TUGAS RESUME

METABOLISME DAN ZAT GIZI

Nama : Andi Kumala

Nim : J1A120003

Kelas : Gizi Kesehatan Masyarakat

Judul Jurnal Internasional :

1. Cross-Sectional Associations Between Dietary Antioxidant Vitamins

C, E and Carotenoid Intakes and Sarcopenic Indices in Women
Aged 18–79 Years
2. Metabolism of Dietary and Microbial Vitamin B Family in the Regulation of Host
Immunity

Vitamin adalah zat gizi mikro yang memiliki efek fisiologis pada berbagai
respons biologis, termasuk kekebalan inang. Oleh karena itu, kekurangan vitamin
menyebabkan peningkatan risiko mengembangkan penyakit menular, alergi, dan
inflamasi. Karena vitamin B disintesis oleh tanaman, ragi, dan bakteri, tetapi tidak
oleh mamalia, mamalia harus memperoleh vitamin B dari sumber makanan atau
mikroba, seperti mikrobiota usus. Demikian pula, beberapa bakteri usus tidak
dapat mensintesis vitamin B dan harus mendapatkannya dari makanan inang atau
dari bakteri usus lain untuk pertumbuhan dan kelangsungan hidup mereka. Hal ini
menunjukkan bahwa komposisi dan fungsi mikrobiota usus dapat mempengaruhi
penggunaan vitamin B inang dan, dengan perluasan, kekebalan inang.

Meskipun vitamin B makanan umumnya diserap melalui usus kecil,

vitamin B bakteri diproduksi dan diserap terutama melalui usus besar (10,11),
menunjukkan bahwa vitamin B yang diturunkan dari makanan dan usus mungkin
ditangani secara berbeda oleh tubuh manusia. Vitamin B adalah kofaktor dan
koenzim penting dalam beberapa jalur metabolisme, dan baru-baru ini dilaporkan
bahwa vitamin B juga memainkan peran penting dalam pemeliharaan homeostasis
imun.

Regulasi imunologi yang dimediasi vitamin B spesifik untuk sel imun dan
respons imun yang berbeda: yaitu, vitamin B yang berbeda diperlukan untuk
respons imun yang berbeda.Pernah terpikir bahwa vitamin B hanya diperoleh dari
makanan; namun, kita tahu sekarang bahwa ini bukan masalahnya dan bahwa
mikrobiota usus juga merupakan sumber vitamin yang penting. Di dalam
mikrobiota usus, tidak semua bakteri menghasilkan vitamin B dan beberapa
bakteri menggunakan vitamin B makanan atau vitamin B yang diproduksi oleh
bakteri usus lain untuk kebutuhan mereka sendiri; oleh karena itu, mungkin ada
persaingan antara inang dan mikrobiota usus untuk vitamin B
komposisi dan fungsi mikrobiota usus. Oleh karena itu,imunologi yang
diperantarai vitamin pemeliharaan juga bervariasi antar individu. Lebih jauh
pemeriksaan di bidang ini diperlukan, dan informasi baru yang ditemukan akan
membantu mengembangkan era baru kesehatan dan nutrisi yang presisi.

vitamin antioksidan diet (A, C, E dan karotenoid) adalah kandidat yang

menjanjikan untuk pencegahan dan pengobatan hilangnya massa dan fungsi
terkait usia. Vitamin ini mempengaruhi otot rangka dan berfungsi melalui
perannya sebagai antioksidan eksogen dan agen anti-inflamasi. Selain itu, vitamin
C terlibat dalam sintesis kolagen dan karnitin dan retinol dalam metabolisme
protein, pembentukan kolagen, dan oksidasi lipid.

vitamin C digunakan untuk sifat antioksidan dan fungsi sistem kekebalan

tubuh. Oksalat adalah produk akhir dari metabolisme vitamin C. Asam askorbat
yang bersirkulasi diubah menjadi antioksidan jaringan, menghasilkan pembentuk
an radikal askorbat. Radikal tersebut kemudian akan diubah menjadi
dehyroascorbate, yang dapat diubah menjadi asam diketogulonat, molekul tidak
stabil yang dapat dengan mudah terurai menjadi oksalat. Peningkatan konsumsi
asam askorbat menyebabkan pembentukan oksalat yang berlebihan. Kenaikan
kecil oksalat urin dapat meningkatkan risiko pembentukan kristal kalsium oksalat,
dan kadar oksalat urin yang lebih tinggi merupakan faktor risiko utama
pembentukan batu ginjal kalsium oksala. Sayuran penyumbang vitamin C terbesar
adalah paprika, kubis Brussel dan brokoli. Untuk karoten sumber nabati utama
adalah wortel dan bayam sedangkan Buah secara keseluruhan adalah penyumbang
terbesar asupan vitamin C.

Ada relevansi mekanistik yang jelas dari vitamin antioksidan pada

kesehatan otot rangka selama penuaan yang juga mendukung temuan pengamatan
kami. Suplementasi likopen,karoten atau vitamin antioksidan campuran pada
hewan percobaan memiliki efek positif pada kekuatan otot atau aktivitas fisik atau
mengurangi stres oksidatif atau hilangnya otot rangka.vitamin C memiliki efek
perlindungan pada biomarker oksidatif, sitokin inflamasi dan CRP, vitamin C juga
merupakan bagian integral dari sintesis kolagen dan karnitin yang penting untuk
metabolisme asam lemak rantai panjang selama aktivitas fisik.

suprafisiologis vitamin C dapat digunakan untuk memperbaiki asupan

makanan rendah pada kelompok rentan yang lebih tua, ini perlu digunakan dengan
hati-hati karena efek prooksidan yang merugikan dari vitamin C dosis tinggi
jangka panjang (> 500 mg per hari) dibandingkan konsentrasi yang ditemukan
secara alami dalam makanan.
Calcified Tissue International (2020) 106:331–342
https://doi.org/10.1007/s00223-019-00641-x

ORIGINAL RESEARCH

Cross-Sectional Associations Between Dietary Antioxidant

Vitamins C, E and Carotenoid Intakes and Sarcopenic Indices in
Women Aged 18–79 Years
A. A. Welch1 · A. Jennings1 · E. Kelaiditi1,2 · J. Skinner1 · C. J. Steves3

Received: 1 August 2019 / Accepted: 22 November 2019 / Published online: 7 December 2019
© The Author(s) 2019

Abstract
The prevalence of sarcopenia is increasing in aging populations, so prevention is critical. Vitamins (A, C, E and
carotenoids) modify skeletal muscle via protein and collagen synthesis and anti-inflammatory activities. Previous studies
have not inves- tigated intake of these vitamins in relation to sarcopenic indices in both younger and older-aged women.
Indices of skeletal muscle mass (as fat-free mass (FFM) relative to body size) were measured using DXA and leg
explosive power (LEP) using the Nottingham Power Rig in 2570 women aged 18–79 years. Adjusted measures of skeletal
muscle were calculated according to quintiles of vitamin C, E, retinol and carotenoid intake, derived from Food Frequency
Questionnaires, after stratification by age. Higher vitamin C intake was associated with significantly higher indices of
FFM and LEP, (Q5-Q1 = 2.0–12.8%, P < 0.01–0.02). Intakes of total and individual carotenoids were significantly
associated with indices of FFM and LEP (Q5-Q1 = 1.0–7.5%). Vitamin E was significantly associated with FFM% and
FFMBMI only. In mutually adjusted analysis with vitamin C, total carotene, vitamin E and protein in the model, the
strongest associations were with vitamin C. These associations were stronger in younger women (< 65 years). For the first
time, our research shows higher dietary intakes of antioxidant vitamins, particularly vitamin C, is associated with higher
skeletal muscle mass and power in free-living women. These findings have relevance for the treatment and prevention of
frailty and sarcopenia throughout adulthood.

Keywords Sarcopenia · Diet · Vitamin C · Vitamin E · Carotenes · Vitamin A · Skeletal muscle · Grip strength

Background and Aims

and mortality, leading to longer and more expensive hospi-
Across the world, populations are aging rapidly leading to tal stays. These conditions also impact adversely on qual-
increased prevalence of sarcopenia and frailty, conditions ity of life for individuals [1]. The estimated prevalence of
that also increase the risk of osteoporosis, falls, fractures sarcopenia in the community is 1–29% in people over the
age of 60 years, which rises to 17.7–87% in those living in
assisted or residential or care facilities [1–3]. Frailty is esti-
mated to affect 25% of people over the age of 80 years. We
Electronic supplementary material The online version of this need, therefore, strategies to prevent the gradual age-
article (https://doi.org/10.1007/s00223-019-00641-x) contains
related decline in skeletal muscle mass and function, or
supplementary material, which is available to authorized
users. sarcopenic indices, that start in middle age to maintain
intrinsic capacity in older adults [4].
 A. A. Welch The established mechanisms of aging of skeletal muscle
a.welch@uea.ac.uk
include increases in circulating cytokines and production
1
Department of Epidemiology & Public Health, Norwich of reactive oxygen species (ROS), with age which have
Medical School, University of East Anglia, Norwich detri- mental effects on synthesis of protein as well as direct
Research Park, Norwich, Norfolk NR4 7TJ, UK cellular damage of skeletal muscle fibres and DNA [5].
2
Faculty of Sport, Health and Applied Science, St Additionally, there are changes to the proportion, quality
Mary’s University, Waldegrave Road, and viscoelastic properties of the different types of
Twickenham, London TW1 4SX, UK collagen that form the important structural component of
3
Department of Twin Research and Genetic skeletal muscle cells, con- nective tissues and tendons
Epidemiology, King’s College, London, UK
13
during aging [6]. As endogenous

13
33 A. A. Welch et al.

antioxidant efficiency is reduced with aging, and skeletal aged 18–103 years who are representative of singleton
muscle generates the greatest quantities of ROS in the
body, exogenous antioxidant vitamins have potential
importance for skeletal muscle health.
Dietary antioxidant vitamins (A, C, E and carotenoids)
are, therefore, promising candidates for the prevention and
treat- ment of age-related loss of mass and function. These
vitamins influence skeletal muscle and function through
their roles as exogenous antioxidant and anti-inflammatory
agents. In addi- tion, vitamin C is involved in collagen and
carnitine synthesis and retinol in protein metabolism,
collagen formation, and lipid oxidation [7, 8].
To date, dietary treatments for sarcopenia and frailty
have focused largely on interventions with protein intake,
with or without resistance exercise, which is important for
skeletal muscle but the effectiveness of intervention studies
in indi- viduals with sarcopenia and frailty have been
mixed [9, 10]. As there are no effective pharmacological
treatments for sar- copenia, frailty and the loss of skeletal
muscle mass and func- tion with age, identifying other
dietary factors to prevent or attenuate losses of muscle
mass and function in middle and early old age is important.
Whilst limited previous research has studied nutritional
intake or blood concentrations of vitamin A, C, E or carot-
enoids and measures of skeletal muscle mass or function in
older-aged populations, none has measured their relative
effec- tiveness in relation to a range of indices of both
skeletal mass and function in young as well as in older-aged
women [11–19]. Therefore, the purpose of this study was to
first, understand the associations between: (i) dietary
vitamins C, E and A and (ii) the full range of dietary
carotenoids; α-carotene, β-carotene, β-cryptoxanthin,
lycopene, lutein and zeaxanthin and indices of skeletal
muscle mass [i.e. Fat Free Mass Index (FFMI), percentage
fat-free mass (FFM%) and fat-free mass adjusted for body
mass index (FFMBMI)], and function [i.e. hand grip
strength, arm muscle quality and leg explosive power
(LEP)] in a population of women with a wide age range.
Second, to understand whether these associations differed
in young women compared with those over the age of 65
years, third to estimate the relationship between the
inflammatory cytokine C-reactive protein (CRP) and indices
of skeletal muscle health and fourthly, to determine the
relative associations of vitamins A, C, E, total carotene and
protein by including all the nutri- ents in the same
statistical models to determine which of the vitamins was
the most strongly associated with the indices of skeletal
muscle health.

Methods

The TwinsUK registry is an ongoing population study

of approximately 12,000 male and female (83%) twins,

13
Cross-Sectional Associations Between Dietary Antioxidant Vitamins C, E and Carotenoid 33
Intakes…
populations in the United Kingdom [20]. Data were used
from 2570 women who had completed a food frequency
questionnaire (FFQ) and attended for dual-energy X-ray
absorptiometry (DXA) measurements between 1996 and
2000. Within this group, there were 1914 individuals with
measures of leg explosive power and 1658 individuals
with measures of high sensitivity C-reactive protein (hs-
CRP). Between 2005 and 2008, 949 women completed an
FFQ and had grip strength measured (including 512
individu- als from the first cohort) (Supplementary Fig.
1). Ethical approval was obtained from St. Thomas’s
Hospital Research Ethics Committee and informed
consent was acquired from all participants.

Dietary Intake

Participants completed a 131 item-validated FFQ with

nutrient values calculated using the UK national nutrient
database and a database of the carotenoid composition of
foods [21, 22]. Individuals were excluded from the
dietary analyses if answers to > 10 food items were left
blank or the ratio of estimated total energy intake to the
estimated basal metabolic rate fell 2 SDs outside the
mean ratio. Energy- reporting quality was estimated using
the ratio of reported energy intake (EI) to estimated
energy expenditure (EER), the EI:EER ratio expressed as
the percentage of EI to EER, was calculated and was
included as a covariate for adjust- ment in the statistical
analyses [23].

Muscle Mass, Strength and Power

Fat-free mass (FFM) was measured by DXA scans (Hol-

ogic QDR-2000 DXA scanner, Hologic Inc., Waltham,
MA, USA) and adjusted to body size using the following
cal- culations: FFMI: FFM (kg)/ height (m2) [1], FFM%:
FFM (kg)/ weight (kg) *100 and FFMBMI: FFM (kg)/ BMI
(kg/ m2). FFM (kg) divided by BMI (kg/m2) (FFMBMI)
takes into account the increase in body size, scaled for
height and adjusting FFM for BMI is likely to set
meaningful cut- off points for clinical use [1]. Isometric
grip strength was assessed using a Jamar hand grip
dynamometer (Sammons, Preston, UK) on the dominant
arm with reproducibility assessed by repeated
measurement on 24 individuals (CV of 11.4%) [24] and
adjusted for mean arm lean mass as arm muscle quality
(grip strength (kg)/ mean arm lean mass (kg). LEP was
measured using the Nottingham Power Rig which
assesses the force and velocity of muscle contraction
from the quadriceps, normalised for body weight (LEP
(watts)/ weight (kg)) [25]. LEP is validated with high
reliability (reli- ability coefficient 0.97, coefficient of
variation 9.4%, over one week period in adults) [25].
13
33 A. A. Welch et al.

C-reactive Protein smoking status (never, former, current), energy

Circulating hs-CRP was measured by a highly sensitive

automated microparticle capture enzyme immunoassay,
standardised on the World Health Organisation
International Reference Standard for CRP immunoassay as
previously described [26].

Covariates

Information on lifestyle, medication use, menopausal

status, and demographic variables were obtained using
standardised nurse-administered questionnaires. Weight
and height were measured to the nearest 0.1 kg and to the
nearest 0.5 cm, respectively. BMI was calculated as weight
in kilograms (kg) divided by height in meters squared (m2).
Leisure- and work-time physical activity was self-reported
using a ques- tionnaire significantly correlated with in
depth assessments of physical activity in a subset of this
cohort [27]. For par- ticipants in the grip strength group (n
= 949 women) who had missing data on physical activity
(N = 10), the General Practice Physical Activity
Questionnaire (GPPAQ) was used. This was based on the
original physical activity index developed by the European
Prospective Investigation into Cancer and Nutrition (EPIC)
cohort. The EPIC physical activity index was previously
validated against heart rate monitoring in two independent
studies [28].

Statistical Analyses

Descriptive statistics (means ± SDs or % (n)) were

analysed for all participants and for those under 65 years (n
= 2346) and ≥ 65 years (n = 224). Multivariate regression
analysis was used to calculate statistical trends and
adjusted val- ues (least square means) for FFM%, FFMI,
FFMBMI, LEP, arm muscle quality and hs-CRP (outcomes)
classified by quintile of dietary vitamins C, E and A and
dietary carot- enoids; α-carotene, β-carotene, β-
cryptoxanthin, lycopene, lutein and zeaxanthin (exposures).
The percentage difference between quintiles 1 and 5 was
calculated as a percentage of the value in quintile 1. As
physical activity and smoking habit were related to indices
of skeletal muscle the analy- ses were adjusted for physical
activity and smoking habit. All models were adjusted for
age (years), physical activity (active, moderately active,
inactive), smoking status (never, former, current), energy
intake (kcal/d) and potential mis- reporting of energy
intake (EI:EER). FFMI was adjusted for age (years),
physical activity (active, moderately active, inactive),

13
Cross-Sectional Associations Between Dietary Antioxidant Vitamins C, E and Carotenoid 33
Intakes… 2
intake (kcal/d), potential mis-reporting of energy intake trend < 0.001), FFMI was 0.4 kg/ m higher (± 0.1 P-trend
(EI:EER) and fat mass (kg). LEP and arm muscle = 0.002), FFMBMI was 0.03 kg/kg/ m2 higher (± 0.01 P-
qual- ity were adjusted for age (years), physical activity trend = 0.023) and LEP was 10.9 w/ kg higher (± 2.6 P-
(active, moderately active, inactive), smoking status trend < 0.001), when comparing extreme
(never, former, current), energy intake (kcal/d), potential
mis-reporting of energy intake (EI:EER) and menopausal
status (pre-meno- pausal/ post-menopausal), use of HRT
(yes/no) and height (m). High sensitivity (hs) CRP was
adjusted for age (years), physical activity (active,
moderately active, inactive), smok- ing status (never,
former, current), energy intake (kcal/d), potential mis-
reporting of energy intake (EI:EER), and BMI, use of
anti-inflammatory medications (yes/no) and HRT
(yes/no). Values for hs-CRP were skewed and therefore,
natural log-transformed values were used for the
analyses.
In analysis stratified by age, we also assessed the
associa- tions of FFM%, FFMI, FFMBMI and LEP with
quintiles of intake for those under 65 years (n = 2346)
and tertiles for those ≥ 65 years (n = 224). The data for
those ≥ 65 years was analysed for tertiles due to the
smaller number of women in this group. For these and
the comparative analyses we only considered vitamins A,
C, E, total carotene and protein intakes as these
demonstrated the greatest associations, as shown in Table
2. In additional analyses, to compare the relative
associations of vitamins A, C, E, total carotene and
protein we included all these nutrients in the same statisti-
cal models and standardised the values for FFM%, FFMI,
FFMBMI and LEP using Z-scores. Values for the FFM
indi- ces and LEP were also adjusted for covariates as
described previously, and as in the models shown in
Table 3.
Values in the text are means ± SE. A P value < 0.05
was considered statistically significant. P trend was
calculated from the multivariable regression across
quintiles of intake of the nutrients. All analyses were
performed with Stata statistical software version 14.0
(Stata Corp, College Sta- tion, TX) and included the
robust cluster regression option in STATA to account for
clustering within twin pairs.

Results

The characteristics of the participants are presented in

Table 1, stratified by age (< 65 years and ≥ 65 years). The
mean age (±SD) of participants was 48.37 ± 12.7 years
and mean (± SD) BMI was 24.9 ± 4.14 kg/m 2. More than
one- half of the participants were moderately active
(53.9%), and 18.2% were current smokers. In
multivariable analyses, women in the highest quintile of
vitamin C intake had sign- ficantly higher FFM%, FFMI,
FFMBMI and LEP, compared to those in the lowest quintile
(Table 2). Specifically, FFM% was 1.4% higher (± 0.4 P-

13
33 A. A. Welch et al.

intake quintiles. In terms of vitamin E, FFM% (Q5-Q1

1.6% ± 0.5 P-trend = 0.002) and FFMBMI (Q5-Q1 0.06 ±
0.02
P-trend = 0.002) were positively associated with intakes.
For total carotene, associations were also significant for
FFM% (Q5-Q1 0.6% ± 0.4 P-trend = 0.028), FFMI (Q5-
Q1 0.2 ± 0.1
P-trend = 0.012) and LEP (Q5-Q1 6.4 ± 2.7 P-trend =
0.011).
The results of the associations between the nutrients and
arm muscle quality were non-significant ranging from a
beta coefficient (95% CI) of − 0.075 (− 0.199, 0.048) per
quintile for vitamin C intake (P-trend = 0.231) to 0.036 (−
0.150, 0.222) per quintile for vitamin E intake (P-trend =
0.705). The results of the associations between the
nutrients and hs-CRP were also non-significant ranging
from a beta coef- ficient (95% CI) of − 0.008 (− 0.046,
0.029) per quintile for vitamin C intake (P-trend = 0.666)
to − 0.023 (− 0.070, 0.025) per quintile for vitamin E
intake (P-trend = 0.349). Also no associations were
observed for vitamin A (measured as retinol).
Age-Stratified Analyses
In age-stratified analyses, we observed that associations
between vitamin C and E with FFM% were observed in
the younger (< 65 years) but not the older (≥ 65 years)
subset (Table 3). Specifically, vitamin C was associ-
ated with a 0.3% (95% CI 0.1, 0.5 P-trend = 0.002) and
vitamin E with a 0.4% (95% CI 0.2, 0.7 P-trend = 0.001)
higher FFM% per quintile of intake. Likewise, associa-
tions between vitamin C (2.3 95% CI 1.1, 3.6 per quintile
P-trend < 0.001) and total carotene (1.7 95% CI 0.4, 2.9
per quintile P-trend = 0.009) and LEP (w/kg) were only
observed in participants aged < 65 years. Similar trends
were also found with FFMI and FFM/BMI, although there
was no significant association between FFM/BMI and vita-
min C in those < 65 years.
Standardised Analyses

In standardised analyses, we compared the relative

Individual Carotenoids
Intakes of all the individual carotenoids, with the excep-
tion of leutein + zeaxanthin, were significantly associated
with FFMI. The strongest association was observed for α-
carotene intake (Q5-Q1 0.24 kg/m 2 ± 0.1 P-trend = 0.03,
1.6%). Significant associations were also found with FFM
% for ß-cryptoxthanin and with FFM% and FFMBMI for
lutein and zeaxthanin, with interquintle differences
ranging from
1.1 to 7.2%. LEP was associated significantly with the
carot- enoids, with the exception of α-carotene, with
differences ranging from 6.3 to 7.5% when comparing
extreme quintiles of intake.
associations of vitamins C, E, retinol and total caro-
tene with FFM% and LEP (Figs. 1, 2). Associations
between vitamin C and FFM% and LEP were maintained
after mutual adjustment for the other nutrients, includ-
ing protein (FFM% 0.17 SD ± 0.07 P-trend = 0.013
and LEP 0.27 SD ± 0.08 P-trend = 0.004). A signifi-
cant negative association with protein was observed for
FFM% (− 0.18 SD ± 0.06 P-trend < 0.001), FFMBMI
– 0.27 ± 0.06 P-trend < 0.001 and LEP (− 0.14 SD ± 0.07
P-trend = 0.007) but the association with FFMI was not

Fig. 1 The relative associa- P-

tions of vitamin C, vitamin E, tren 0.7
retinol, carotene and protein d< 0.6
with percentage fat-free mass 0.05 0.5
in 2570 females aged 18–79
0.4
years, stratified by age1. Values
repre- sent the difference in 0.3
stand- 0.2
ardised values of percentage 0.1
fat-free mass between
SD

partici- 0
pants in Q5 vs Q1 of intake -0.1
(T3- T1 for > 65 years sub- -0.2
group) with all nutrients
-0.3
included in the model. Values
were also adjusted for age, -0.4
physical activity, smoking -0.5
status, energy intake and
-0.6
underreporting. *

13
Cross-Sectional Associations Between Dietary Antioxidant Vitamins C, E and Carotenoid 33
Intakes…
-0.7 Protein

* *

* *

A
ll
(n
=
2
5
7
0)

<
6
5
y
(n
=
2
3
4
6)

≥
6
5
y
(n
=
2
2
4)
Vita
min
C

Vita
min
E

Tot
al
caro
tene

Reti
nol

13
33 A. A. Welch et al.

Table 1 Characteristics and

dietary intakes of n = 2570
C haracteristic All < 65 years ≥ 65 years
females aged 18–79 years,
stratified by agea n= 2570 2346 224
Age (years) 48.3 ± 12.7 46.5 ± 11.6 68.0 ± 2.81
BMI (kg/m2) 24.9 ± 4.14 24.8 ± 4.11 26.1 ± 4.23
Fat mass (kg) 22.7 ± 7.87 22.5 ± 7.90 25.0 ± 7.26
Fat free mass (%) 61.1 ± 6.50 61.4 ± 6.46 58.0 ± 6.08
Fat Free Mass Index (kg/m2) 15.0 ± 1.72 15.0 ± 1.71 15.0 ± 1.80
Fat-free mass/ BMI (kg/ kg/m2) 1.62 ± 0.23 1.63 ± 0.23 1.47 ± 0.19
Leg explosive powerb (watts) 89.8 ± 36.8 91.3 ± 36.7 67.5 ± 30.8
Leg explosive powerb (watts/kg) 90.9 ± 36.5 92.4 ± 36.3 68.3 ± 31.6
hs-CRPc (mg/L) 2.49 ± 2.30 2.47 ± 2.30 2.72 ± 2.26
Grip strengthd (kg) 28.8 ± 5.95 30.1 ± 5.66 25.3 ± 5.20
Energy intake (kcal/d) 1979 ± 524 1978 ± 533 1989 ± 415
Protein (% energy) 16.6 ± 2.62 16.6 ± 2.62 16.9 ± 2.68
Vitamin C (mg/d) 155 ± 80.2 154 ± 80.4 165 ± 78.3
Vitamin E (mg/d) 11.4 ± 4.57 11.3 ± 4.60 11.7 ± 4.26
Total carotene (µg/d) 3448 ± 1944 3416 ± 1964 3777 ±
1688
α-Carotene (µg/d) 559 ± 416 554 ± 418 621 ± 389
β-Carotene (µg/d) 3091 ± 1757 3062 ± 1775 3398 ±
1529
Retinol (µg/d) 559 ± 789 545 ± 793 700 ± 728
β-Cryptoxanthin (µg/d) 200 ± 194 201 ± 197 192 ± 166
Lycopene (µg/d) 1347 ± 958 1364 ± 968 1165 ± 820
Lutein + zeaxanthin (µg/d) 2267 ± 1478 2253 ± 1501 2413 ±
1204
Underreporting (EI:EER, %) 87.4 ± 24.6 86.7 ± 24.6 95.0 ± 24.2
Physical activity (active, %) 24.2% (622) 23.9% (561) 27.2% (61)
Moderately active (%) 53.9% (1385) 54.2% (1271) 50.9% (114)
Inactive (%) 21.9% (563) 21.9% (514) 21.9% (49)
Smoking status (current, %) 18.2% (468) 19.1% (449) 8.48% (19)
Menopausal status (post-menopausal, %) 47.4% (1218) 42.5% (997) 98.7% (221)
Anti-inflammatory medicationc (yes, %) 6.15% (102) 6.06% (90) 6.98% (12)
Hormone replacement therapyc (yes, %) 6.33% (105) 6.33% (94) 6.40% (11)

EI:EER ratio of reported energy intake to estimated energy requirements

a
Values are mean ± SD or % (n), n = 2570
Values for a subset of b1914
c
1658
d
949 participants

significant 0.05 SD ± 0.06 P-trend = 0.390. The results vitamin E, the food groups cakes and biscuits and whole
for vitamin E, not shown in the figures were: FFMI grain cereals contributed more to intake than vegetables.
– 0.05 SD ± 0.08 P-trend = 0.487 and FFMBMI 0.19 ± 0.09
P-trend = 0.025.
Total vegetable intake was a significant source of vita-
mins C, E and carotene intake, however, the specific veg-
etables that contributed differed for each nutrient (Fig. 3).
For vitamin C the main contributors were peppers, Brus-
sels sprouts and broccoli, for vitamin E avocado, mush-
rooms and spinach, and for carotene, carrots and spinach.
Fruit intake also contributed to vitamin C intake, and for

13
Cross-Sectional Associations Between Dietary Antioxidant Vitamins C, E and Carotenoid 33
Intakes…
Discussion

Our findings of significant associations between higher

intakes of vitamin C, total carotene and specific carote-
noids and improved sarcopenic indices of skeletal
muscle; FFM and LEP were found in a cohort of women
with a wide age range (18–79 years). The scale of the
associa- tions for these vitamins ranged from 1.0 to 3.2%
for indi- ces of FFM and from 6.3 to 12.8% for LEP,
comparing the highest and lowest intakes and adjusting
for relevant covariates including age. This highlights the
potential clin- ical importance of the findings as the
yearly losses of FFM

13
33 A. A. Welch et al.

Table 2 Fat free mass indices

Intakeb FFM (%) FFM/BMI FFMI (kg/mb) LEPc (w/kg)
and leg explosive power by
quintile of nutrient intake in Vitamin C (mg/d)
2570 females aged 18–79
yearsa Q1 68.8 ± 16.4 60.4 (59.8,60.9) 1.60 (1.58,1.62) 14.8 (14.7,15.0) 85.2 (81.8,88.6)
Q2 108 ± 9.43 60.9 (60.4,61.4) 1.61 (1.59,1.63) 15.0 (14.9,15.2) 90.8 (87.2,94.4)
Q3 141 ± 10.1 61.1 (60.6,61.6) 1.62 (1.60,1.64) 15.0 (14.9,15.1) 90.1 (86.4,93.8)
Q4 181 ± 13.8 61.3 (60.8,61.8) 1.62 (1.60,1.64) 15.1 (14.9,15.2) 92.0 (88.2,95.9)
Q5 276 ± 80.3 61.8 (61.3,62.3) 1.63 (1.61,1.65) 15.2 (15.1,15.3) 96.1 (92.3,99.9)
P-trend – < 0.01 0.02 < 0.01 < 0.01
Q5-Q1% – 2.36 1.99 2.40 12.79
Vitamin E (mg/d)
Q1 5.90 ± 1.16 60.3 (59.7,60.9) 1.58 (1.56,1.61) 14.9 (14.8,15.1) 89.2 (84.8,93.6)
Q2 8.59 ± 0.63 60.7 (60.2,61.2) 1.60 (1.58,1.62) 15.1 (14.9,15.2) 90.6 (86.9,94.3)
Q3 10.7 ± 0.64 61.3 (60.8,61.8) 1.63 (1.61,1.65) 15.1 (14.9,15.2) 91.9 (88.1,95.8)
Q4 13.3 ± 0.81 61.2 (60.7,61.8) 1.62 (1.60,1.64) 15.0 (14.9,15.2) 90.9 (87.3,94.5)
Q5 18.3 ± 3.39 61.9 (61.2,62.5) 1.64 (1.62,1.66) 15.0 (14.8,15.2) 91.6 (87.1,96.2)
P-trend – < 0.01 < 0.01 0.70 0.54
Q5-Q1% – 2.60 3.54 0.42 2.77
Total carotene (µg/d)
Q1 1342 ± 377 60.8 (60.3,61.3) 1.61 (1.59,1.63) 14.9 (14.7,15.0) 85.6 (81.8,89.3)
Q2 2379 ± 320 60.9 (60.4,61.4) 1.61 (1.59,1.63) 15.0 (14.9,15.1) 91.2 (87.7,94.8)
Q3 3223 ± 189 60.8 (60.3,61.3) 1.61 (1.59,1.63) 15.0 (14.9,15.2) 90.8 (87.0,94.5)
Q4 3991 ± 321 61.6 (61.1,62.1) 1.63 (1.61,1.65) 15.1 (15.0,15.3) 94.8 (91.1,98.4)
Q5 6303 ± 2107 61.4 (60.9,62.0) 1.62 (1.60,1.64) 15.1 (14.9,15.2) 91.9 (88.2,95.7)
P-trend – 0.03 0.11 0.01 0.01
Q5-Q1% – 1.03 0.96 1.44 7.45
α-Carotene (µg/d)
Q1 131 ± 62.8 61.1 (60.6,61.7) 1.62 (1.60,1.64) 14.9 (14.8,15.0) 88.1 (84.6,91.7)
Q2 265 ± 88.2 61.1 (60.6,61.6) 1.62 (1.60,1.64) 15.0 (14.9,15.1) 90.8 (86.9,94.8)
Q3 605 ± 9.02 60.8 (60.3,61.2) 1.60 (1.58,1.62) 15.1 (14.9,15.2) 91.8 (88.3,95.3)
Q4 638 ± 11.7 61.0 (60.5,61.6) 1.62 (1.59,1.64) 15.0 (14.9,15.2) 92.8 (89.2,96.5)
Q5 1158 ± 467 61.4 (60.9,62.0) 1.62 (1.60,1.64) 15.1 (15.0,15.3) 90.7 (86.9,94.4)
P-trend – 0.62 0.94 0.03 0.24
Q5-Q1% – − 0.46 – 0.02 – 1.61 2.85
β-Carotene (µg/d)
Q1 1194 ± 344 60.7 (60.2,61.2) 1.60 (1.58,1.63) 14.9 (14.7,15.0) 85.6 (81.8,89.4)
Q2 2139 ± 272 61.0 (60.6,61.5) 1.61 (1.60,1.63) 15.0 (14.9,15.2) 91.4 (87.8,94.9)
Q3 2861 ± 175 60.9 (60.4,61.4) 1.61 (1.59,1.63) 15.0 (14.9,15.1) 90.1 (86.4,93.8)
Q4 3589 ± 294 61.4 (60.9,61.9) 1.62 (1.60,1.64) 15.1 (15.0,15.3) 95.3 (91.6,98.9)
Q5 5673 ± 1911 61.4 (60.9,62.0) 1.62 (1.60,1.65) 15.1 (14.9,15.2) 91.9 (88.2,95.7)
P-trend – 0.05 0.17 0.01 < 0.01
Q5-Q1% – 1.20 1.23 1.50 7.38
Retinol (µg/d)
Q1 137 ± 43.1 61.2 (60.7,61.8) 1.61 (1.59,1.63) 15.1 (14.9,15.3) 92.1 (88.1,96.2)
Q2 245 ± 25.1 60.8 (60.3,61.3) 1.60 (1.58,1.62) 15.0 (14.9,15.2) 90.7 (87.0,94.5)
Q3 347 ± 36.6 61.0 (60.5,61.5) 1.61 (1.59,1.63) 15.0 (14.8,15.1) 89.7 (86.2,93.2)
Q4 639 ± 187 61.4 (60.8,61.9) 1.63 (1.60,1.65) 15.1 (15.0,15.3) 90.8 (87.2,94.3)
Q5 1426 ± 1412 61.0 (60.5,61.6) 1.62 (1.61,1.64) 14.9 (14.8,15.1) 91.0 (87.0,95.0)
P-trend – 0.85 0.14 0.23 0.78
Q5-Q1% − 0.36 0.91 – 1.30 – 1.24
β-Cryptoxanthin (µg/d)
Q1 38.3 ± 16.8 60.7 (60.2,61.2) 1.60 (1.58,1.62) 14.9 (14.8,15.0) 87.7 (84.0,91.4)
Q2 82.9 ± 12.1 61.0 (60.5,61.5) 1.62 (1.60,1.64) 15.0 (14.8,15.1) 89.1 (85.6,92.5)

13
Cross-Sectional Associations Between Dietary Antioxidant Vitamins C, E and Carotenoid 33
Intakes…
Table 2 (continued)
Intakeb FFM (%) FFM/BMI FFMI (kg/mb) LEPc (w/kg)
Q3 146 ± 25.2 61.0 (60.5,61.4) 1.61 (1.59,1.63) 15.1 (15.0,15.2) 90.6 (87.1,94.1)
Q4 234 ± 36.4 61.5 (61.0,62.0) 1.63 (1.61,1.65) 15.0 (14.9,15.2) 93.7 (90.2,97.3)
Q5 499 ± 230 61.4 (60.8,61.9) 1.62 (1.60,1.64) 15.1 (15.0,15.3) 93.2 (89.4,97.0)
P-trend – 0.03 0.22 0.03 < 0.01
Q5-Q1% – 1.09 0.87 1.50 6.30
Lycopene (µg/d)
Q1 412 ± 168 61.0 (60.4,61.5) 1.61 (1.59,1.63) 15.0 (14.8,15.1) 87.2 (83.7,90.7)
Q2 821 ± 99.2 61.3 (60.8,61.8) 1.63 (1.61,1.65) 15.0 (14.8,15.1) 91.5 (87.4,95.5)
Q3 1161 ± 100 61.4 (60.9,61.8) 1.63 (1.61,1.65) 15.0 (14.9,15.1) 93.3 (89.6,97.1)
Q4 1573 ± 154 61.2 (60.7,61.6) 1.62 (1.60,1.63) 15.1 (14.9,15.2) 89.8 (86.2,93.4)
Q5 2771 ± 1121 60.6 (60.1,61.2) 1.59 (1.57,1.61) 15.1 (15.0,15.3) 92.5 (88.5,96.5)
P-trend – 0.33 0.05 0.03 0.18
Q5-Q1% – – 0.58 – 1.57 1.29 6.05
Lutein + zeaxanthin (µg/d)
Q1 835 ± 261 60.5 (60.0,61.1) 1.59 (1.57,1.61) 15.0 (14.8,15.1) 88.4 (84.6,92.2)
Q2 1462 ± 153 60.6 (60.2,61.1) 1.60 (1.58,1.62) 15.0 (14.9,15.2) 87.7 (84.4,91.0)
Q3 1970 ± 152 61.1 (60.6,61.6) 1.62 (1.60,1.64) 15.0 (14.9,15.2) 90.3 (86.8,93.8)
Q4 2602 ± 216 61.3 (60.8,61.9) 1.62 (1.61,1.64) 15.1 (14.9,15.2) 93.1 (89.4,96.8)
Q5 4464 ± 1744 61.9 (61.3,62.4) 1.64 (1.62,1.66) 15.0 (14.9,15.2) 94.8 (91.0,98.6)
P-trend – < 0.01 < 0.01 0.37 < 0.01
Q5-Q1% – 2.27 3.15 0.52 7.23

FFM fat-free mass, FFMI Fat Free Mass Index and LEP leg explosive power
a
Values are adjusted means (least square means) ± SE, n = 2570. Means were adjusted for age, physical
activity, smoking status, energy intake, protein intake and underreporting and Fat Free Mass Index was
additionally adjusted for fat mass. Participant numbers by quintile were Q1 = 514; Q2 = 514; Q3 =
514;
Q4 = 514; Q5 = 514
b
Intake values are unadjusted means ± SD
c
Values are mean (least square means) ± SE, n = 1914. Means were adjusted for age, physical activ-
ity, smoking status, energy intake, protein intake, underreporting, menopausal status, hormone replace-
ment therapy and height. Participant numbers by quintile were Q1 = 383; Q2 = 383; Q3 = 383; Q4 = 383;
Q5 = 382

and strength range from 1%, for skeletal muscle mass, and
3% for grip strength, for people over the age of 50 years
[29]. Moreover, using standardised analyses to compare
the different vitamins and protein, the associations were
greatest for vitamin C, despite mutual adjustment for all
the nutrients investigated in our study, including protein
intake. This research highlights the importance of dietary
associations with muscle mass in individuals of all ages,
supporting previous research that has shown that notable
changes in skeletal muscle mass occur earlier in adult life
(between 30 and 45 years of age) [30, 31].
The range of nutrients across quintiles of intake investi-
gated in our study varied from 3.1 fold for vitamin E, and
fourfold for vitamin C, to 4.6 fold for total carotene and
to larger differences for retinol (10.4 fold). However, the
associations with vitamin E intake were only found with
FFM expressed either as a percentage or divided by BMI,
and no associations were found with retinol.
Different foods contributed to intake of the nutrients
inves- tigated. Although vegetables were the most significant
source, the specific vegetables contributing to these nutrients
differed. The largest vegetable contributors to vitamin C
were peppers, Brussels sprouts and broccoli. For carotene
the major veg- etable sources were carrots and spinach
whereas vitamin E was supplied by avocado and
mushrooms. Overall fruit was the greatest contributor to
vitamin C intakes, and whole grain cereals were the greatest
contributor to vitamin E intake. Our findings highlight the
importance of eating a broad range of vegetables, fruits and
whole grain cereal foods to achieve opti- mal intakes of
vitamin C, carotenes and vitamin E.

13
33 A. A. Welch et al.

Table 3 Relative associations between indices of fat-free mass and leg explosive power calculated according to quantile of nutrient intake in
2570 females aged 18–79 years, stratified by agea

All (n = 2570) < 65 years (n = 2346) ≥ 65 years (n = 224)

Quintile of nutrient intake β (95% CI) P-trend β (95% CI) P-trend β (95% CI) P-trend

FFM (%)
Vitamin C (mg/d) 0.28 (0.1, 0.5) < 0.01 0.30 (0.1, 0.5) < 0.01 0.50 (− 0.5, 1.5) 0.32
Vitamin E (mg/d) 0.41 (0.2, 0.6) < 0.01 0.41 (0.2, 0.7) < 0.01 0.31 (− 0.7, 1.4) 0.55
Total carotene (µg/d) 0.14 (0.0, 0.3) 0.12 0.13 (− 0.1, 0.3) 0.16 0.63 (− 0.2, 1.5) 0.14
Retinol (µg/d) 0.00 (− 0.2, 0.2) 0.99 – 0.01 (− 0.2, 0.2) 0.92 0.32 (− 0.6, 1.3) 0.51
Protein (%E/d) – 0.32 (− 0.5, − 0.2) < 0.01 – 0.34 (− 0.5, − 0.2) < 0.01 – 0.50 (− 1.5, 0.5) 0.34
LEPb (w/kg)
Vitamin C (mg/d) 2.14 (0.9, 3.3) < 0.01 2.32 (1.1, 3.6) < 0.01 – 1.06 (− 9.0, 6.9) 0.79
Vitamin E (mg/d) 0.66 (− 1.0, 2.3) 0.43 0.60 (− 1.2, 2.4) 0.51 – 0.76 (− 9.4, 7.8) 0.86
Total carotene (µg/d) 1.35 (0.1, 2.6) 0.03 1.67 (0.4, 2.9) < 0.01 – 6.68 (− 15.0, 1.6) 0.11
Retinol (µg/d) – 0.23 (− 1.5, 1.1) 0.73 – 0.39 (− 1.8, 1.0) 0.57 0.26 (− 8.8, 9.4) 0.95
Protein (%E/d) – 1.18 (− 2.3, 0.0) 0.04 – 1.18 (− 2.4, 0.0) 0.05 – 2.64 (− 8.6, 3.3) 0.38
FFMI (kg/m2)
Vitamin C (mg/d) 0.07 (0.0, 0.1) < 0.01 0.08 (0.03, 0.12) < 0.01 0.02 (− 0.3, 0.3) 0.91
Vitamin E (mg/d) 0.00 (− 0.05, 0.06) 0.85 – 0.01 (− 0.07, 0.05) 0.70 0.23 (− 0.1, 0.6) 0.17
Total carotene (µg/d) 0.06 (0.02, 0.1) < 0.01 0.05 (0.01, 0.1) 0.02 0.22 (− 0.03, 0.46) 0.09
Retinol (µg/d) – 0.03 (− 0.08, 0.02) 0.26 – 0.04 (− 0.1, 0.01) 0.11 0.13 (− 0.16, 0.43) 0.38
Protein (%E/d) 0.03 (− 0.01, 0.07) 0.15 0.03 (− 0.01, 0.1) 0.14 0.05 (− 0.23, 0.33) 0.73
FFM/BMI
Vitamin C (mg/d) 0.00 (− 0.0, 0.01) 0.08 0.01 (− 0.00, 0.01) 0.11 0.02 (− 0.01, 0.05) 0.26
Vitamin E (mg/d) 0.01 (0.0, 0.02) < 0.01 0.02 (0.01, 0.03) < 0.01 – 0.00 (− 0.04, 0.04) 0.96
Total carotene (µg/d) 0.00 (− 0.0, 0.0) 0.49 0.00 (− 0.00, 0.01) 0.54 0.02 (− 0.01, 0.05) 0.24
Retinol (µg/d) 0.00 (− 0.0, 0.01) 0.23 0.00 (− 0.00, 0.01) 0.24 0.01 (− 0.02, 0.05) 0.43
Protein (%E/d) – 0.02 (− 0.02, − 0.01) < 0.01 – 0.02 (− 0.02, − 0.01) < 0.01 – 0.02 (− 0.05, 0.02) 0.38

FFM fat free mass and LEP leg explosive power

a
Values are adjusted beta coefficients (95% CI) per quantile of intake, n = 2570 (Quintiles for all ages and < 65 years and tertiles for
those ≥ 65 year). Models were adjusted for age, physical activity, smoking status, energy intake and underreporting
b
Subset analysis n = 1914 (< 65 years n = 1794; ≥ 65 years n = 120). Means were adjusted for age, physical activity, smoking status, energy
intake, underreporting, menopausal status, hormone replacement therapy and height. Dietary variables are expressed per quintile for all partici-
pants and participants < 65 years and tertiles for participants ≥ 65 years

Comparison with Other Studies
The positive associations we found between higher intakes
of vitamin C, carotenoids and vitamin E are, in the main,
supported by the few previous human studies that investi-
gated the relationships between intake or circulating con-
centrations of vitamin C, E or carotene and sarcopenic
indices in cohorts of older people [11–14]. Dietary vitamin
C was associated with measures of skeletal muscle func-
tion in the InCHIANTI Study, and in women only in the
UK Hertfordshire Cohort Study (HCS) [11–14]. Circulat-
ing vitamin C and sarcopenic indices were only previously
examined in elderly Japanese women where positive asso-
ciations were found between measures of physical function
but not FFM [13]. The only study investigating vitamin C
13
intake with FFM found positive associations but only after
2.6 years of follow-up [11].
Circulating vitamin E was investigated in only three pre-
vious studies where plasma α-tocopherol was positively
associated with measures of grip and knee strength, perfor-
mance or decline in physical function. However, two stud-
ies found no associations between intake of vitamin E and
either grip strength or function [12, 14, 17, 32]. Our
finding of a positive relationship with dietary vitamin E
and certain indices of fat-free mass further supports the
relevance of vitamin E to skeletal muscle health.
For carotenoids, two previous studies found associations
with either circulating β or total carotene and greater
muscle strength or physical activity, with only one other
study find- ing positive associations between higher
intakes of carotene and strength or physical activity, in
women [12, 14–19]. Our
Cross-Sectional Associations Between Dietary Antioxidant Vitamins C, E and Carotenoid 33
Intakes…
Fig. 2 The relative associa-
tions of vitamin C, vitamin E, 0.8
retinol, carotene and protein 0.7
with leg explosive power in 0.6
1914 females aged 18–79 0.5 *
*
years, stratified by age1. Values 0.4
repre- sent the difference in 0.3
stand- 0.2
ardised values of leg explosive 0.1
power between participants in 0
Q5 vs Q1 (T3-T1 for > 65 years

SD
-0.1
sub-group) of intake with all -0.2
nutrients included in the model. -0.3
Values were also adjusted for -0.4 * *
age, physical activity, smoking -0.5
status, energy intake, underre- -0.6
porting, height, menopausal sta- -0.7
tus and use of hormone replace- -0.8
ment therapy. * P-trend < 0.05 -0.9
-1
All (n=2570) <65 y (n=2346) ≥65 y (n=224)
Vitamin C Vitamin E Total carotene Retinol Protein

Fig. 3 Foods that contributed 80

to at least 10% of vitamin C, 70
vitamin E, carotene and retinol
intakes in 2570 females aged 60
18–79 years1. Values are the 50
percentage contribution of 40
%

individual foods to total nutrient 30

intake. The main contributors to 20
vegetable intakes were peppers, 10
Brussels sprouts and broc-
0
coli for Vitamin C, avocado,
Fruits

Offal
Cakes and biscuits
Wholegrain cereals

Wholegrain cereals

Dairy
Vegetables

Vegetables

Vegetables
mushrooms and spinach for
Vitamin E and carrots and
spinach for carotene. For Vita-
min C the main contributors to
fruit intakes were strawberries,
oranges and grapefruit
due to LEP being a direct measure of lower limb strength
Vitamin C Vitamin E Carotene Retinol

findings extend the previous research by identifying that

the full range of carotenoids also have importance for
skeletal muscle health.
The lack of association with retinol intake and sarco-
penic indices found in our study contrasts with the only
other human study that found greater intakes of retinol
were significantly related to loss of appendicular lean mass
[11]. In recent studies, relating intakes of either vitamin C,
E,
or circulating vitamin E, to sarcopenia no associations
were found [33–35] although greater prevalence of frailty
was associated with either lower concentrations of
carotenoids, retinol or α-tocopherol, in a further three
studies [36–39].
The reason for the lack of association between the vita-
mins in our study and grip strength, compared with the
strong associations with LEP are not clear but may be
13
34
and power which may be more sensitive to the effects of
diet [29].
The lack of association between the vitamins and
CRP in our study may be due to the relatively high
intakes of vitamins, coupled with the relatively low blood
concentra- tions of hsCRP in this population.
13
A. A. Welch et al.
Although we also found smaller associations with the
sarcopenic indices and protein intake than the vitamins in
our standardised analyses, other studies have found protein
is important for skeletal muscle. However, there has been
variability in effectiveness of supplementation with protein
in intervention studies, and protein intake in our study was
higher, 1.3 g protein/kg/d, than in other previous studies
which may explain our findings [10, 40]. However, our
findings relating to protein intake require further explora-
tion. Nevertheless, our findings suggest that consuming
Cross-Sectional Associations Between Dietary Antioxidant Vitamins C, E and Carotenoid 34
Intakes…
sufficient antioxidant vitamins in addition to protein is by a different food groups. Given the shortfall in intakes of
important.
Despite the few previous studies, to our knowledge, our
study is the first to investigate dietary intakes of vitamin C,
E, A (retinol), individual carotenoids and protein, as well
as circulating CRP with the full range of sarcopenic indices
in both younger and older women.

Physiological Mechanisms

There is clear mechanistic relevance of the antioxidant

vitamins on skeletal muscle health during aging which
also supports our observational findings. Supplementation
of lycopene, β-carotene or mixed antioxidant vitamins in
animal experiments either had positive effects on muscle
force or physical activity or attenuated oxidative stress or
loss of skeletal muscle [41]. Vitamin E also had protective
effects on exercise-induced oxidative damage in both
young and older adults [7], and vitamin C had protective
effects on oxidative biomarkers, inflammatory cytokines
and CRP [42]. Beyond roles as an antioxidant and anti-
inflammatory agent vitamin C is also integral to synthesis
of collagen and of carnitine which is important for the
metabolism of long- chain fatty acids during physical
activity [8].

Deficiency of Vitamins and Supplementation

Although the significant associations between intake of

vitamins C, E and carotenoids were found across the usual
range of consumption, the women in the lowest quintile of
vitamin C intake consumed less than the average require-
ment for vitamin C intake of 80 mg/d [43]. The prevalence
of low intakes of vitamin C, or circulating concentrations
of vitamin C indicative of scurvy, is high in vulnerable
older populations living in community and residential care
~ 40% [3, 44]. Moreover, consumption of 5 or more
portions of fruits and vegetables a day is only 19% of those
over the age of 75 years in the UK [45].
Whilst supraphysiological doses of vitamin C could be
used to rectify low dietary intakes in older vulnerable
groups these need to be used with caution due to the
detrimental, prooxidant, effects of long-term high doses
of vitamin C (> 500 mg per day) compared the
concentrations found natu- rally in foods [46]. There is also
debate surrounding poten- tial negative effects of
supraphysiological doses of vitamin C on skeletal muscle
function in athletes [47].
Our research has found that intake of antioxidant vita-
mins within the normal dietary range has important effects
on sarcopenic indices, in the whole cohort and stratified
analysis showed associations in younger women, provided

13
34 A. A. Welch et al.
the antioxidant vitamins it is important to encourage Trust (Grant WT081878MA). Medical Research Council, European
greater intakes of the foods that supply them; namely Union, the National Institute for Health Research (NIHR)-funded
BioResource,
vegetables, fruits and whole grain cereal foods, in
addition protein for maintaining skeletal muscle health in
populations of all ages. This is particularly important for
vulnerable older people in the community and in
residential care.
Strengths of this study include comprehensive meas-
urements of diet in a well characterised large cohort
which allowed us to examine for the first time
associations between an extensive range of sarcopenic
indices, including DXA-measured FFM, LEP and grip
strength and a range of antioxidant vitamins, including
all the carotenoids. We were also able to undertake a
comparative analysis of the different nutrients allowing
us to examine the independ- ent associations with intake
of each vitamin and protein. Limitations include the small
number of women aged over 65 an imbalance between
younger and older women in our cohort which future
studies should address. In addition, this study was cross-
sectional which limits inference for causa- tion. We also
did not account for the nutrients supplied by food
supplements, however, our analyses indicate the impor-
tance of intake of vitamins supplied by diet alone. A
further limitation is that we did not have directly
measured physical activity, however, we used validated
questionnaires, which although less precise than objective
measures, do distinguish across the range of activity
levels in individuals [28].
Although we were able to determine what dietary intakes
are relevant for skeletal muscle health we lacked data on
dietary biomarkers meaning we were unable to confirm
our findings with blood or urinary levels.

Conclusion

For the first time in a free-living population, our research

shows that higher dietary intake of the antioxidant
vitamins, particularly vitamin C, could be protective for
both loss of skeletal muscle mass and power during
aging, and have rel- evance for treatment and prevention
of frailty and sarcopenia in women. It was notable that the
observed associations were identified in women younger
than 65 years. Our findings provide further
encouragement for intervention trials as well as for
following the healthy eating guidelines for protection of
skeletal muscle mass and power in people of all ages.

Author Contributions Concept and design of study AAW, AJ, EK.

Statistical analysis AJ. Drafting of manuscript AAW, AJ. All
authors commented on the manuscript and contributed to the study.

Funding This study was funded by PhD funding from the

University of East Anglia. TwinsUK is funded by the Wellcome
13
Cross-Sectional Associations Between Dietary Antioxidant Vitamins C, E and Carotenoid
Intakes…
Clinical Research Facility, and Biomedical Research Centre based
at Guy’s and St Thomas’ NHS Foundation Trust in partnership with
King’s College London.
Compliance with Ethical Standards
Conflict of interest Ailsa Welch, Amy Jennings, Eirini Kelaiditi, Jane
Skinner and Claire Steves declare no conflicts of interest.
Human and Animal Rights and Informed Consent This study was
approved by the St Thomas’s Hospital Research Ethics Committee
and all participants provided written informed consent.
Open Access This article is licensed under a Creative Commons Attri-
bution 4.0 International License, which permits use, sharing, adapta-
tion, distribution and reproduction in any medium or format, as long
as you give appropriate credit to the original author(s) and the source,
provide a link to the Creative Commons licence, and indicate if
changes were made. The images or other third party material in this
article are included in the article’s Creative Commons licence, unless
indicated otherwise in a credit line to the material. If material is not
included in the article’s Creative Commons licence and your intended
use is not permitted by statutory regulation or exceeds the permitted
use, you will need to obtain permission directly from the copyright
holder.To view a copy of this licence, visit
https://creativecommons.org/licenses/by/4.0/.
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Edited by:
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Reviewed by:
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*Correspondence:
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kunisawa@nibiohn.go.jp
Specialty section:
This article was submitted to
Nutrition and Microbes,
a section of the journal
Frontiers in Nutrition
Received: 05 December 2018
Accepted: 01 April 2019
Published: 17 April 2019
Citation:
Yoshii K, Hosomi K, Sawane K and
Kunisawa J (2019) Metabolism of
Dietary and Microbial Vitamin B Family
in the Regulation of Host Immunity.
Front. Nutr. 6:48.
doi: 10.3389/fnut.2019.00048
Frontiers in Nutrition | www.frontiersin.org
REVIEW
published: 17 April 2019
doi: 10.3389/fnut.2019.00048
Metabolism of Dietary and Microbial
Vitamin B Family in the Regulation of
Host Immunity
Ken Yoshii 1,2, Koji Hosomi 1, Kento Sawane 1,3,4 and Jun Kunisawa 1,2,3,5,6,7*
1
Laboratory of Vaccine Materials, Center for Vaccine and Adjuvant Research, and Laboratory of Gut Environmental System,
National Institutes of Biomedical Innovation, Health and Nutrition, Osaka, Japan, 2 Graduate School of Medicine, Osaka
University, Osaka, Japan, 3 Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan, 4 Innovation
Center, Nippon Flour Mills Co., Ltd., Atsugi, Japan, 5 Graduate School of Dentistry, Osaka University, Osaka, Japan,
6
Department of Microbiology and Immunology, Graduate School of Medicine, Kobe University, Hyogo, Japan, 7 Division of
Mucosal Vaccines, International Research and Development Center for Mucosal Vaccines, The Institute of Medical Science,
The University of Tokyo, Tokyo, Japan
Vitamins are micronutrients that have physiological effects on various biological responses,
including host immunity. Therefore, vitamin deficiency leads to increased risk of developing
infectious, allergic, and inflammatory diseases. Since B vitamins are synthesized by plants,
yeasts, and bacteria, but not by mammals, mammals must acquire B vitamins from dietary or
microbial sources, such as the intestinal microbiota. Similarly, some intestinal bacteria are unable
to synthesize B vitamins and must acquire them from the host diet or from other intestinal
bacteria for their growth and survival. This suggests that the composition and function of the
intestinal microbiota may affect host B vitamin usage and, by extension, host immunity.
Here, we review the immunological functions of B vitamins and their metabolism by
intestinal bacteria with respect to the control of host immunity.
Keywords: absorption, gut microbiota, intestinal immunity, nutrition, vitamin
INTRODUCTION
The gut is continuously exposed both to toxic (e.g., pathogenic microorganisms) and beneficial
(e.g., dietary components, commensal bacteria) compounds and microorganisms; therefore, the
intestinal immune system must maintain a healthy balance between active and suppressive immune
responses. This balance is controlled not only by host immune factors such as cytokines but also by a
variety of environmental factors such as dietary components and the composition of the commensal
bacteria. Furthermore, several lines of evidence have demonstrated that immune homeostasis
in the intestine is related not only to intestinal health but also to the health of the whole body
(1– 3). Therefore, immune regulation by environmental factors is attracting attention as a means
of maintaining immunological health and preventing many diseases.
Nutrients are essential for the development, maintenance, and function of the host immune
system (4, 5). Vitamins are essential micronutrients that are synthesized by bacteria, yeasts, and
plants, but not by mammals. Therefore, mammals must obtain vitamins from the diet or rely on
their synthesis by commensal bacteria in the gastrointestinal tract. Some vitamins are water-soluble
(e.g., vitamin B family and vitamin C), whereas others are fat-soluble (e.g., vitamins A, D, E, and
K). Water-soluble vitamins are not stored by the body and any excess is excreted in the urine;
therefore, it is important to consume a diet containing the necessary amounts of these
vitamins. Vitamin
1 April 2019 | Volume 6 | Article 48
Yoshii et al. Vitamin B Family and Host Immunity

deficiency associated with insufficient dietary intake occurs not induced by vitamin B1
only in developing countries but also in developed countries as
a result of increased use of unbalanced diet (6).
In addition to the diet, the commensal bacteria are recognized
as important players in the control of host health (7–9). From
the point of view of vitamins, commensal bacteria are both
providers and consumers of B vitamins and vitamin K.
Although dietary B vitamins are generally absorbed through
the small intestine, bacterial B vitamins are produced and
absorbed mainly through the colon (10, 11), indicating that
dietary and gut microbiota- derived B vitamins are possibly
handled differently by the human body. B vitamins are
important cofactors and coenzymes in several metabolic
pathways, and it has been reported recently that B vitamins
also play important roles in the maintenance of immune
homeostasis (12, 13). Thus, both dietary components and the
gut microbiota modulate host immune function via B
vitamins. Here, we review the metabolism and function of
dietary and gut microbiota-derived B vitamins in the control of
host immunity.

VITAMIN B1
Vitamin B1 (thiamine) is a cofactor for several enzymes,
including pyruvate dehydrogenase and α-ketoglutarate
dehydrogenase, which are both involved in the tricarboxylic
acid (TCA) cycle (14, 15). World Health Organization
(WHO)/Food and Agriculture Organization (FAO) recommend
a daily vitamin B1 intake of 1.1–1.2 mg for adult (16). Vitamin
B1 deficiency causes lethargy and, if left untreated, can
develop into beriberi, a disease that affects the peripheral
nervous system and cardiovascular system. Accumulating
evidence suggests that energy metabolism—particularly the
balance between glycolysis and the TCA cycle—is associated
with the functional control of immune cells, in what is now
referred to as immunometabolism (17).
Immunometabolism is well studied in T cells and
macrophages; quiescent or regulatory-type cells (e.g., naive
T cells, Treg cells, and M2 macrophages) use the TCA cycle for
energy generation, whereas activated or pro-inflammatory
cells (e.g., Th1, Th2, Th17, and M1 macrophages) use
glycolysis (18, 19).
Previously, we examined B cell immunometabolism in the
intestine. In the intestine, naïve immunoglobulin (Ig) M + B cells
differentiate into IgA+ B cells in Peyer’s patches (PPs) by class
switching, and then IgA+ B cells differentiate into IgA-
producing plasma cells in the intestinal lamina propria (20).
Naïve B cells
in PPs preferentially use a vitamin B1-dependent TCA cycle for
the generation of ATP. However, once the B cells differentiate
into IgA-producing plasma cells, they switch to using glycolysis
for the generation of ATP and shift to a catabolic pathway for
the production of IgA antibody (Figure 1). Consistent with the
importance of vitamin B1 in the maintenance of the TCA cycle,
mice fed a vitamin B1-deficient diet show impaired
maintenance of naïve B cells in PPs, with little effect on IgA-
producing plasma cells. Since PPs are the primary sites of
induction of antigen- specific IgA responses, PP regression
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Yoshii et al. Vitamin B Family and Host Immunity

deficiency leads to decreased IgA antibody responses to oral

vaccines (21).
Vitamin B1 is found in high concentrations as thiamine
pyrophosphate (TPP) in meat (particularly pork and chicken);
eggs; cereal sprouts and rice bran; and beans. Dietary TPP
is hydrolyzed by alkaline phosphatase and converted to free
thiamine in the small intestine (22). Free thiamine is absorbed
by the intestinal epithelium via thiamine transporters (e.g.,
THTR-1, THTR-2) and transported to the blood for distribution
throughout the body (11). Free thiamine is converted back to TPP
and is used for energy metabolism in the TCA cycle.
Various types of intestinal bacteria, mostly in the colon,
also produce vitamin B1 as both free thiamine and TPP
(11, 23). In the colon, free bacterial thiamine is absorbed
mainly by thiamine transporters, transported to the blood,
and distributed throughout the body; this mechanism is
similar to how free dietary thiamine is taken up in the small
intestine. However, unlike in the small intestine, TPP produced
by the gut microbiota is not converted to free thiamine,
because alkaline phosphatase is not secreted in the colon
(24). Instead, TPP is absorbed directly by the colon via TPP
transporters (e.g., TPPT-1) that are highly expressed on the
apical membrane of the colon (25). The absorbed TPP
enters the mitochondria via MTPP-1, a TPP transporter that
is expressed in the mitochondrial inner membrane and is used
as a cofactor for ATP generation (26). This suggests that
bacterial TPP is important for energy generation in the colon.
Thus, dietary and bacterial vitamin B1 appears to have
different roles in the host.
The vitamin B1 structure consists of a thiazole moiety
linked to a pyrimidine moiety. Bacteria obtain the thiazole
moiety from glycine or tyrosine and 1-deoxy-D-xylulose-5-
phosphate, and plants and yeasts synthesize it from glycine
and 2-pentulose (27– 30). In both bacteria and plants, the
pyrimidine moiety is derived from 5-aminoimidazole
ribonucleotide, an intermediate in the purine pathway (29).
Metagenomic analyses of the human gut microbiota predict
that Bacteroides fragilis and Prevotella copri (phylum
Bacteroidetes); Clostridium difficile, some Lactobacillus spp.,
and Ruminococcus lactaris (Firmicutes); Bifidobacterium spp.
(Actinobacteria); and Fusobacterium varium are vitamin B1
producers (Table 1) (10, 46), implying that many intestinal
bacteria possess a complete vitamin B1 synthesis pathway, which
includes pathways for the synthesis of thiazole and
pyrimidine. Indeed, Lactobacillus casei produces thiamine
during the production of fermented milk drinks (31), and
Bifidobacterium infantis and B. bifidum produce thiamine in
culture supernatant (32). However, Faecalibacterium spp.
(Firmicutes) lack a vitamin B1 synthesis pathway even though
they require vitamin B1 for their growth (10). Therefore, these
bacteria must obtain their vitamin B1 from other bacteria or
from the host diet via a thiamine transporter, suggesting that
there is competition for vitamin B1 between the host and
certain intestinal bacteria.

VITAMIN B2
Vitamin B2 (riboflavin) and its active forms (flavin adenine
dinucleotide [FAD] and flavin mononucleotide [FMN]) are

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Yoshii et al.
cofactors for enzymatic reactions in the TCA cycle and in fatty
acid oxidization (also known as β-oxidization) (15). WHO/FAO
recommends a daily
FIGURE 1 | Vitamin vitamin
B1 and B2 intake
B2-mediated of 1.0–1.3inmg
immunometabolism fordifferentiation
B cell
(16). Vitamin B2 deficiency suppresses the activity of acyl-
CoA dehydrogenases involved in the oxidation of fatty acids to
generate acetyl-CoA, which is used by mitochondria to produce
ATP via the TCA cycle. Fatty acid oxidization is involved
in the activation, differentiation, and proliferation of immune
cells through the generation of acetyl-CoA and its entry into
TCA cycle (47). This suggests that vitamin B2 is associated
with the control of differentiation and function of immune
cells through regulation of fatty acid oxidization (Figure 1);
however, the immunological roles of vitamin B2 in the control
of host immunity remain to be investigated. In addition to
energy generation, vitamin B2 is associated with reactive
oxygen species (ROS) generation in immune cells through the
priming of NADPH oxidase 2 (48); ROS are important effector
and signaling molecules in inflammation and immunity.
Vitamin B2 is found at high levels in leafy green vegetables,
liver, and eggs. Dietary vitamin B2 exists as FAD or FMN and is
converted to free riboflavin by FAD pyrophosphatase and FMN
phosphatase in the small intestine (49, 50). Free riboflavin is
absorbed via riboflavin transporter expressed on the
epithelium of the small intestine and is then released into the
blood. In the blood, free riboflavin is converted back to FAD or
FMN and distributed throughout the body (51–53).
Bacterial vitamin B2 is synthesized from guanosine
triphosphate (GTP) and D-ribulose 5-phosphate (54). Bacterial
vitamin B2 exists as free riboflavin, which is directly absorbed
in the large intestine, converted to FAD or FMN, and
distributed throughout the body as described above (23). A
metagenome analysis of the human gut microbiota by
Magnúsdóttir et al.
(10) has predicted that Bacteroides fragilis and Prevotella copri
(Bacteroidetes); Clostridium difficile, Lactobacillus plantarum,
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Vitamin B Family and Host Immunity
L. fermentum, and Ruminococcus lactaris (Firmicutes) express
factors essential for vitamin B2 synthesis, suggesting that these
adults inbacteria
the intestine.are an B1
Vitamin important source
acts as a cofactor of vitamin
for enzymes such as B2 in the
pyruvate large and α-ketogl
dehydrogenase
intestine (Table 1). In contrast, Bifidobacterium spp., and
Collinsella spp. (Actinobacteria) lack a vitamin B2 pathway.
Supplementation of fermented soymilk containing
Lactobacillus plantarum with riboflavin deficient diet has been
shown to promote vitamin B2 production and prevent vitamin
B2 deficiency in mice (35). L. fermentum isolated from
sourdough can synthesize riboflavin in vitro (36). Furthermore,
recent evidence indicates that some species in Bacteroidetes
phylum produce more riboflavin than do Actinobacteria and
Firmicutes phyla (55). However, Actinobacteria and
Firmicutes phyla still express riboflavin transporter and the
enzymes necessary for FAD and FMN generation from free
riboflavin (i.e., FAD synthases and flavin kinases) (10, 56),
suggesting that all bacteria, including those that are unable to
synthesize vitamin B2 themselves, require FAD and FMN for
their growth and survival. Thus, as with vitamin B1, there is
likely competition for riboflavin between the host and the
commensal bacteria.
In addition to being able to produce vitamin B2, some
bacteria (e.g., commensals such as Lactobacillus acidophilus
and pathogens such as Mycobacterium tuberculosis and
Salmonella typhimurium) produce the vitamin B2 intermediate
(57–59), 6- hydroxymethyl-8-D-ribityllumazine (60, 61). 6-
Hydroxymethyl- 8-D-ribityllumazine binds to major
histocompatibility complex class I-related gene protein (MR1)
on antigen-presenting cells; this causes mucosal-associated
invariant T (MAIT) cells, an abundant population of innate-
like T cells, to produce cytokines such as interferon gamma
and interleukin (IL) 17, which contribute to host defense
against pathogens (Figure 2) (62). It is thought that
stimulation by commensal bacteria contributes to the
development and activation of MAIT cells for immunological
surveillance against pathogens. MAIT cells
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Yoshii et al.
TABLE 1 | Vitamin B family producing bacteria.
Vitamins Forms Bacteria
B1 Thiamin Bacteroides fragilis
pyrophosphate Prevotella copri
(TPP) Clostridium difficile
Lactobacillus casei
Lactobacillus curvatus
Lactobacillus plantarum
Ruminococcus lactaris
Bifidobacterium infantis
Bifidobacterium bifidum
Fusobacterium varium
B2 Flavin adenine Bacteroides fragilis
dinucleotide (FAD) Prevotella copri
Flavin
Clostridium difficile
mononucleotide Lactobacillus plantarum
(FMN) Lactobacillus fermentum
Ruminococcus lactaris
B3 Nicotinic acid Bacteroides fragilis
Nicotinamide Prevotella copri
Ruminococcus lactaris
Clostridium difficile
Bifidobacterium infantis
Helicobacter pylori
Fusobacterium varium
B5 Free pantothenic
Bacteroides fragilis
acid Prevotella copri
Ruminococcus lactaris
Ruminococcus torques
Salmonella enterica
Helicobacter pylori
B6 Pyridoxal
Bacteroides fragilis
phosphate (PLP) Prevotella copri
Bifidobacterium longum
Collinsella aerofaciens
Helicobacter pylori
B7 Free biotin Bacteroides fragilis
Lactobacillus helveticus
Fusobacterium varium
Campylobacter coli
B9 Tetrahydrofolate Bacteroides fragilis
(THF) Prevotella copri
Clostridium difficile
Lactobacillus plantarum
Lactobacillus delbrueckii
ssp. bulgaricus
Lactobacillus reuteri
Streptococcus thermophilus
Bifidobacterium
pseudocatenulatum
Bifidobacterium
adolescentis
Fusobacterium varium
Salmonella enterica
B12 Adenosylcobalamin Bacteroides fragilis
Prevotella copri
Clostridium difficile
Faecalibacterium prausnitzii
Ruminococcus lactaris
Propionibacterium
freudenreichii
Lactobacillus plantarum
Lactobacillus coryniformis
Lactobacillus s reuteri
Bifidobacterium animalis
Bifidobacterium infantis
Bifidobacterium longum
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Vitamin B Family and Host Immunity
also produce inflammatory cytokines and have tissue-homing
References properties, suggesting that these cells are also involved in the
development of autoimmune and inflammatory diseases (63).
(10, 31–33)
VITAMIN B3
Vitamin B3 (niacin) is generally known as nicotinic acid and
nicotinamide. These compounds are precursors of
nicotinamide adenine dinucleotide (NAD), a coenzyme in the
cellular redox reaction with a central role in aerobic
respiration. WHO/FAO recommends a daily vitamin B3 intake
(10, 34–36) of 11–12 mg for adults (16).
Vitamin B3 is also a ligand of GPR109a, a G-protein coupled
receptor expressed on several types of cells, including immune
cells (64). Vitamin B3–GPR109a signaling induces
differentiation of regulatory T cells and suppresses colitis in a
(10, 32) GPR109a- dependent manner (65). Vitamin B3 also inhibits the
production of the pro-inflammatory cytokines IL-1, IL-6, and
tumor necrosis factor alpha (TNF-α) by macrophages and
monocytes (Figure 3) (66). Thus, vitamin B3 has anti-
inflammatory properties by modulating host immune cells
and playing an important role in the maintenance of
(10)
immunological homeostasis. Indeed, in humans, vitamin B3
deficiency causes pellagra, which is a disease characterized by
intestinal inflammation, diarrhea, dermatitis, and dementia
(67).
Unlike the other B vitamins, vitamin B3 can be generated
(10, 32)
by mammals via an endogenous enzymatic pathway from
tryptophan and is stored in the liver, although it is also
obtained from the diet (68). Animal-based foods such as fish
and meat contain vitamin B3 as nicotinamide, and plant-based
foods such as beans contain vitamin B3 as nicotinic acid. Both
nicotinamide
(10, 37) and nicotinic acid are directly absorbed through the small
intestine, where nicotinic acid is converted to nicotinamide.
Vitamin B3 is also synthesized from tryptophan by intestinal
(10, 38–41) bacteria (69, 70). Bacteroides fragilis and Prevotella copri
(Bacteroidetes); Ruminococcus lactaris, Clostridium difficile
(Firmicutes); Bifidobacterium infantis (Actinobacteria);
Helicobacter pylori (Proteobacteria); and Fusobacterium varium
(Fusobacteria) possess a vitamin B3 biosynthesis pathway
(Table 1) (10, 71). Thus, many intestinal bacteria from various
genera can produce vitamin B3, suggesting that both dietary
and commensal bacteria-derived vitamin B3 are important for
host immunity.
VITAMIN B5
(10, 32, 33,
Fusobacterium
42–45)
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Yoshii et al. Vitamin B Family and Host Immunity

Vitamin B5 (pantothenic acid) is a precursor of

coenzyme A (CoA), which is an essential
cofactor for the TCA cycle and fatty acid
oxidation (72). WHO/FAO recommends a daily
vitamin B5 intake of 5.0 mg for adults (16). Like
vitamins B1 and B2, vitamin B5 is involved in
the control of host immunity via energy
generation by immune cells. Vitamin B5 deficiency
causes immune diseases such as dermatitis, as
well as non-immune- related conditions such as
fatigue and insomnia (73). In a randomized,
double-blind, placebo-controlled study in
adults, dietary supplementation with vitamin
B5 was shown to improve facial acne (74),
suggesting that epithelial barrier function

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Yoshii et al. Vitamin B Family and Host Immunity

FIGURE 2 | Regulation of MAIT cells by microbial metabolites derived from vitamin B2 and B9. Commensal bacteria/pathogens produce the vitamin B2 metabolite 6-hydroxymethyl-8-D-rib
pathogens and conversely are associated with inflammation. In contrast, the vitamin B9 metabolite acetyl-6-formylpterin binds as an antagonist to MR1, thus inhibiting
the activation of MAIT cells.

FIGURE 3 | Pivotal roles of vitamins B3, B7, and B9 in maintenance of immunological homeostasis. Vitamin B3 binds to GPR109a in dendritic cells and macrophages, and GPR109a signaling lead

improves via the promotion of keratinocyte proliferation and differentiation into fibroblasts (75). To maintain vitamin B5
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Yoshii et al. Vitamin B Family and Host Immunity

levels during times of deficiency, CoA is converted back cysteamine inhibits peroxisome proliferator-activated receptor
to vitamin B5 or cysteamine via pantetheine (76). However, gamma (PPARγ) signaling, causing inflammation (77). Indeed,
colitis has been improved in pantetheine-producing-enzyme
knockout mice (78). Thus, vitamin B5 deficiency causes

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Yoshii et al.
inflammation through both dysfunction of the epithelial barrier
and the production of pro-inflammatory molecules.
In terms of immune responses, vitamin B5 enhances
protective activity against Mycobacterium tuberculosis infection
by promoting innate immunity and adaptive immunity. In mice,
vitamin B5 supplementation activates phagocytosis and
cytokine production (including IL-6 and TNF-α) by macrophages
and subsequently promotes Th1 and Th17 responses for the
clearance of M. tuberculosis from the lungs (79). Thus, vitamin
B5 contributes to host defense by activating immune
responses, suggesting that this vitamin has an important role
as a natural adjuvant.
Vitamin B5 is found in high concentrations as CoA or
phosphopantetheine in liver, eggs, chicken, and fermented
soybeans. CoA and phosphopantetheine are converted to free
pantothenic acid by endogenous enzymes such as phosphatase
and pantetheinase in the small intestine. Free pantothenic acid
is absorbed via sodium-dependent multivitamin transporter
(SMVT) expressed on the epithelium of the small intestine and
is then released into the blood (80). Finally, free pantothenic acid
is converted back to CoA and distributed throughout the body,
particularly to the liver and kidney.
Bacterial vitamin B5 is synthesized from 2-dihydropantoate
and β-alanine via de novo synthesis pathways (10). Bacterial
vitamin B5 exists as free pantothenic acid, which is directly
absorbed in the large intestine, converted to CoA, and
distributed in the same way as dietary vitamin B5. According
to a genomic analysis, Bacteroides fragilis and Prevotella copri
(Bacteroidetes); some Ruminococcus spp. (R. lactaris and R.
torques) (Firmicutes); Salmonella enterica and Helicobacter pylori
(Proteobacteria) possess a vitamin B5 biosynthesis pathway,
indicating that intestinal commensal bacteria can produce
vitamin B5. In contrast, most Fusobacterium (Fusobacteria)
and Bifidobacterium spp. (Actinobacteria) and some strains of
Clostridium difficile, Faecalibacterium spp., and Lactobacillus spp.
(Firmicutes) lack such a pathway (Table 1), although some of
them do express pantothenic acid transporter to utilize vitamin
B5 for energy generation (10), suggesting that these bacteria
compete with the host for vitamin B5.
VITAMIN B6
Vitamin B6 exists in several forms, including as pyridoxine,
pyridoxal, and pyridoxamine. These forms of vitamin B6 are
precursors of the coenzymes pyridoxal phosphate (PLP) and
pyridoxamine phosphate (PMP), which are involved in a variety
of cellular metabolic processes, including amino acid, lipid, and
carbohydrate metabolism (81). WHO/FAO recommends a daily
vitamin B6 intake of 1.3–1.7 mg for adults (16). Vitamin B6
deficiency is associated with the development of inflammatory
diseases such as allergy and rheumatoid arthritis, as well as
with neuronal dysfunction (82–84). Vitamin B6 deficiency
disrupts the Th1–Th2 balance toward an excessive Th2
response, resulting in allergy (85). Moreover, low plasma
vitamin B6 levels, together with increased levels of pro-
inflammatory cytokines such as TNF-α and IL-6, have been
observed in patients with rheumatoid
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Vitamin B Family and Host Immunity
arthritis (86). However, the mechanism underlying the regulation
of inflammation by vitamin B6 is currently unknown. Vitamin
B6 contributes to intestinal immune regulation through the
metabolism of the lipid mediator sphingosine 1-phosphate (S1P).
S1P regulates lymphocyte trafficking into the intestines, especially
in the large intestine. Lymphocyte trafficking is dependent on S1P
gradient which is created by S1P production and its
degradation. S1P degradation is mediated by S1P lyase that
requires vitamin B6 as a cofactor. The administration of vitamin
B6 antagonist impairs S1P lyase activity and creates an
inappropriate S1P gradient, resulted in impairing lymphocyte
migration from lymphoid tissues and reducing the numbers
of lymphocytes in the intestines (87). The lymphocytes located
between gut epithelial cells are known as intraepithelial cells
(IELs) which are involved in the protection against pathogens
(88). Therefore, vitamin B6 is important role for
immunosurveillance in the intestines.
Vitamin B6 is abundant in fish, chicken, tofu, sweet potato,
and avocado. Dietary vitamin B6 exists as PLP or PMP; it
is converted to free vitamin B6 by endogenous enzymes such
as pyridoxal phosphatase and is then absorbed by the small
intestine. Although absorption of vitamin B6 through acidic pH-
dependent and carrier-mediated transport has been shown, an
intestinal pyridoxine transporter is yet to be identified in any
mammalian species (11). After the absorption of free vitamin
B6, it enters the blood and is converted back to PLP or PMP.
Microbial vitamin B6 is synthesized as PLP from
deoxyxylulose 5-phosphate and 4-phosphohydroxy-L-threonine
or from glyceraldehyde-3-phosphate and D-ribulose 5-phosphate
(10). In the large intestine, bacteria-derived PLP is converted
to free vitamin B6, which is absorbed by passive transport,
transported to the blood, and distributed throughout the body.
Metagenomic analysis has shown that Bacteroides fragilis
and Prevotella copri (Bacteroidetes), Bifidobacterium longum
and, Collinsella aerofaciens (Actinobacteria), and Helicobacter
pylori (Proteobacteria) possess a vitamin B6 biosynthesis
pathway. Bacteroidetes and Proteobacteria likely produce
vitamin B6 starting from deoxyxylulose 5-phosphate and 4-
phosphohydroxy-L-threonine, whereas Actinobacteria likely
start from glyceraldehyde-3-phosphate and D-ribulose 5-
phosphate. In contrast, most Firmicutes genera (Veillonella,
Ruminococcus, Faecalibacterium, and Lactobacillus spp.),
except for some Clostridium (C. bartlettii, C. leptum, C.
methylpentosum, and C. sporogenes) and Lactobacillus spp. (L.
brevis and L. ruminis) lack a vitamin B6 biosynthesis pathway
(10) (Table 1).
VITAMIN B7
Vitamin B7 (biotin) is a cofactor for several carboxylases that
are essential for glucose, amino acid, and fatty acid metabolism
(89). For example, vitamin B7 is an essential cofactor for acetyl-
CoA carboxylase and fatty acid synthase, which are enzymes
involved in fatty acid biosynthesis (90, 91). Thus, vitamin B7
likely influences immunometabolism. WHO/FAO recommends
a daily vitamin B7 intake of 30 µg for adults (16). Vitamin
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Yoshii et al.
B7 suppresses gene expression by binding to (biotinylating)
histones; these genes include that encoding NF-κB, which is
a major signaling molecule for the production of several pro-
inflammatory cytokines (e.g., tumor necrosis factor alpha, IL-
1, IL-6, IL-8) (92, 93). Nuclear transcription of NF-κB is
activated in response to vitamin B7 deficiency (94), suggesting
that biotinylation of histones suppresses the expression of
genes encoding pro-inflammatory cytokines in NF-κB signaling
(Figure 3). Therefore, vitamin B7 has anti-inflammatory effects
by inhibiting NF-κB activation, and dietary vitamin B7
deficiency causes inflammatory responses via enhanced
secretion of pro- inflammatory cytokines (95, 96).
Vitamin B7 is abundant in foods such as nuts, beans, and
oilseed. However, raw egg-white contains a large amount of
avidin, which binds strongly to vitamin B7 and prevents its
absorption in the gut (97). Therefore, vitamin B7 deficiency can
be caused not only by insufficient vitamin B7 intake, but also by
excessive intake of raw egg-white. Dietary biotin exists as a
free protein-bound form or as biocytin (11). In the small
intestine, biotinidase releases free biotin from the bound
protein and the free biotin is absorbed via the biotin
transporter SMVT (98).
Vitamin B7 is also produced by intestinal bacteria as
free biotin synthesized from malonyl CoA or pimelate via
pimeloyl-CoA (99, 100). Bacterial free biotin is absorbed
by SMVT expressed in the colon (23, 101). Metagenomic
analysis has shown that Bacteroides fragilis and Prevotella
copri (Bacteroidetes); Fusobacterium varium (Fusobacteria)
and Campylobacter coli (Proteobacteria) possess a vitamin
B7 biosynthesis pathway (10). In contrast, Prevotella spp.
(Bacteroidetes), Bifidobacterium spp. (Actinobacteria), and
Clostridium, Ruminococcus, Faecalibacterium, and Lactobacillus
spp. (Firmicutes) lack such a pathway (Table 1); however, they
do express free biotin transporter (10, 102), suggesting that
these bacteria also utilize dietary and bacterial vitamin B7 and
therefore may compete with the host. Thus, free biotin may
influence the composition of the gut microbiota, because biotin is
necessary for the growth and survival of the microbiota.
Indeed, biotin deficiency leads to gut dysbiosis and the
overgrowth of Lactobacillus murinus, leading to the
development of alopecia (103). Furthermore, vitamin B7
production appears to proceed in a cooperative manner among
different intestinal bacteria; Bifidobacterium longum in the
intestine produces pimelate, which is a precursor of vitamin B7
that enhances vitamin B7 production by other intestinal
bacteria (104).
VITAMIN B9
Vitamin B9 (folate), in its active form as tetrahydrofolate,
is a cofactor in several metabolic reactions, including DNA
and amino acid synthesis. WHO/FAO recommends a daily
vitamin B9 intake of 400 µg for adults (16). Owing to the
high requirement of vitamin B9 by red blood cells, vitamin
B9 deficiency leads to megaloblastic anemia (23). Vitamin B9
deficiency also inhibits the proliferation of human CD8 + T cells
in vitro by arresting the cell cycle in the S phase and increasing
the frequency of DNA damage (105). Moreover, vitamin B9
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Vitamin B Family and Host Immunity
contributes to the maintenance of immunologic homeostasis.
Regulatory T cells (Treg) express high levels of vitamin B9
receptor (folate receptor 4 [FR4]). Administration of anti-FR4
antibody results in specific reduction in the Treg cell population
(106), suggesting that the vitamin B9–FR4 axis is required for
Treg cell maintenance. In vitro culture of Treg cells under
vitamin B9-reduced conditions leads to impaired cell survival,
with decreased expression of anti-apoptotic Bcl2 molecules,
although naïve T cells retain the ability to differentiate into
Treg cells; this suggests that vitamin B9 is a survival factor for
Treg cells (87). Consistent with these findings, deficiency of
dietary vitamin B9 results in reduction of the Treg cell
population in the small intestine (107, 108). Since Treg cells
play an important role in the prevention of excessive immune
responses (109), mice fed a vitamin B9-deficient diet exhibit
increased susceptibility to intestinal inflammation (107).
Foods such as beef liver, green leafy vegetables, and asparagus
contain high levels of vitamin B9. Vitamin B9 exists as both
mono- and polyglutamate folate species in the diet (110).
Folate polyglutamate is deconjugated to the monoglutamate
form and then absorbed in the small intestine via folate
transporters such as proton-coupled folate transporter (PCFT)
(11, 111). In the intestinal epithelium, folate monoglutamate is
converted to tetrahydrofolate (THF), an active form and cofactor,
before being transported to the blood (111).
Intestinal bacteria synthesize vitamin B9 as THF from GTP,
erythrose 4-phosphate, and phosphoenolpyruvate (38, 112).
Bacterial THF is directly absorbed in the colon via PCFT and
distributed through the body by the blood (113). Metagenomic
analysis has shown that Bacteroides fragilis and Prevotella
copri (Bacteroidetes); Clostridium difficile, Lactobacillus
plantarum,
L. reuteri, L. delbrueckii ssp. bulgaricus, and Streptococcus
thermophilus (Firmicutes), some species in Bifidobacterium spp
(Actinobacteria); Fusobacterium varium (Fusobacteria) and
Salmonella enterica (Proteobacteria) possess a folate biosynthesis
pathway (Table 1) (10, 40). This suggests that almost all species
in all phyla produce folate. Indeed, dietary supplementation
with Bifidobacterium probiotic strains (B. adolescentis and
B. pseudocatenulatum) enhances folate production in the
large intestine of folate-deficient rats and folate-free culture
medium (38, 41, 114). Furthermore, Lactobacillus plantarum,
L. delbrueckii ssp. bulgaricus, and L. reuteri enhance folate
production in bacterial culture supernatant lacking the
components needed for folate synthesis (38, 39, 115).
In commensal bacteria, a vitamin B9 metabolite, 6-
formylpterin (6-FP), is produced by photodegradation
of folic acid (116). Like the vitamin B2 metabolite 6-
hydroxymethyl-8-D-ribityllumazine, 6-FP binds to MR1,
but unlike 6-hydroxymethyl-8-D-ribityllumazine it cannot
activate MAIT cells (62, 117). An analog of 6-FP, acetyl-6-FP,
is an antagonist of MR1, which inhibits MAIT cell activation
(118). As mentioned in the section on vitamin B2, 6-
hydroxymethyl-8- D-ribityllumazine activates MAIT cells, which
provide defense against pathogens, so vitamin B9 metabolites
may suppress excess MAIT cell responses and prevent
excessive allergic and inflammatory responses (Figure 2). The
quantitative balance between dietary vitamin B2 and B9 and
the composition of
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Yoshii et al.
the microbiota and its ability to metabolize these vitamins may
be keys to understanding MAIT-cell-mediated homeostasis in
the intestine.
FIGURE 4 | Schematic representation of B-vitamin-mediated interaction between commensal bacteria andduring
VITAMIN B12
Vitamin B12 (cobalamin) is a cobalt-containing vitamin that,
in its active forms of methylcobalamin and adenosylcobalamin,
catalyzes methionine synthesis (119). WHO/FAO recommends
a daily vitamin B12 intake of 2.4 µg for adults (16). Together
with vitamin B6 and B9, vitamin B12 plays important roles
in red blood cell formation and nucleic acid synthesis,
especially in neurons. Therefore, vitamin B12 deficiency causes
megaloblastic anemia and nervous system symptoms such
as numbness of the hands and feet (119). In terms of
host immunity, dietary vitamin B12 deficiency decreases the
number of CD8+ T cells and suppresses natural killer T-
cell activity in mice; supplementation with methylcobalamin
improves these conditions (120), suggesting that vitamin B12
contributes to the immune response via CD8+ T cells and
natural killer T cells.
Beef liver, bivalves, fish, chicken, and eggs contain high
levels of vitamin B12. Dietary vitamin B12 exists in complex
with dietary protein and is decomposed to free vitamin B12 by
pepsin in the stomach. Free vitamin B12 is absorbed by the
epithelial cells of the small intestine via intrinsic factor (IF), a
gastric glycoprotein. Inside the epithelial cells, IF-vitamin B12
complex is decomposed to free vitamin B12 by lysosome and then
released into the blood, where it is converted to the active
form and distributed throughout the body (121, 122).
Bacterial vitamin B12 is synthesized from precorrin-
2 to produce adenosylcobalamin (10), which is absorbed
directly by the large intestine and distributed throughout the
body; the mechanism underlying this absorption is currently
unclear. Metagenomic analysis has predicted that Bacteroides
fragilis and Prevotella copri (Bacteroidetes); Clostridium
difficile, Faecalibacterium prausnitzii and Ruminococcus
lactaris (Firmicutes); Bifidobacterium animalis, B.infantis,
and B.longum (Actinobacteria); Fusobacterium varium
(Fusobacteria) possess a vitamin B12 biosynthesis pathway
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Vitamin B Family and Host Immunity
(Table 1) (10, 32, 42, 43, 45). Indeed, Lactobacillus plantarum
and L. coryniformis isolated from fermented food produce
vitamin B12 (33), and Bifidobacterium animalis synthesizes
host immunity.
vitamin B12 milk fermentation (123).
CONCLUSION
B-vitamin-mediated immunological regulation is specific
to different immune cells and immune responses: that is,
different B vitamins are required for different immune
responses (Figure 4). It was once thought that B vitamins were
obtained only from the diet; however, we know now that this is
not the case and that the intestinal microbiota is also an
important source of vitamins. Within the intestinal microbiota,
not all bacteria produce B vitamins and some bacteria utilize
dietary B vitamins or B vitamins produced by other intestinal
bacteria for their own needs; therefore, there may be
competition between the host and the intestinal microbiota for
B vitamins (Figure 4). Research in this field is complicated,
because not only does the composition of the intestinal
microbiota vary among individuals, but also the composition of
the diet can alter both the composition and function of the
intestinal microbiota. Therefore, vitamin-mediated
immunological maintenance also varies among individuals.
Further examinations in this field are needed, and the new
information uncovered will help to develop a new era of
precision health and nutrition.
AUTHOR CONTRIBUTIONS
KY and KH wrote the draft of review article which was
corrected by JK. KY, KH, and KS drew figures and JK performed
correction.
ACKNOWLEDGMENTS
This review article contains results obtained from our studies
that were supported at least in part by grants from the
Japan Agency for Medical Research and Development
[AMED; 17fk0108223h0002 (JK), 17ek0410032s0102
(JK), 17fk0108207h0002 (JK), 17ek0210078h0002 (JK),
1 April 2019 | Volume 6 | Article 48
0
Yoshii et al.
17ak0101068h0001 (JK), 17gm1010006s0101 (JK), and
19ek0410062h0001 (JK)]; Cross-ministerial Strategic Innovation
Promotion Program; the Ministry of Health, Labor, and
Welfare of Japan; the Science and Technology Research
Promotion Program for Agriculture, Forestry, Fisheries, and
Food Industry; the Ministry of Education, Culture, Sports,
Science, and Technology of Japan; the Japan Society for the
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Conflict of Interest Statement: KS was employed by Nippon Flour Mills Co., Ltd.
The remaining authors declare that the research was conducted in the absence
of any commercial or financial relationships that could be construed as a
potential conflict of interest.
Copyright © 2019 Yoshii, Hosomi, Sawane and Kunisawa. This is an open-access
article distributed under the terms of the Creative Commons Attribution License (CC
BY). The use, distribution or reproduction in other forums is permitted, provided
the original author(s) and the copyright owner(s) are credited and that the
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practice. No use, distribution or reproduction is permitted which does not comply
with these terms.
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