Vitamin C: Sources, Functions, Sensing and Analysis

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Chapter 1
Vitamin C: Sources, Functions, Sensing and Analysis
Sudha J. Devaki and Reshma Lali Raveendran
Additional information is available at the end of the chapter
http://dx.doi.org/10.5772/intechopen.70162
Abstract
Vitamin C is a water-soluble compound found in living organisms. It is an essential nutri-
ent for various metabolism in our body and also serves as a reagent for the prepara-
tion of many materials in the pharmaceutical and food industry. In this perspective, this
chapter can develop interest and curiosity among all practicing scientists and technolo-
gists by expounding the details of its sources, chemistry, multifunctional properties and
applications.
Keywords: vitamin C, biomarker, antioxidant, sensors
1. Introduction
Vitamin C also known as ascorbic acid (AA) is an essential nutrient in many multicellular organ-
isms, especially in humans. Ascorbic acid is a water-soluble vitamin and is found in variable
quantities in fruits and vegetables and organ meats (e.g. liver and kidney). Deciency of vitamin
C causes scurvy, widespread connective tissue weakness and capillary fragility. Among chem-
ists, it is used as a reagent for the preparation of ne chemicals, enzymatic reagent and nanoma-
terials. Consequently, the detection and quantication of ascorbic acid in food samples, products
and nutraceuticals is receiving overwhelming importance among researchers, medical practitio-
ners and also in the pharmaceutical and food industry. Figure 1 shows the schematic representa-
tion of the sources and multifunctional applications of vitamin C in the metabolism of our body.
2. Sources
Vitamin C (Figure 2) is abundantly available in many natural sources, including fresh fruits
and vegetables. The richest sources of ascorbic acid including Indian gooseberry, citrus fruits
© 2017 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons
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such as limes, oranges and lemons, tomatoes, potatoes, papaya, green and red peppers, kiwi-
fruit, strawberries and cantaloupes, green leafy vegetables such as broccoli, fortied cereals
and its juices are also rich sources of vitamin C.
Another source of vitamin C is animals. They usually synthesize their own vitamin C and are
highly concentrated in the liver part [1, 2]. Sources of vitamin C and its content are given in
Figure 3. Average daily recommended amounts of vitamin C for dierent ages are also given
in Table 1.
Figure 1. Sources and multifunctional role of vitamin C in metabolic processes in human body.
Figure 2. Structure of vitamin C.
Vitamin C4

Figure 3. Amount of vitamin C in dierent sources.
Infants
Age Adequate intake (AI) mg/day Upper intake level
(UL)
0–6 monthsa25 –
7–12 monthsb30 –
Children and adolescents
Age EARcRDId
mg/day mg/day mg/day
1–3 25 35 400
4–8 25 35 650
9–14 28 40 1200
14–18 28 40 1800
Adults
Men Women Both
Age EARcRDIdEARcRDId
mg/day mg/day mg/day mg/day mg/day
19–30 30 45 30 45 2000
31–50 30 45 30 45 2000
51–70 30 45 30 45 2000
>70 30 45 30 45 2000
Vitamin C: Sources, Functions, Sensing and Analysis
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3. Functions
Vitamin C plays an important role in many physiological processes in humans. It is needed
for the repair of tissues in all parts of the body. The important functions of vitamin C include
the formation of protein used to make skin, tendons, ligaments, and blood vessels for healing
wounds and forming scar tissue, for repairing and maintaining cartilage, bones, and teeth
and aid in the absorption of iron. It can also act as a reducing and capping agent for metal
nanoparticles.
3.1. As reducing and capping agent
Ascorbic acid acts as a reducing and capping agent for the synthesis of metal nanoparticles
such as silver, gold, copper, etc. Ascorbic acid molecules can cap or surround the particle
and prevent the uncontrolled growth of the particles to micron-sized dimensions. A study
by Khan et al., in 2016 reported the synthesis of copper nanoparticle by using ascorbic acid
as both the reducing agent [6]. Sun et al. reported in Journal of Materials Science in 2009 that
gold nanoparticles can be synthesised in reverse micelles without the addition or introduc-
tion of any other reducing or capping reagent [7]. In analytical methods in 2014, D’souza
et al. reported the use of AA-Au NPs as a colorimetric probe for detection of dichlorvos in
water and wheat samples. The inuence of AA concentration on the aggregation induced by
dichlorvos in AA-AuNPs (Figure 4) and the optical property of the AA-Au NPs was investi-
gated by UV-Vis spectroscopy [8].
Infants
Age Adequate intake (AI) mg/day Upper intake level
(UL)
Motherhood
Pregnancy Lactation Both
Age EARcRDIdEARcRDId
mg/day mg/day mg/day mg/day mg/day
14–18 38 55 58 80 2000
19–30 40 60 60 85 2000
31–50 40 60 60 85 2000
aCalculated as per the average intake of breast milk.
bCalculated on a body weight basis of infants.
cEstimated average requirement.
dRecommended dietary intake.
Table 1. Dietary intake of vitamin C [3–5].
Vitamin C6

3.2. Antioxidant activity
One of the important properties of vitamin C is its antioxidant activity. Antioxidant activity of
vitamin C helps to prevent certain diseases such as cancer, cardiovascular diseases, common
cold, age-related muscular degeneration and cataract.
3.3. In cancer treatment
Since 1970, it has been known that high dose of vitamin C has benecial eects on the sur-
vival time in patients with terminal cancer, which was reported by Cameron, Campbell, and
Pauling. Research is undergoing in detail for using vitamin C in cancer treatment [9–11]. One
of the studies suggests that pharmacologic doses of vitamin C might show promising eects
on the treatment of tumours [12]. Vitamin C can act as pro-oxidant and it can generate hydro-
gen peroxide [13, 14]. Administration of high dose of vitamin C gives long survival times for
patients with advanced cancers.
The continuing aack of DNA by unquenched reactive oxygen species is believed to cause
cancer. As a physiological antioxidant, ascorbic acid plays a role in the prevention of oxida-
tive damage to DNA, which is elevated in cells at sites of chronic inammation and in many
pre-neoplastic lesions. Most DNA damage is repaired metabolically; however, the frequency
of elevated steady-state levels of oxidized DNA bases is estimated to be sucient to cause
mutational events. The base damage product 8-hydroxy-2′-deoxyguanosine has been found
to be elevated in individuals with severe vitamin C deciency and to be reduced by supple-
mentation with vitamins C and E [15].
Figure 4. Analytical process for detecting dichlorvos using AA-Au NPs as a colorimetric probe [8].
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Oxidative stress, arising as a result of an imbalance between free radical production and anti-
oxidant defences, causes damage to a wide range of molecular species including lipids, pro-
teins and nucleic acids. Oxidative stress caused by free radicals is shown in Figure 5. The best
way to ensure adequate intake of the antioxidant nutrients, such as vitamins C and E and
various kinds of minerals, is through a balanced diet consisting of 5–8 servings of fruits and
vegetables per day.
3.4. In cardiovascular diseases
The antioxidant property of vitamin C helps for the treatment of cardiovascular diseases.
Vitamin C has the capability for reducing monocyte adherence to the endothelium, improv-
ing endothelium-dependent nitric oxide production and vasodilation and reducing vascular
smooth-muscle-cell apoptosis, which prevents plaque instability in atherosclerosis [15]. The
oxidative damage including the oxidative modication of low-density lipoproteins is a major
cause of cardiovascular disease. The antioxidant property of vitamin C helps to reduce this to
a certain extent [16, 17].
3.5. In common cold
Pauling in 1970 suggested that vitamin C can be used for the treatment of common cold [18].
There are so many reports in Cochrane Database Syst. Review showing the use of prophylactic
Figure 5. Oxidative stress by free radicals.
Vitamin C8

vitamin C reduces the cold duration in adults and children [19]. The use of vitamin C might
reduce the duration of common cold due to its anti-histamine eect of high dose of vitamin C
[20]. However, the results are inconsistent and still research is undergoing in this eld. There
are Database Syst. Review showing the use of prophylactic vitamin C reduces the cold dura-
tion in adults and children [19]. The use of vitamin C might reduce the duration of common
cold due to its anti-histamine eect of high dose of vitamin C [20].
3.6. In age-related macular degeneration (AMD) and cataract
Age-related macular degeneration (AMD) and cataracts are two of the main causes of vision
loss in older. Oxidative stress might contribute to the aetiology of both conditions. Thus,
researchers have taken interest in the role of vitamin C and other antioxidants in the develop-
ment and treatment of these diseases. There are many reports to study the role of vitamin C
in AMD and cataract [21–23]. Results from two studies indicate that vitamin C intakes greater
than 300 mg/day reduce the risk of cataract formation by 75% [16, 24, 25].
All the studies indicate that the vitamin C formulations might slow AMD progression and
reduce the high risk of developing advanced AMD. AMD is shown in Figure 6.
3.7. Antioxidant mechanism
Vitamins C can protect the body against the destructive eects of free radicals. Antioxidants
neutralize free radicals by donating one of their own electrons, ending the electron-stealing
reaction, as shown in Figure 7. The antioxidant nutrients themselves do not become free radi-
cals by donating an electron because they are stable in either form or act as scavengers, help-
ing to prevent cell and tissue damage that could lead to cellular damage and disease.
Ascorbic acid reacts with free radicals undergoing single-electron oxidation to produce a rela-
tively poor reactive intermediate, the ascorbyl radical, which disproportionates to ascorbate
and dehydroascorbate. Thus, ascorbic acid can reduce toxic, reactive oxygen species superox-
ide anion (O2
•) and hydroxyl radical (OH•), as well as organic (RO2) and nitrogen (NO2
•) oxy
Figure 6. Age-related macular degeneration.
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radicals. Those reactions are likely to be of fundamental importance in all aerobic cells. This
reaction is the basis of most of the biological functions of ascorbic acid. The mechanism of free
radical action on DNA is shown in Figure 8.
Vitamin C also has role in protecting other vitamins (vitamin A and vitamin E) from the harm-
ful eects of oxidation. Vitamin C helps in protecting gums and retards ageing. It strengthens
the general physical condition by removing toxic metals from the body. Vitamin C reduces
the formation of cataract and hence useful in the treatment of glaucoma.
Figure 7. Antioxidant mechanism.
Figure 8. Mechanism of free radical action on DNA.
Vitamin C10

3.8. Synthesis of protein
Another important function of vitamin C is its role in the synthesis of protein. Vitamin C helps
in the synthesis of collagen. Collagen protects our skin from wrinkling and makes our skin
rm and strong. Collagen also protects and supports organs and other soft tissues. One of the
amino acids used to build collagen—hydroxyproline—is only synthesized when vitamin C is
available. Functions of vitamin C on skin.
3.9. Functions of vitamin C on skin
3.9.1. Photoprotection
Vitamin C reduces the damage caused by UV-light exposure. It cannot act as a sunscreen
since it cannot absorb UV light. But the antioxidant activity of vitamin C helps to protect UV
damage caused by free radicals [26]. In response to UV light, vitamin C transport proteins are
increased, suggesting an increased need for vitamin C uptake for adequate protection [27, 28].
Addition of keratinocytes on vitamin C reduces damages caused by UV light and lipid peroxi-
dation, limits the release of pro-inammatory cytokines and protects against apoptosis [29].
Many studies have suggested that vitamin C consumption alone will not reduce the eect of
UV exposure; however, a combination of vitamin C and E eectively increases minimal ery-
themal dose (MED) (the lowest dose of ultraviolet radiation (UVR) that will produce a detect-
able erythema 24 hours after UVR exposure) and decreases erythema-induced blood ow
to damaged areas of skin [30]. Thus, interactions between the two antioxidant vitamins may
be necessary to achieve UV protection. The topical application of vitamin C also reduces the
eect of UV exposure and skin wrinkling and skin tumour [31]. Vitamin C reduced the num-
ber of sunburned cells, decreased erythema response and reduced DNA damage induced by
UV exposure [27]. The combination of antioxidant vitamins decreased the immunosuppres-
sive eects of UV exposure, increased MED and decreased cell damage [32, 33].
3.9.2. Wound healing
Vitamin C plays a key role in healing wound by the formation of collagen, connective tissue
[34–36]. The new tissue is rebuilt with the help of collagen framework. This function is sup-
ported by its co-factor vitamin C. Besides this, vitamin C performs as a strong antioxidant and
immune system modulator [37, 38].
3.10. Deciency of vitamin C
Deciency of vitamin C in humans causes a major disease called scurvy. The major signs
of the disease occur primarily in mesenchymal tissues. It leads to impaired wound healing;
oedema; haemorrhage (due to decient formation of intercellular substance) in the skin,
mucous membranes, internal organs, and muscles; and weakening of collagenous struc-
tures in bone, cartilage, teeth and connective tissues. Those who suer from scurvy have
swollen, bleeding gums with tooth loss. They also show lethargy, fatigue, rheumatic pains
in the legs, muscular atrophy and skin lesions, massive sheet hematomas in the thighs,
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and ecchymoses and haemorrhages in many organs, including the intestines, sub-periosteal
tissues and eyes. All these features are accompanied by psychological changes: hysteria,
hypochondria and depression. In children, the syndrome is called Moeller-Barlow disease;
it is seen in non-breastfed infants usually at about 6 months of age and is characterized
by widening of bone-cartilage boundaries, stressed epiphyseal cartilage of the extremities,
severe joint pain and frequently, anaemia and fever. Children having this disease present
with a limp or inability to walk, tenderness of the lower limbs, bleeding of the gums and
petechial haemorrhages.
4. Detection and sensing
Many analytical techniques are used for the determination of vitamin C in dierent matrices,
such as titrimetric [39], uorimetric [40], spectrophotometric [41], high-performance liquid
chromatography [42], enzymatic [43], kinetic [44, 45] and electrochemical, etc.
4.1. High-performance liquid chromatography (HPLC)
High-performance liquid chromatography (HPLC) methods are preferred earlier because
they are faster and more eective than spectrophotometric, titration or enzymatic methods,
and they do not usually need derivatization [46]. In pharmaceutical and cosmetic industries,
HPLC is used, which is considered as a sensitive and selective method.
Detection of vitamin C through HPLC has been done by many groups. Racz et al. in 1990
reported that HPLC used the HPLC method for the determination of ascorbic acid in fruits
and vegetables. The method has been used to deep-frozen raspberry cream analysis together
with three commonly used chemical methods of vitamin C analysis. The HPLC method has
been compared with the chemical methods from several aspects, and the superiority of HPLC
method has been concluded [47].
Snezana et al. reported the HPLC method for the determination of vitamin C in pharmaceu-
tical samples in Tropical Journal of Pharmaceutical Research in 2011 [48]. The simplicity of this
low-cost, rapid technique and its high specicity to ascorbic acid, even in the presence of a
variety of excipients, demonstrate that this HPLC method would be particularly suitable for
the determination of ascorbic acid (Figure 9). It can be used in pharmaceutical/veterinary
formulations without prior sample preparation.
Another work carried out by Franco et al., which was to optimize and validate a new analytic
strategy for the determination of vitamin C in strawberries by UV–HPLC [49].
For the accurate and reliable measurement of ascorbic acid and dehydroascorbic acid, HPLC
can be used in combination with electrochemical or ultraviolet detection. Line and Hoer
have reported a method for the determination of vitamin C in plasma. It can be analysed
by HPLC in connection with electrochemical or ultraviolet light detection. Electrochemical
HPLC has advantages over UV-HPLC for plasma total vitamin C analysis [50].
Vitamin C12

4.2. Spectrophotometric method
Among many analytical methods, spectrophotometric methods are very simple and low-cost.
Several studies used the spectrophotometric method for the determination of ascorbic acid.
Güçlü et al. [51] have proposed a spectrophotometric method based on ascorbic acid oxida-
tion to dehydroascorbic acid, by using the Cu(II)-neocuproine complex, which is reduced to
Cu(I)-bis(neocuproine), the absorbance of the laer being determined at 450 nm. Other opti-
cal methods for vitamin C estimation include spectrophotometrical determination of iodine
reacted with ascorbic acid [52] and chemiluminescence [53].
A sensitive, simple and low-cost spectrophotometric method was introduced by Kobra and
Somayye in 2015. The present method was successfully applied to determine the ascorbic acid
in food and pharmaceutical samples. The samples were multivitamin tablet, eervescent tab-
let, vitamin C injection, natural orange juice, orange syrup powdered and commercial orange
liquid. The method is based on the reaction of AgNO3 with ascorbic acid in the presence of
polyvinyl pyrrolidone (PVP) and slightly basic medium to prepare silver nanoparticles [54].
Kapur et al. in 2015 reported a method that is based on the oxidation of ascorbic acid to
dehydroascorbic acid by bromine water in the presence of acetic acid. In this method, the
total ascorbic acid (ascorbic acid + dehydroascorbic acid) has been determined in 21 dierent
samples of fruits and vegetables by the spectrophotometric method [55]. Mohammed and
Hazim in 2016 reported a UV-spectrophotometric method for the determination of ascorbic
acid in fruits and vegetables from hill region with 2,4-dinitrophenylhydrazine [56].
A new sensitive colorimetric method for the determination of ascorbic acid tablet in aqueous
solution was reported by Ahmed and Mohamed in 2013. The method is based on the forma-
tion of coloured azo dye by diazotization of 2,4-dichloroaniline, followed by azo-coupling
reaction between the resulting product and ascorbic acid [57].
Figure 9. HPLC plot of ascorbic acid detection.
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Figure 10. Cyclic voltammogram of electrochemical sensing of ascorbic acid using a polyaniline-modied platinum
electrode [70].
4.3. Electrochemical sensing
Electrochemical sensing methods are more widely applied as they are simple, sensitive and
moderate methods. Ascorbic acid (AA), dopamine and catecholamine are important neu-
rotransmiers in the human body. Hence, the low-level detection of ascorbic acid is very
important. All the methods mentioned above involve complicated pretreatment techniques
and expensive instruments. Hence, researchers have developed a more simple, sensitive
and accurate electrochemical method for the detection of ascorbic acid. Several studies have
reported the electrochemical sensing methods for detection of ascorbic acid. Aempts to sim-
plify such methods have resulted in the development of new methods such as the nickel
hexacyanoferrate lm-modied aluminium electrode [58] and graphite epoxy electrode [59].
These methods use specic and modied working electrode systems that are complicated
[60]. These electrochemical methods are widely used to monitor AA metabolites in vivo or
in vitro [61, 62]. For example, batch injection amperometric methods can achieve a detection
limit as low as 450 mg/L [63]. The metallic nickel electrode (2610–6 M), chemiluminometric
ow method (5l M) [64], carbon paste mixed electrode (1.08610–5 M) [65], polyviologen-mod-
ied technique (0.38l M) [66] and NiGCNF plating-modied electrode (2610–6 M) [67] achieve
very sensitive detection ranges.
Suw et al. in 2004 developed a stripping voltammetric technique that manifests faster response,
more cost-eective and sensitive preconcentration techniques, which was used along with a
simpler glassy carbon electrode [68]. In this method, low ascorbic acid concentrations were
detected by the square wave stripping voltammetry and a glassy carbon electrode. The lower
detection limit reported as 0.30l g/L (S/N = 3). This method can be used to detect biological
materials, pharmaceuticals, food and drugs.
Many reports are there for the simultaneous electrochemical detection of ascorbic acid in the
presence of dopamine and uric acid on the glassy carbon electrode. From our own group,
we have synthesized electrically conducting poly (3,4-ethylenedioxythiophene) nanospindles
(PEDOTSs) and used for the electrochemical sensing of ascorbic acid [69] (Figure 10).
Vitamin C14

In another study, our group developed a low-cost electrochemical sensor based on a platinum
electrode for ascorbic acid conductive polyaniline-based composite. This unique low-cost and
user-friendly sensor was validated for the nanomolar detection of AA. The lower detection
limit for AA was observed at 0.1 nm. The aqueous PPICS/Pt electrode could serve as a pro-
spective low-cost and ecient electrochemical sensor and provide promising and outstand-
ing contributions to the food, beverage and medical industries [70].
Electrochemical methods have aracted much aention from clinical diagnostic perspectives
because of their easy operation, low cost, rapid response, high sensitivity and good selectivity.
5. Conclusion
Vitamin C plays a pivotal role in body-building process and in disease prevention. The
various functions of vitamin C, including the antioxidant activity, formation of protein,
tendons, ligaments and blood vessels, for healing wounds and form scar tissue, for repair-
ing and maintaining cartilage, bone, and teeth, and aiding in the absorption of iron, were
discussed. This chapter will denitely benet the students, researchers and technologists
globally.
Author details
Sudha J. Devaki* and Reshma Lali Raveendran
*Address all correspondence to: sudhajd2001@yahoo.co.in
Chemical Sciences and Technology Division, CSIR-National Institute for Interdisciplinary
Science and Technology (CSIR-NIIST), Thiruvananthapuram, India
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