academy of Sciences, 258, 7–23. Losonczy, K. G., Harris, T. B., & Havlik, R. J. (1996). Vitamin E and vitamin C supplement use and risk of all-cause and coronary heart disease mortality in older persons: The established populations for epidemiologic studies of the elderly. The American Journal of Clinical Nutrition, 64, 190–196. Maeda, N., Hagihara, H., Nakata, Y., Hiller, S., Wilder, J., & Reddick, R. (2000). Aortic wall damage in mice unable to synthesize ascorbic acid. Proceedings of the National Academy of Sciences of the United States of America, 97, 841–846. Mandl, J., Szarka, A., & Banhegyi, G. (2009). Vitamin C: Update on physiology and pharmacology. British Journal of Pharmacology, 157, 1097–1110. Mangels, A. R., Block, G., Frey, C. M., Patterson, B. H., Taylor, P. R., Norkus, E. P., et al. (1993). The bioavailability to humans of ascorbic acid from oranges, orange juice and cooked broccoli is similar to that of synthetic ascorbic acid. The Journal of Nutrition, 123, 1054–1061. Michels, K. B., Holmberg, L., Bergkvist, L., Ljung, H., Bruce, A., & Wolk, A. (2001). Dietary antioxidant vitamins, retinol, and breast cancer incidence in a cohort of Swedish women. International journal of cancer, 91, 563–567. Milton, K. (1999). Nutritional characteristics of wild primate foods: Do the diets of our closest living relatives have lessons for us? Nutrition, 15, 488–498. Muntwyler, J., Hennekens, C. H., Manson, J. E., Buring, J. E., & Gaziano, J. M. (2002). Vitamin supplement use in a low-risk population of US male physicians and subsequent cardiovascular mortality. Archives of Internal Medicine, 162, 1472–1476. Myint, P. K., Luben, R. N., Welch, A. A., Bingham, S. A., Wareham, N. J., & Khaw, K. T. (2008). Plasma vitamin C concentrations predict risk of incident stroke over 10 y in 20 649 participants of the European Prospective Investigation into Cancer Norfolk prospective population study. The American Journal of Clinical Nutrition, 87, 64–69. Naidu, K. A. (2003). Vitamin C in human health and disease is still a mystery? An overview. Nutrition Journal, 2, 7. Nelson, P. J., Pruitt, R. E., Henderson, L. L., Jenness, R., & Henderson, L. M. (1981). Effect of ascorbic acid deficiency on the in vivo synthesis of carnitine. Biochimica et Biophysica Acta, 672, 123–127. Nishiyama, I. (2007). Fruits of the actinidia genus. Advances in Food and Nutrition Research, 52, 293–324.
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Nishiyama, I., Yamashita, Y., Yamanaka, M., Shimohashi, A., Fukuda, T., & Oota, T. (2004). Varietal difference in vitamin C content in the fruit of kiwifruit and other actinidia species. Journal of Agricultural and Food Chemistry, 52, 5472–5475. Nyyssonen, K., Poulsen, H. E., Hayn, M., Agerbo, P., Porkkala-Sarataho, E., Kaikkonen, J., et al. (1997). Effect of supplementation of smoking men with plain or slow release ascorbic acid on lipoprotein oxidation. European Journal of Clinical Nutrition, 51, 154–163. Ogiri, Y., Sun, F., Hayami, S., Fujimura, A., Yamamoto, K., Yaita, M., et al. (2002). Very low vitamin C activity of orally administered L-dehydroascorbic acid. Journal of Agricultural and Food Chemistry, 50, 227–229. Ozer, A., & Bruick, R. K. (2007). Non-heme dioxygenases: Cellular sensors and regulators jelly rolled into one? Nature Chemical Biology, 3, 144–153. Padayatty, S. J., & Levine, M. (2001). New insights into the physiology and pharmacology of vitamin C. Canadian Medical Association Journal, 164, 353–355. Padayatty, S. J., & Levine, M. (2008). Fruit and vegetables: Think variety, go ahead, eat!. The American Journal of Clinical Nutrition, 87, 5–7. Padayatty, S. J., Sun, A. Y., Chen, Q., Espey, M. G., Drisko, J., & Levine, M. (2010). Vitamin C: Intravenous use by complementary and alternative medicine practitioners and adverse effects. PLoS One, 5, e11414. Padayatty, S. J., Sun, H., Wang, Y., Riordan, H. D., Hewitt, S. M., Katz, A., et al. (2004). Vitamin C pharmacokinetics: Implications for oral and intravenous use. Annals of Internal Medicine, 140, 533–537. Parsons, K. K., Maeda, N., Yamauchi, M., Banes, A. J., & Koller, B. H. (2006). Ascorbic acid-independent synthesis of collagen in mice. American Journal of Physiology, Endocrinology and Metabolism, 290, E1131–E1139. Pelletier, O., & Keith, M. O. (1974). Bioavailability of synthetic and natural ascorbic acid. Journal of the American Dietetic Association, 64, 271–275. Rautenkranz, A., Li, L., Machler, F., Martinoia, E., & Oertli, J. J. (1994). Transport of ascorbic and dehydroascorbic acids across protoplast and vacuole membranes isolated from barley (Hordeum vulgare L. cv Gerbel) leaves. Plant Physiology, 106, 187–193. Roberts, L. J., 2nd, Traber, M. G., & Frei, B. (2009). Vitamins E and C in the prevention of cardiovascular disease and cancer in men. Free Radical Biology and Medicine, 46, 1558. Rush, E. C., Patel, M., Plank, L. D., & Ferguson, L. R. (2002). Kiwifruit promotes laxation in the elderly. Asia Pacific Journal of Clinical Nutrition, 11, 164–168. Sauberlich, H. E. (1997). A history of scurvy and vitamin C. In L. Packer & J. Fuchs (Eds.), Vitamin C in health and disease (pp. 1–24). (1st ed). New York: Marcel Dekker, Inc. Schleicher, R. L., Carroll, M. D., Ford, E. S., & Lacher, D. A. (2009). Serum vitamin C and the prevalence of vitamin C deficiency in the United States: 2003–2004 National Health and Nutrition Examination Survey (NHANES). The American Journal of Clinical Nutrition, 90, 1252–1263. Schofield, C. J., & Ratcliffe, P. J. (2004). Oxygen sensing by HIF hydroxylases. Nature Reviews Molecular Cell Biology, 5, 343–354. Sesso, H. D., Buring, J. E., Christen, W. G., Kurth, T., Belanger, C., MacFadyen, J., et al. (2008). Vitamins E and C in the prevention of cardiovascular disease in men: The Physicians’ Health Study II randomized controlled trial. The Journal of the American Medical Association, 300, 2123–2133. Smirnoff, N., & Pallanca, J. E. (1996). Ascorbate metabolism in relation to oxidative stress. Biochemical Society Transactions, 24, 472–478. Tanaka, K., Hashimoto, T., Tokumaru, S., Iguchi, H., & Kojo, S. (1997). Interactions between vitamin C and vitamin E are observed in tissues of inherently scorbutic rats. The Journal of Nutrition, 127, 2060–2064.
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The Bioavailability of Vitamin C from Kiwifruit Margreet C.M. Vissers1, Anitra C. Carr, Juliet M. Pullar, Stephanie M. Bozonet Pathology Department, Centre for Free Radical Research, University of Otago, Christchurch, New Zealand 1 Corresponding author: e-mail address: [email protected]
Contents 1. Vitamin C and Scurvy 2. The Biological Activity of Vitamin C 3. The Recommended Dietary Intake 4. Vitamin C and the Prevention of Chronic Diseases 5. Vitamin C and Respiratory Diseases 6. Food Sources of Vitamin C 7. Vitamin C Content of Kiwifruit 8. Effect of Kiwifruit Supplementation on Vitamin C Intake 9. Effect of Kiwifruit Intake on Plasma Vitamin C 10. Effect of Kiwifruit Intake on Tissue Vitamin C Levels 11. Animal Studies with Kiwifruit 12. Effect of Other Plant Components on Uptake of Vitamin C 13. Natural versus Synthetic Vitamin C 14. Conclusion References
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Abstract Vitamin C is an essential component of the diet for humans, and an adequate intake is important not only for the prevention of scurvy but also to limit the risk of developing chronic diseases such as heart disease and cancer. To achieve a regular and adequate intake, daily consumption of fresh fruit and vegetables is recommended. The vitamin C content of food varies widely, however, and plasma levels generally reflect the amount consumed, regardless of its origin. Kiwifruit are one of the premier dietary sources of vitamin C, with all commercially important varieties having high content, and with one serving delivering the bulk of the recommended dietary intake. Recent studies have shown that the addition of kiwifruit to a marginal vitamin C diet markedly improves plasma vitamin C levels and can increase them to both healthy and optimal levels.
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Vitamin C (ascorbic acid (AA), ascorbate) is a small, water soluble molecule synthesized from simple sugars in plants and animals (Levine, 1986; Loewus, Wagner, & Yang, 1975; Tsao, 1997). It is found in all cells at relatively high (1–10 mM) concentrations (Tsao, 1997). The biosynthetic pathway differs for plants and animals, but the end product remains the same (Loewus et al., 1975; Tsao, 1997). Most animals make ascorbate in the liver and from there it is transported around the body via the circulation and taken up into other tissues (Banhegyi, Braun, Csala, Puskas, & Mandl, 1997; Chatterjee, Majumder, Nandi, & Subramanian, 1975). Humans, however, have lost this ability, due to mutation in the gene for L-gulonolactone oxidase, the enzyme that catalyzes the terminal step in the biosynthetic pathway (Levine, 1986; Sauberlich, 1997). We share this mutation with a number of other species, notably other primates, guinea pigs, and fruit bats (Burri & Jacob, 1997). Together with these animals, we must obtain ascorbate from the diet and thus it is a vitamin. In plants, ascorbate is associated with chloroplasts, together with an ascorbate peroxidase enzyme (Loewus et al., 1975). It is thought to play an important role in the synthesis of the plant cell wall and may also have an antioxidant role, but its functions in plants are poorly understood (Rautenkranz, Li, Machler, Martinoia, & Oertli, 1994; Smirnoff & Pallanca, 1996). In animals, ascorbate plays a many-faceted role as a result of its activity as an enzyme cofactor, with purported involvement in hormone, neurotransmitter, and carnitine synthesis, and collagen cross-linking (Burri & Jacob, 1997; Diliberto, Daniels, & Viveros, 1991; Eipper & Mains, 1991; Levine, 1986; Nelson, Pruitt, Henderson, Jenness, & Henderson, 1981). It is also a versatile one- and two-electron reducing agent, and as such, is thought to be a major antioxidant (Levine, 1986; Tsao, 1997). However, despite extensive documentation of its ability to act as an antioxidant in vitro, there is little direct evidence that demonstrates that this is a major activity in vivo (Halliwell & Gutteridge, 1989). Dietary vitamin C can come from any number of foods containing ascorbate, be they plant or animal in origin, and intake depends on the type of food eaten. This chapter will first briefly explain the human need for adequate dietary intake of vitamin C and will then examine the qualities of kiwifruit that make it an exceptional source of this vitamin.
1. VITAMIN C AND SCURVY Vitamin C became well known in the 1930s when it was identified as the single agent that could alleviate scurvy, an acute disease that had plagued humanity when fresh food was in limited supply, notably in times of war and
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famine (Sauberlich, 1997). Long sea voyages were feared because they were associated with the devastation of scurvy, which begins with depression, general malaise, and joint pain, but rapidly progresses to intestinal bleeding, difficulty with breathing, and ulceration in the gums and other parts of the body and ultimately results in death (Lind, 1753; Sauberlich, 1997). The original report written by James Lind states “It is not easy to conceive a more dismal and diversified scene of misery” (Lind, 1753). Although it is commonly thought that scurvy is a condition limited to seafarers of old, it should be noted that it was probably more easily recognized in this group because of its synchronous onset and devastating loss of life. Its occurrence in individuals in the general population is less easily diagnosed, but scurvy will occur in anyone whose intake of vitamin C from food is about 10 mg/day or less (Sauberlich, 1997). Case reports do appear in the modern literature and are a reminder that vitamin deficiencies still occur (Ho, Prinsloo, & Ombiga, 2007), thereby emphasizing the need for constant daily replenishment of the vitamin. A more common condition is likely to be hypovitaminosis C that occurs when plasma levels fall below about 25 mM for any extended period and this can be associated with poor wound healing, mood disorders, and general lassitude (Hodges, Hood, Canham, Sauberlich, & Baker, 1971; Kinsman & Hood, 1971; Schleicher, Carroll, Ford, & Lacher, 2009; Zhang, Robitaille, Eintracht, & Hoffer, 2011).
2. THE BIOLOGICAL ACTIVITY OF VITAMIN C The scurvy symptoms described above are associated with the function of ascorbate as a cofactor for a number of enzymes with critical functions throughout the body (summarized in Table 7.1). Prominent among these are the copper-containing hydroxylases present in the brain and adrenal glands that catalyze the synthesis of adrenalin and peptide hormones (Diliberto et al., 1991; Eipper & Mains, 1991) and the iron-containing proline and lysine hydroxylases that form collagen cross-links (Burri & Jacob, 1997; Levine, 1986) and generate carnitine (Nelson et al., 1981). The very high concentrations of ascorbate in the adrenals, brain, and other tissues are thought to reflect the requirement for ascorbate to maintain these vital functions. In recent years, a number of new enzymes have been identified that also require ascorbate as a cofactor (Knowles et al., 2003; Ozer & Bruick, 2007; Schofield & Ratcliffe, 2004). Most of these belong to the family of Fe-containing 2-oxoglutrate-dependent dioxygenases, and they have important roles to play in the maintenance of normal metabolism and cellular responses to environmental stresses (Table 7.1).
Table 7.1 Enzymes for which ascorbate acts as a cofactor and their biological functions Enzyme groups Activity/function Biological activities affected
References
Proline hydroxylase C-P4H-1, lysine hydroxylase
Cross-linking of collagen
Maintenance of interstitial tissue structures. Development of scar tissue, wound healing. Integrity of blood vessels and cartilage.
Burri and Jacob (1997)
g-Butyrobetaine hydroxylase, trimethyllysine hydroxylase
Synthesis of carnitine
Transport of long chain fatty acids across the mitochondrial inner membrane. Energy metabolism.
Nelson et al. (1981) and Levine (1986)
Dopamine b-hydroxylase
Generates norepinephrine from dopamine
Synthesis of adrenalin in the central nervous system and adrenal medulla. Effects on energy and vigor.
Diliberto et al. (1991)
Peptidyl-glycine alpha-amidating monooxygenase
Synthesis of neurotransmitters and neuroendocrine peptides
Essential functioning of the nervous and endocrine systems.
Eipper and Mains (1991)
HIF hydroxylases (a family of three proline hydroxylases and one asparagine hydroxylase)
Hydroxylation of the alpha chain of hypoxia-inducible factor 1
Regulation of the transcription of genes under the control of hypoxia-inducible factor. This affects cell responses to hypoxic and metabolic stress with particular modification of cell life and death pathways, angiogenesis, and energy metabolism. Highly implicated in cancer survival and development of chronic diseases.
Knowles, Raval, Harris, and Ratcliffe (2003) and Schofield and Ratcliffe (2004)
Jumonji histone demethylating dioxygenases (up to 12 identified)
Histone demethylation
Regulation of the interaction between histones and DNA, affecting chromatin organization and also gene expression. Wide-ranging consequences as yet uncharacterized.
Ozer and Bruick (2007)
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3. THE RECOMMENDED DIETARY INTAKE Since the identification of ascorbate deficiency, there has been debate as to what amount should be recommended for the daily diet (Carr & Frei, 1999; Levine, 1986; Levine, Dhariwal, Welch, Wang, & Park, 1995). The principle behind the original recommended dietary intake (RDI) recommendations has been that intake should be sufficient to prevent the development of scurvy and that excess is to be avoided. Hence, the recommended amount has been set at a level that ensures there is minimal excretion in the urine while still preventing the development of scurvy. However, a significant case has been made that this is insufficient for good health (Carr & Frei, 1999; Levine et al., 1995); and in 2000, the recommended level in the United States was increased from 60 mg/day to 90 mg for men and from 45 to 75 mg/day for women. Higher levels are recommended for smokers and pregnant and lactating women to offset additional demand. In Australasia, the RDI (Vitamin C, 2006) remains at 45 mg/day. Much of the discussion around the setting of the RDI has focused on the pharmacodynamics of ascorbate in other species. It is notable that those animals that synthesize vitamin C produce enormously high levels daily (Chatterjee, 1973). A goat of 70 kg body mass—equivalent to the average human adult—produces 13,000 mg of ascorbate daily, as well as eating a plant diet rich in vitamin C (Chatterjee, 1973). The other species that share gulonolactone oxidase deficiency consume 20- to 80-fold more vitamin C than do humans, on a body weight basis (Milton, 1999). While this is thought to indicate that the RDI is currently set too low, it has been argued that human utilization of vitamin C is much lower than for other species and that therefore a much lower daily amount should be recommended. Human studies from the United States have indicated that saturation of both plasma and tissues occurs with a daily intake of 200–400 mg/day (Carr & Frei, 1999; Levine, Wang, Padayatty, & Morrow, 2001; Levine et al., 1996). There is concern that excessive intake of vitamin C will result in toxicity. In support of this, there is some evidence that high intake is contraindicated for individuals with iron overload conditions such as hemochromatosis, or with kidney failure (Cook & Reddy, 2001; Naidu, 2003; Padayatty et al., 2010). However, with the exception of these relatively uncommon conditions, it appears that extraordinarily high doses of vitamin C are well tolerated and that any excess is efficiently cleared through the kidneys
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(Naidu, 2003; Padayatty et al., 2010). Basal plasma levels do not rise above 100 mM even when gram doses are consumed, and this has given rise to the idea that taking vitamin C supplements will not be effective in raising body stores (Mandl, Szarka, & Banhegyi, 2009; Padayatty et al., 2004). However, if plasma levels are less than saturating, supplementation with vitamin C will result in increased body status (Carr & Frei, 1999; Mandl et al., 2009).
4. VITAMIN C AND THE PREVENTION OF CHRONIC DISEASES Whereas a small amount of vitamin C daily will prevent the onset of scurvy, there is a growing body of evidence indicating that maintaining plasma levels at closer to saturation levels offers protection from some chronic diseases (Carr & Frei, 1999; Levine & Eck, 2009; Ye & Song, 2008). It is fair to say that there has been enormous confusion about the evidence to support this view and that many population-based studies are difficult to interpret (Kushi, Fee, Sellers, Zheng, & Folsom, 1996; Losonczy, Harris, & Havlik, 1996; Muntwyler, Hennekens, Manson, Buring, & Gaziano, 2002). Most of the claims come from studies in which the intake of vitamin C is monitored and compared with the disease incidence. These prospective studies have generally supported an association of higher vitamin C intake with decreased incidence of cancer, heart disease, and cataract formation (Knekt et al., 2004; Roberts, Traber, & Frei, 2009; Sesso et al., 2008). A meta-analysis of 14 cohort studies has concluded that low dietary vitamin C resulted in increased risk of coronary heart disease (Ye & Song, 2008). However, the results of intervention studies in which healthy individuals were given supplementary vitamin C over and above their normal intake, are more equivocal (Sesso et al., 2008). Only a few have shown any health benefits (Roberts et al., 2009). This is thought to be related to the fact that the study populations were already healthy, making the supplement redundant (Roberts et al., 2009). Many of the studies referred to above have shown that protection from cardiovascular disease requires an average daily intake of 400 mg vitamin C per day. This is equivalent to the amount required for fully saturated plasma and tissue levels and supports the case being made for an increased RDI (Frei, Birlouez-Aragon, & Lykkesfeldt, 2012; Levine & Eck, 2009). Similar data have been obtained for the prevention of stroke and many types of
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cancer, with studies showing a decrease in risk for these diseases generally requiring higher daily intake, often between 100 and 500 mg daily (Cho et al., 2006; Kromhout, 1987; Michels et al., 2001; Myint et al., 2008; Yokoyama et al., 2000; Zhang et al., 1999).
5. VITAMIN C AND RESPIRATORY DISEASES Possibly, the most discussed and the most investigated aspect of vitamin C has been its effect on the common cold. Although the conditions investigated have varied widely, there is a general consensus that supplemental vitamin C does not protect an already well-nourished population from contracting the cold virus, but may shorten the duration of symptoms (Douglas, Hemila, Chalker, & Treacy, 2007; Hemila, 2009). Supplementation with vitamin C has been found to decrease the susceptibility to a cold in people who are stressed by extreme exercise (marathon runners) or cold exposure (arctic explorers) (Douglas et al., 2007). A few studies have shown a more marked effect with severe respiratory illness, indicating that vitamin C may have a significant benefit in both preventing and treating pneumonia (Hemila & Louhiala, 2007). More research is needed in this area, but the link with acute respiratory illness is of interest.
6. FOOD SOURCES OF VITAMIN C All living things contain vitamin C, and therefore, it is available to some extent in most food sources (The Natural Food Hub, 2001; U.S. Department of Agriculture, 2011). Its availability is determined by both the content of the food and also the method of preparation. Ascorbate is readily oxidized and can be destroyed by exposure to oxygen, heat, and some metal ions. Thus, cooking can decrease the amount of available vitamin C and meat, which can contain substantial amounts of vitamin C, does not deliver significant vitamin C as this vitamin is destroyed in the preparation process (U.S. Department of Agriculture, 2011). In addition, organ meats such as liver and brain, which contain more vitamin C than muscle, are not often eaten in the Western diet. Fresh food ensures the best delivery and, as fruit is generally consumed fresh, this is by far the most effective way to achieve an adequate daily intake (The Natural Food Hub, 2001). It should be noted
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that many foods deliver some vitamin C, and the key to effective intake is to ensure a good variety of fruit and vegetables in the daily diet. The vitamin C content of some common foods is shown in Table 7.2. From this list, it can be seen that kiwifruit is rated as one of very few foods that are considered to be an exceptional source of vitamin C, with a single Table 7.2 The vitamin C content of some common foods, based on content and serving size Rating as a source Vitamin C content Vitamin C per of vitamin C Food (mg/100 g) servinga (mg)
Guava
183
165
Exceptional
Kiwifruit, gold
105–120
95–108
Exceptional
Capsicum
180
95
Exceptional
Acerola
1677
80
Exceptional
Kiwifruit, green
65–90
50–74
Exceptional
Orange
53
70
Excellent
Mango
28
57
Excellent
Broccoli
90
51
Excellent
Rosehip
1500
45
Excellent
Grapefruit
34
44
Excellent
Brussels sprouts
80
40
Excellent
Persimmon
40
40
Excellent
Watermelon
10
27
Very good
Mandarin
31
26
Very good
Tamarillo
31
22
Very good
Tomato
19
23
Very good
Cabbage greens
30
20
Very good
Feijoa
31
16
Very good
Potato
20
15
Very good
Pineapple
15
13
Good
Banana
9
11
Good
Strawberry
57
7
Good
Apple
6
8
Fairly good
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Table 7.2 The vitamin C content of some common foods, based on content and serving size—cont'd Vitamin C content Vitamin C per Rating as a source Food (mg/100 g) serving (mg) of vitamin C
Grape
11
6
Fairly good
Carrot
9
5
Good
Lettuce
4
4
–
Cucumber
3
1
–
Avocado
8
2
–
Onion
7
1
–
Cow’s milk (fresh)
2
–
–
Nuts
–
–
–
Grains
–
–
–
Seeds (including peas and beans)
–
–
–
Data in this table are taken from The Natural Food Hub, 2001 and U.S. Department of Agriculture, 2011 a One serving is defined as the amount normally eaten and varies for different food types, for example, one slice of watermelon or avocado, two mandarins, ½ cup cooked vegetables, ½ cup grapes.
serving providing almost the entire RDI as determined in the United States, and twice the Australasian RDI (Vitamin C, 2006). The exceptional label is a composite of the amount of vitamin C present in the food per unit weight and the likely serving size (The Natural Food Hub, 2001; U.S. Department of Agriculture, 2011). Therefore, some foods that have very high vitamin C content, such as blackcurrants, strawberries, or lemons, are not considered a highly effective dietary source because less is consumed at any one time. It should also be noted that most fruits generally have higher vitamin C content than vegetables. Legumes such as peas and beans, nuts, grains, and seeds do not contain measureable amounts of vitamin C. This, together with the destruction of vitamin C in meat upon cooking, means that delivery of vitamin C to the diet is mostly, if not entirely, from fruit and vegetables, with fruit being the major contributor.
7. VITAMIN C CONTENT OF KIWIFRUIT There are many varieties of kiwifruit and their vitamin C content differs markedly. An analysis of more than 20 varieties reported a range of 25–205 mg/ 100 g (Hunter, Greenwood, Zhang, & Skinner, 2011; Nishiyama et al., 2004).
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These levels reflect the total AA content of the fruit, which is a combination of both AA and its oxidized form, dehydroascorbic acid (DHAA) (Nishiyama et al., 2004). Because it is known to be readily reduced in the body, DHAA is generally considered as part of the available vitamin C. However, animal studies have indicated that, when given orally, DHAA has limited antiscorbutic properties, and may therefore not be biologically available (Ogiri et al., 2002). For this reason, it is probably more accurate to consider only the AA content when estimating the effective vitamin C content. There is a large degree of variability in the proportion of DHAA present in the different kiwifruit cultivars (Nishiyama et al., 2004). The reasons for this variation are unknown, but may reflect levels of plant stress or inherent differences in the amount of the enzyme ascorbate oxidase, which catalyzes the oxidation of AA to DHAA. In the most common commercial cultivar, Actinidia deliciosa cv. ‘Hayward’ (green kiwifruit), approximately 15% of the total AA is present as DHAA, and a higher level (up to 30%) is found in the Actinidia chinensis cv. ‘Hort 16A’ (gold kiwifruit) (Nishiyama et al., 2004). This means that the effective concentration of AA for these two commercial cultivars is likely to be around 55–65 mg/100 g for the green and 75–85 mg/100 g for the gold. In agreement with this, our laboratory has measured 89 mg AA/ 100 g of gold kiwifruit, which is equivalent to about 80 mg per fruit. The vitamin C content of fruit is often found to decrease upon storage. This was shown not to be a significant factor in the main kiwifruit varieties, with levels remaining stable during postharvest ripening (Nishiyama et al., 2004). However, we have measured a decline in AA levels of around 30% after extended long-term chilled storage of around 9 months postharvest (M. Vissers and L. Braithwaite, unpublished data).
8. EFFECT OF KIWIFRUIT SUPPLEMENTATION ON VITAMIN C INTAKE The information above suggests that the addition of kiwifruit to the diet would contribute significantly to overall vitamin C intake. We have recently been able to demonstrate just how effective this can be with a human supplementation study. The study population comprised 14 nonsmoking healthy male university students aged between 18 and 30 years who had an average baseline fasting plasma ascorbate level of 38 mM. The participants kept detailed food and beverage diaries, which indicated a mean baseline intake of 3.9 0.5 servings of fruit and vegetables per day (Fig. 7.1A). Based on the specifics of the food consumed, this was estimated
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Figure 7.1 The effect of kiwifruit consumption on dietary vitamin C intake in humans. Data from a human intervention study involving 15 males aged 18–35 are shown. (A) Daily servings of fruit and vegetables consumed by the participants were estimated from weekly food diaries. Addition of kiwifruit to the diet increased the baseline consumption from four to six serves with the addition of two or three kiwifruit (KF) daily. (B) Vitamin C intake was significantly increased with the addition of even one half a KF per day. After washout, levels returned to baseline. Data represent means SEM from a group of 15 participants. indicates statistical significance (P < 0.05) from baseline as determined by one-way repeated measures analysis of variance with Fisher LSD pairwise multiple comparison procedure.
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to contribute 44 5 mg ascorbate per day, which is the equivalent of the Australasian RDI and indicates an average of around 11 mg vitamin C per serving of fruit or vegetables. When kiwifruit was given daily, it appeared that some individuals consumed this in addition to their daily diet, whereas others reduced their intake of other fruit and vegetables to accommodate their increased kiwifruit consumption. Consequently, addition of one half or one kiwifruit per day did not alter the participants’ average number of fruit and vegetable servings. Addition of two and three kiwifruit to the daily diet resulted in only a small increase, to a maximum of six serves per day at the three kiwifruit dose. Despite the lack of an increase in fruit and vegetable servings at the lowest doses, supplementation with one half and one kiwifruit per day significantly increased the average daily intake of vitamin C to around 80 and 130 mg, respectively (Fig. 7.1B). The latter is equivalent to the amount recommended for prevention of chronic disease (Carr & Frei, 1999; Padayatty & Levine, 2008) and reflects the high ascorbate content in the fruit. Total intake reached 220 mg/day when two kiwifruit per day were consumed (Fig. 7.1B). This is comparable to the intake required to achieve plasma saturation and is now considered by some to be optimal (Frei et al., 2012; Levine & Eck, 2009; Padayatty & Levine, 2001). These results clearly demonstrate the value and effectiveness of adding a high vitamin C-containing food like kiwifruit to the diet.
9. EFFECT OF KIWIFRUIT INTAKE ON PLASMA VITAMIN C In recent years, several studies have monitored plasma vitamin C in individuals supplemented with kiwifruit. In most of these studies, the objective was to monitor specific biological effects of nutrition and in these studies, kiwifruit was found to improve digestive health (Rush, Patel, Plank, & Ferguson, 2002), enhance iron uptake (Beck, Conlon, Kruger, Coad, & Stonehouse, 2011), increase DNA repair activity (Brevik et al., 2011, see also Chapter 16), modulate lipid profiles (Chang & Liu, 2009), and reduce platelet aggregation (Duttaroy & Jorgensen, 2004). Most of these studies showed that consuming kiwifruit resulted in an increase in plasma vitamin C, with saturation levels being reached at a dose of two or three kiwifruit per day. Collins et al. reported a dose-dependent effect upon supplementation with one to three kiwifruit per day (Brevik et al., 2011; Collins, Harrington, Drew, & Melvin, 2003). The increase was rather modest, however, with plasma steady
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137
state levels increasing by 7–16 mM (Brevik et al., 2011). These small increases are likely to reflect the fact that the participants already had saturating plasma vitamin C levels of about 60 mM at baseline (Brevik et al., 2011). To more accurately measure the contribution of kiwifruit to dietary vitamin C intake, we recently monitored plasma vitamin C levels in a group of 14 male students with an average baseline plasma vitamin C level of 38 mM (Carr, Pullar, Moran & Vissers, 2012). We were able to measure a strong effect of kiwifruit supplementation on plasma vitamin C levels, with increases of around 25 mM (Fig. 7.2A). When a subpopulation with baseline plasma vitamin C levels of 28 mM was investigated, we measured an increase of 22–43 mM with supplementation of one half to three kiwifruit per day. The effect was dose-dependent and was seen after the first week of supplementation. At two kiwifruit per day, plasma levels approached saturation with no further increase with three per day, although some noncompliance was evidenced at this highest dose. Plasma saturation was confirmed by monitoring urinary excretion, which showed that there was no increase until a dose of two kiwifruit per day (Fig. 7.2B). This coincided with plasma levels reaching around 60 mM. During the washout period of the study, plasma and urine levels rapidly returned to baseline (Fig. 7.2A and B), which strongly suggests significant daily turnover of body stores.
10. EFFECT OF KIWIFRUIT INTAKE ON TISSUE VITAMIN C LEVELS Tissues throughout the body concentrate ascorbate from the plasma via active transport. The uptake is generally thought to reflect plasma concentrations and some tissues saturate at lower vitamin C intake than is required for plasma saturation (Levine et al., 1996; Levine et al., 2001). Because of difficulty in accessing human tissue, most information about tissues has been obtained from the measurement of white blood cell levels. Leukocytes preferentially take up ascorbate to millimolar intracellular concentrations with saturation levels being reached with a daily intake of around 100 mg/day (Carr & Frei, 1999; Levine et al., 1996; Levine et al., 2001). This corresponds to plasma levels of around 50 mM (Lee, Hamernyik, Hutchinson, Raisys, & Labbe, 1982; Levine et al., 1996; Levine et al., 2001). In agreement with this, we found that the supplementation of one kiwifruit per day to our study
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Figure 7.2 Increase in plasma vitamin C as a result of kiwifruit consumption. (A) Plasma vitamin C levels of a group of 14 young men (aged 18–30 years) with an average baseline plasma vitamin C level of 38 mM were measured after increased intake of kiwifruit in addition to baseline diet. Study participants consumed kiwifruit as indicated for a period of 4–6 weeks and fasting plasma vitamin C levels were monitored weekly. (B) Urinary excretion of vitamin C by the same study participants as a function of daily kiwifruit intake. Data represent mean SEM. indicates statistical significance (P < 0.05) from baseline as determined by one-way repeated measures analysis of variance with Fisher LSD pairwise multiple comparison procedure.
Vitamin C from Kiwifruit
139
population resulted in optimal leukocyte vitamin C, with no further increase seen with two kiwifruit per day. Addition of one kiwifruit corresponded to a total dietary vitamin C intake of around 130 mg/day. That we were able to detect a statistically significant increase in leukocyte vitamin C from baseline levels in our study population is likely due to their low initial levels.
11. ANIMAL STUDIES WITH KIWIFRUIT Monitoring vitamin C levels in tissues other than white blood cells is difficult in humans, and animal studies have provided most of this information. A recently developed transgenic mouse model that lacks gulonolactone oxidase and displays the same phenotype as the human deficiency condition has provided a useful tool with which to assess the pharmacodynamics of vitamin C uptake and metabolism (Maeda et al., 2000; Parsons, Maeda, Yamauchi, Banes, & Koller, 2006). We used this model, the Gulo / mouse, to monitor the effect of variable dietary ascorbate on uptake into the tissues and also to investigate the bioavailability of ascorbate from kiwifruit (Vissers, Bozonet, Pearson, & Braithwaite, 2011). We found that the ascorbate levels differed in the various mouse tissue compartments (Vissers et al., 2011) and were in agreement with the expected relative distributions in the same tissues from studies of other vitamin C-deficient animals, including humans (Tsao, 1997). We monitored plasma, brain, heart muscle, liver, kidney, and white cells in response to varied dietary intake administered through the drinking water. Uptake for all tissues followed a sigmoid-shaped dose–response curve, similar to that observed in human studies. The rate of uptake differed significantly, and in particular, liver, heart, and kidney required around five times more ascorbate than brain tissue to reach saturation levels. We also determined the effectiveness of vitamin C uptake from kiwifruit in the Gulo / mouse and compared this with delivery of reagent vitamin C delivered through the drinking water or embedded in a gel. When mice were fed a fresh kiwifruit gel preparation (either green or gold varieties) of known ascorbate content for 4 weeks, the ascorbate levels measured in blood, liver, kidney, and heart were markedly higher than those measured in animals taking an equivalent dose of purified ascorbate via the water supply or in a gel form (Vissers et al., 2011). A period of 4 weeks was required to reach tissue steady state, which is similar to the time needed for humans. The
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increased bioavailability of vitamin C from kiwifruit was clearly apparent when we compared the dose response. When kiwifruit was used, a significantly (p < 0.05) lower daily ascorbate intake was required to reach 50% of the maximum levels (ED50) in serum, heart, kidney, and liver. This difference was around fivefold for some tissues (e.g., liver). Both varieties of kiwifruit delivered significantly more ascorbate to most tissues when compared directly with an equivalent dose of ascorbate supplied in the drinking water (Table 7.3). These results were somewhat surprising and the suggestion that kiwifruit could be a much better source of dietary vitamin C than a purified supplement is of interest for human nutrition. Our present studies aim to address this question in humans, and the results with the university student population discussed above suggest that the addition of kiwifruit to the diet is certainly a highly effective way to increase plasma and white cell levels. Whether kiwifruit would be more effective in humans than a purified ascorbate supplement is yet to be determined. Table 7.3 Direct comparison of vitamin C levels after supplementation with either reagent ascorbate or green or gold kiwifruit in the Gulo / mouse Gulo / mice Daily vitamin C intake and source Supplement ascorbate 1.7 mg (n ¼ 5)
Green Supplement kiwifruit ascorbate 1.8 mg (n¼4) 5.0 mg (n ¼ 5)
Gold kiwifruit 3.0 mg (n ¼ 4)
Serum
15.0 4.9
34.1 7.9*
24.6 7.8
64.9 8.9*
Brain
51.5 6.2
37.5 1.1*
55.8 1.4
64.7 3.2*
White blood cells
2.3 0.6
3.5 0.7
5.8 0.7
5.0 0.4
Liver
2.5 0.8
9.6 1.7** 10.7 0.7
Heart muscle
1.2 0.4
1.4 0.5
1.6 0.4
1.4 0.5
Kidney
2.4 0.9
4.6 1.1*
3.3 0.2
9.4 2.0*
20.6 2.8**
Vitamin C was supplied in kiwifruit or as a purified compound (sodium ascorbate) through the water supply (supplement) for a period of 4 weeks (Vissers et al., 2011). Measured concentrations as given are mM for serum; nmol/108 cells for white blood cells; and mg/100 mg wet weight for tissues from brain, liver, kidney, and heart. Means SE are shown and significant differences between kiwifruit and ascorbate supplement, as determined using the student’s t-test for unpaired samples, are indicated (*p < 0.05 or **p < 0.01).
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12. EFFECT OF OTHER PLANT COMPONENTS ON UPTAKE OF VITAMIN C In the mouse study described above, we also found that eating kiwifruit could elevate serum and tissue vitamin C levels beyond those seen in a wild-type control mouse that maintains high and stable levels by synthesis in the liver. This suggests that there may be a difference in the efficiency of intestinal absorption and/or clearance through the kidneys in the kiwifruitfed animal. This observation is reminiscent of a speculation often made in the mid twentieth century when it was proposed that some foods contain an unknown factor that enhances the bioavailability of vitamin C. Reference was made to a hypothetical factor described as an “AA-economizing factor,” “vitamin P,” or “vitamin C2” which affected the efficiency of ascorbate uptake (Cotereau, Gabe, et al., 1948). Alternatively, there may be other factors present that could affect the delivery of vitamin C. Kiwifruit are a rich source of vitamin E (up to 10% of the RDI per serving) (Celik, Ercisli, & Turgut, 2007), as well as a range of flavonoids (mostly in the skin) and have a high content of dietary fiber (Hunter et al., 2011; Nishiyama, 2007). Some early studies with people or guinea pigs suggested that ascorbate uptake is enhanced by catechin and other flavonoids (Cotereau et al., 1948), but very little good evidence can be found that this actually occurs (Nishiyama, 2007). In contrast, vitamin E deficiency has been shown to accelerate vitamin C loss in the ODS rat (Tanaka, Hashimoto, Tokumaru, Iguchi, & Kojo, 1997) and, given the high content of vitamin E in kiwifruit (Nishiyama, 2007), it is possible that an additional source of vitamin E may affect vitamin C pharmacokinetics. Thus, it will be of interest to further investigate the effects of vitamin C uptake from kiwifruit in humans. (Note from editors: The ODS, or Osteogenic Disorder Shionogi rat is a wellestablished model animal for studying vitamin C metabolism.) The effect of bioflavonoids on the uptake of ascorbate has been directly investigated in a few studies. In one study, a mixture of synthetic ascorbate and a citrus extract containing a mixture of bioflavonoids, proteins, and carbohydrates was administered to a group of human volunteers (Vinson & Bose, 1988). The ascorbate from the food mix was found to be more slowly absorbed and was effectively 35% more bioavailable, determined by measuring plasma levels and urinary excretion over a 24-h period. Another study found no difference between a synthetic ascorbate
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dose of 500 mg with or without added bioflavonoids (Johnston & Luo, 1994). The main difference between these two apparently contradictory studies was the ratio of bioflavonoids to ascorbate (4:1 in the first study and 0.05:1 in the second). This highlights the importance of determining the optimum study conditions before embarking on human intervention trials, and the results suggest that further investigations may be warranted.
13. NATURAL VERSUS SYNTHETIC VITAMIN C The suggestion above that there may be differences in the uptake and metabolism of vitamin C from foods has been a topic of frequent discussion and some investigations have provided useful information. Chemically speaking, synthetic AA is indistinguishable from the same chemical that is present in plants and animals. There have been a small number of human studies that have directly compared the uptake of synthetic ascorbate and an equivalent amount in citrus fruit and these have found that both contributed equally to plasma vitamin C levels (Gregory, 1993; Mangels et al., 1993; Pelletier & Keith, 1974). Different formulations of synthetic vitamin C are often found in supplements, with the claim being made that some will have better bioavailability (Nyyssonen et al., 1997; Yung, Mayersohn, & Robinson, 1982). These include AA preparations presented as powders, in capsules or tablet form, or as slow-release formulations. It remains unclear whether any one form is more absorbable than another. The biggest difference between these forms may well be the stability of the product during storage, as ascorbate is readily oxidized by exposure to air, heat, and light. For this reason, it is better kept as a capsule or tablet than as a powder when stored over longer periods. When present in a liquid, ascorbate is often more labile and can be destroyed by heating in the commercial preparation of fruit juices. To ensure consistency of these products, synthetic ascorbate is often added back prior to sale. Because vitamin C supplements are often taken in higher bolus amounts than would be found in food, formulations are often made to mitigate the effect of the free acid and to attempt to ensure slower and more prolonged uptake. Mineral salts of AA, most commonly sodium or calcium ascorbate, are neutral and this is thought to lessen bowel irritation, but there is little actual evidence that this does occur. “Ester C” is a mixture of mostly calcium ascorbate, together with some metabolites (DHAA and calcium threonate) and differs from esterified ascorbate, which include mixtures of vitamin C and lipids that are claimed to increase bioavailability. However, in vivo data that would
Vitamin C from Kiwifruit
143
determine the relative effectiveness of these products compared with AA either in purified form or from a food source is currently lacking.
14. CONCLUSION Despite the many and varied formulations of vitamin C supplements, it should be noted that all clinical information to date suggests that the ascorbate present in fresh food is bioavailable and readily absorbed and is the preferred source, as it will be delivered together with other important nutrients and minerals. Hence, the best advice that can be given is to encourage the consumption of a wide range of fresh foods and to include in the mix one to two serves per day of a high vitamin C-containing food such as kiwifruit.
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