Essentiality of Manganese for Bone Health: An Overview and Update

Abstract

The evidence regarding a deficiency of manganese (Mn) in humans is scarce. So the aim of this narrative review was to consider the state of the art on the relation between manganese and bone health in humans and the effectiveness of manganese supplementation (alone or with other micronutrients) on bone mineralization. This review included 4 eligible studies. All the literature published is in agreement in showing that osteoporotic women have lower serum Mn levels than women with normal bone mineral density, thus confirming the essential role of manganese in the synthesis of cartilage and bone collagen, as well as in bone mineralization and confirming the studies on the animal model. Considering the human studies that evaluated the effectiveness of an oral Mn supplement for a long period (2 years) on the bone mineral density of menopausal women, both of the clinical trials showed that bone loss was significantly greater in the placebo group than in the group taking supplementation, equal to 5.0 mg Mn/day in the study by Strause, and equal to 2.5 mg Mn/day in the study by Saltman, considering, however, that supplementation was represented by a set of microelements (Mn, copper, and zinc) and by calcium.

Full text

1IRCCS Mondino Foundation, Pavia, Italy
2Department of Public Health, Experimental and Forensic Medicine, Unit of
Human and Clinical Nutrition, University of Pavia, Italy
3Endocrinology and Nutrition Unit, Azienda di Servizi alla Persona ‘‘Istituto
Santa Margherita’’, University of Pavia, Italy
4General Management, Azienda di Servizi alla Persona ‘‘Istituto Santa
Margherita’’, Pavia, Italy
5Department of Biology, College of Science, University of Bahrain, Sakhir,
Bahrain
6Research and Development Unit, Indena SpA, Milan, Italy
Corresponding Author:
Gabriella Peroni, Endocrinology and Nutrition Unit, Azienda di Servizi alla
Persona ‘‘Istituto Santa Margherita’’, University of Pavia, Pavia 27100, Italy.
Email: gabriella. peroni01@ universitadipavia. it
Review
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Essentiality of Manganese for Bone Health:
An Overview andUpdate
MariangelaRondanelli1,2, Milena AnnaFaliva3, GabriellaPeroni3 ,
VittoriaInfantino2, ClaraGasparri3, GiancarloIannello4, SimonePerna5, AntonellaRiva6,
GiovannaPetrangolini6, and AliceTartara3
Abstract
The evidence regarding a deficiency of manganese (Mn) in humans is scarce. So the aim of this narrative review was to consider
the state of the art on the relation between manganese and bone health in humans and the effectiveness of manganese supplemen-
tation (alone or with other micronutrients) on bone mineralization. This review included 4 eligible studies. All the literature pub-
lished is in agreement in showing that osteoporotic women have lower serum Mn levels than women with normal bone mineral
density, thus confirming the essential role of manganese in the synthesis of cartilage and bone collagen, as well as in bone mineral-
ization and confirming the studies on the animal model. Considering the human studies that evaluated the effectiveness of an oral
Mn supplement for a long period (2 years) on the bone mineral density of menopausal women, both of the clinical trials showed
that bone loss was significantly greater in the placebo group than in the group taking supplementation, equal to 5.0 mg Mn/day in
the study by Strause, and equal to 2.5 mg Mn/day in the study by Saltman, considering, however, that supplementation was repre-
sented by a set of microelements (Mn, copper, and zinc) and by calcium.
Keywords
manganese, bone, dietary supplementation, bone mineral density, bioactivity
Received: February 15th, 2021; Accepted: April 16th, 2021.
Manganese (Mn) is a metal element present both naturally and
as a consequence of contamination of the soil, sediments and
water; it can exist in different oxidation states, of which Mn2 +
and Mn3 + are the most important from a biological point of
view.1 Food represents the main source of Mn exposure for the
general population; its level in drinking water varies from 0.001
to 0.1 mg/L, while daily intake through the ambient air is sig-
nificantly lower. Mn has proven essential for various species; it
is a component of arginase, pyruvate carboxylase and superox-
ide dismutase, plays the role of cofactor of various enzyme
systems and activates others (glycosyl transferases, for exam-
ple, are specifically activated by Mn).1,2 Due to its poor solubil-
ity, only a small percentage of food Mn is absorbed; this
element is largely excreted very quickly in the intestine via bile
and minimally with urine. It tends to accumulate in tissues rich
in mitochondria, such as the liver and pancreas, and in the
brain, in particular in the globus pallidus, striatum and substan-
tia nigra.1,3
Mn deficiency in animals can lead to several adverse effects,
including growth alterations, skeletal abnormalities, reproduc-
tive defects, ataxia, and lipid and carbohydrate metabolism
defects. On the contrary, the evidence about a deficiency of
Mn in humans is scarce and no specific manganese deficiency
syndrome has been described so far (effects such as hair color
change from black to red, slow nail growth and scaly dermatitis
have been observed only in experimental conditions).1,2
Studies on Animal and in Vitro Models
According to data in the literature, bone represents one of the
main deposition tissues of Mn (containing about 43% of total

Natural Product Communications
2
body Mn).4 Mn, in fact, seems to be an important osteotropic
element: In addition to stimulating the synthesis of the bone
matrix, it seems to have an effect on calcification in general.5 In
a 1986 study, 150 female rats were divided into 3 groups
according to the quantities of Mn and/or copper (Cu) possibly
added to their diet: (1) group N, normal intake of Mn (66 ppm)
and Cu (5 ppm); (2) group L, low intake of Mn (2.5 ppm) and
Cu (0.5 ppm); (3) group D, with no addition of Mn and normal
supply of Cu (5 ppm). After a period of 12 months, the rats
belonging to group D (without Mn intake) showed significantly
higher serum calcium and phosphorus levels than the rats of
group N. On the other hand, calcium and Mn concentrations
were found to be significantly lower in group D than in group
N, with an inverse correlation between femoral concentration
and serum calcium concentration in rats in groups L and D.
These observations could be the consequence of alterations in
the regulation of calcium at bone level (decreased mineraliza-
tion) associated with increased bone resorption, in turn caused
by long- term manganese and copper deficiencies.6
In order to investigate the possible relationship between Mn
deficiency and bone development, Liu and collaborators
administered to 3 groups of chicks the same basal diet, supple-
mented with 60 (control group), 40 (Mn I deficient group) and
8.7 mg Mn/kg of food (group deficient in Mn II), respectively.
Compared to the control group, the Mn deficient groups
showed, in a dose- dependent manner, a markedly less trabecu-
lar reticulum at the tibial level, with a reduction in the number
and thickness of the trabeculae. At an ultrastructural level, the
presence of damaged osteoblasts (rupture of the nuclear and
external mitochondrial membranes, loss of the mitochondrial
crests, and alterations in the endoplasmic reticulum) was
observed in both groups with low Mn intake, with a higher
incidence, in particular, in group II. Furthermore, in these 2
groups (and to a greater extent in II), a reduced level of expres-
sion of the OPG (osteoprotegerin) and RANKL (receptor
activator of nuclear factor kB ligand) genes was found with an
increase in the RANKL/OPG mRNA ratio, an index of an
alteration in the dynamic balance between resorption and bone
formation in favor of the former. Overall, therefore, Mn defi-
ciency could interfere with normal bone development by inhib-
iting the viability of osteoblasts and altering the RANKL/
OPG mRNA ratio.7 The same authors, in a second study car-
ried out on chicks, divided in a similar way to the previous one,
in 3 groups based on the Mn content of their diet, observed a
significant reduction in the size of the proliferative zone and an
increase in the rate of apoptosis of chondrocytes at the level
of the tibial growth cartilage in animals deficient in Mn. The
deficit of this element would therefore be related to disorders
in bone development and growth.8
A study conducted in 1998 on young male rats on a poor
Mn diet (- Mn: 0.5 µg/g of food) for 3 months had already
shown a reduction in the growth rate, with a significant decrease
in the body weight of these animals, compared to the control
group (+Mn: rats fed a diet containing 45 µg Mn/g of food).
The deficiency status induced by the poor Mn diet (confirmed
by the finding of a 15- fold reduction in the concentration of
Mn in the liver) was associated with an alteration in the metab-
olism of GH and IGF-1: in -Mn rats, a decrease in circulating
concentrations of IGF-1 and insulin and an increase in GH
levels compared to controls had been observed, suggesting
how these modifications could, at least in part, be responsible
for the growth anomalies observed in –Mn animals.9
Using ovariectomized rats as the model of osteopenia, Rico
et al. investigated the effects of Mn supplementation, alone
and in association with Cu, on vertebral and femoral bone
mass. The animals were divided into 4 groups of each of 15
subjects: non- ovariectomized (Sham- OVX), ovariectomized
but without supplements (OVX), ovariectomized and receiving
integration with 40 mg Mn/kg of food (OVX + Mn) and inte-
gration with Mn and 15 mg/kg of Cu (OVX + Mn + Cu) for
30 days. BMD (assessed by DXA) and bone mineral content
(BMC) at the femoral level were significantly lower in the OVX
group than in the Sham- OVX group and the 2 groups receiv-
ing supplementation with Mn and Mn +Cu; overlapping results
were also found at the vertebral level (fifth lumbar vertebra).
The authors therefore concluded that Mn supplementation is
able to inhibit effectively the loss of bone mass induced by
ovariectomy, both at the level of the axial skeleton and at the
appendiceal level, an effect not further accentuated by the addi-
tion of Cu, assuming that this inhibitory action of Mn can be
explained by the fact that this element represents an indispens-
able co- factor for some enzymes involved in carbohydrate
metabolism, in particular for the synthesis of mucopolysaccha-
rides in the bone, and is able to counteract the bone resorption
induced by free radicals.10
In another rat study, the level of Mn in the teeth and man-
dibular bone, which was significantly decreased following ova-
riectomy, increased again after administration of 17 β-estradiol,
suggesting that the deposition of Mn at the bone level could be
influenced by the estrogenic state.5,11
In a Bae & Kim study, a group of ovariectomized rats (OVX)
and a group of non- ovariectomized rats (sham- operated) were fur-
ther divided into a group fed a diet with an adequate concentration
of Mn (Mn 0.001%) and one fed a high concentration diet (0.01%
Mn) for 12 weeks. Lumbar BMD was significantly higher in sham
rats supplemented with Mn than in non- supplemented sham rats;
femoral BMD showed an increase following Mn supplementation
in both the OVX and sham groups. Furthermore, serum levels of
osteocalcin, a sensitive marker of bone formation, were consider-
ably higher in the groups with the highest intake of Mn. These
results indicate how Mn plays an important role in the formation
of bone tissue and in the increase of BMD, suggesting its possible
use in the treatment of postmenopausal osteoporosis.12
That Mn can stimulate the growth of osteoblasts and bone
regeneration has also emerged from in vitro studies on bioac-
tive glass, in which it has been observed how the addition of
Mn is able to promote, in cultures of human osteoblasts, the
expression of alkaline phosphatase (ALP) and some bone
morphogenetic proteins (BMPs),13 and to stimulate the osteo-
genetic differentiation of human mesenchymal stem cells

Rondanelli etal. 3
(hMSCs) and influence the mineralization process by increas-
ing the expression of markers of osteogenetic differentiation,
such as type I collagen, osteopontin and osteocalcin.14
Daily Intake of Manganese
Mn is one of the most abundant metals on the earth’s crust15; it
occurs naturally in surface and underground waters, especially in
conditions of anaerobiosis or low oxidation, and in numerous
foods, which are the main source of Mn for humans.16
Nuts, chocolate, cereal products, crustaceans, molluscs,
legumes and fresh fruit are foods rich in Mn; the greatest con-
tribution to the daily intake of this element is provided by
whole grains and derivatives, vegetables and fruit, while dairy
products, meat and fish provide minimal amounts.
Among drinks, tea is particularly rich in Mn, while 2 L/day
of drinking water provide on average only 40‐64 μg of Mn
(corresponding to about 2% to 3% of the quota supplied by
food)15,17,18 The absorption of Mn from foods introduced with
the diet is quite low, about 3% to 4%,15 and occurs in the small
intestine with an active mechanism and by passive diffusion.19
In humans, it has been reported that Mn deficiency can be
associated with hypocholesterolemia, bone demineralization
and reduced growth in children; however, to date, the defi-
ciency has not been clearly correlated with reduced diet intake.20
Since Mn is an abundant element in food, especially of plant
origin, the consumption of a varied and balanced diet will be
reasonably sufficient to ensure the proper contribution of this
microelement.18 On the basis of these observations, together
with the uncertain validity of the studies carried out to deter-
mine the need for Mn, in 1989 a daily dietary intake for adults
equal to 2.0‐5.0 mg of Mn was provisionally recommended.18
In 2013, the European Food Safety Agency (EFSA), judging
the available data still insufficient to be able to derive an aver-
age requirement (Average Requirement, AR) and a reference
intake for the population (Population Reference Intake, PRI)
for Mn, proposed an adequate intake (Adequate Intake, AI) for
adults (including pregnant and breastfeeding women) equal to
3.0 mg/day (considering it is not necessary to provide sex-
specific values) based on the average contributions observed in
the countries of the EU.17 The Italian Society of Human
Nutrition (SINU), on the other hand, in the IV Review of
LARN (Reference Assumption Levels of Nutrients and Energy
for the Italian population), established for adulthood, based on
the median value of the content of Mn emerged from studies
conducted on the total diet in Italy, an AI equal to 2.7 mg Mn/
day for males and 2.3 mg/day for females.20
Toxicity phenomena (specifically neurotoxicity) in humans
have been observed only in workers professionally exposed to high
concentrations of Mn in inhaled dusts and fumes, but not as a
consequence of its high intake through diet; an occasional intake
of 10 mg/day was considered safe for humans.18 However, con-
sidering the available data concerning humans insufficient and not
definitive, EFSA has not established any tolerable upper intake
Level (UL) for Mn.2 Sharing the recommendations of EFSA, and
also the SINU, the IV Review of the LARN did not consider it
possible to indicate a UL for Mn, either for the various age groups
or for pregnancy and breastfeeding.20
The constancy found in the concentration values of Mn in
different human tissues and in different ages of life seems to
suggest an adequate dietary intake together with a strong
homeostatic control by the human body.18
From the data collected through national dietary question-
naires, duplicate diet studies, total diet studies and market bas-
ket studies in several European countries, an estimated average
daily intake of Mn between 2 and 6 mg in adults emerges, with
most values around 3 mg/day; inter- individual variations may
be due to the characteristics and dietary habits of the individual
subject (eg., vegetarian diet vs mixed diet).17
In a recent Italian study, after determining, through inductively
coupled plasma mass spectrometry, the Mn content of different
foods and drinks usually consumed by the population, the average
daily intake was estimated in a sample of inhabitants of the region
Emilia- Romagna (719 subjects, 319 males and 400 females, aged
between 18 and 87 years). Each participant was subjected to a val-
idated, semi- quantitative Food Frequency Questionnaire (FFQ),
specifically developed for the Italian population of Central and
Northern Italy. From the data collected, an average intake of 2.34
mg Mn/day emerged, with the major sources represented by cere-
als, vegetables, legumes (in particular chickpeas, soy and beans),
sweets (especially dark chocolate) and drinks such as coffee and
tea.21 This value appears in line with what is estimated, in the
United States, through the Total Diet Study (average daily dietary
intake of Mn equal to 2.7 mg for men and 2.2 mg for adult
women)22 and with the figure calculated for the French adult pop-
ulation (2.5 mg/day, with 75% of the total Mn supplied by cereals
and vegetables).15 From a duplicate meal study conducted in
Belgium, however, higher average daily intake values emerged (3.1
mg Mn/day, SD 1.8; range 0.6‐8.8),23 as well as from a total diet
study that investigated the intake of Mn through the diet in the
United Kingdom (approximately 4.6 mg/day), where almost half
of this microelement was found to derive from drinks, especially
from tea (with a further 30% coming from cereals and derivatives
and about 15% from vegetables and fruit).24
Given this background, the aim of this narrative review was
to consider the state of the art on the relation between Mn and
bone health in humans and the effectiveness of Mn supple-
mentation (alone or with other micronutrients) on bone
mineralization.
Results and Discussion
Manganese and Bone: Human Studies
Several authors have investigated the possible relationship
between Mn and bone health in humans. Osteoporotic women,
for example, showed significantly lower serum Mn levels than
healthy subjects (0.02, 0.04 mg/L, respectively).25 In a prospec-
tive, placebo- controlled, double- blind study, Saltman & Strause
then assessed the importance of calcium supplementation (Ca,
250 mg in the form of a citrate- malate complex 4 times/day)

Natural Product Communications
4
with or without the addition of copper (Cu, 5.0 mg), Mn (Mn,
2.5 mg) and zinc (Zn, 15 mg) (collectively referred to as TMIN,
trace minerals, and contained in a single tablet taken once/day)
in a sample of 137 healthy Caucasian postmenopausal women
over 50 years of age (mean age 64.6 years, SD 7). The subjects
were divided into 4 groups: placebo, integration with only Ca,
integration with only TMIN and integration with Ca +TMIN.
After 2 years of supplementation, a greater loss of lumbar
BMD (L2- L4) was observed in the placebo group in women
not undergoing hormone replacement therapy (n = 76), with
significantly different values compared to the baseline and with
the only difference between statistically significant groups
found between placebo and Ca +TMIN. As for women under-
going estrogen therapy during the study, however, no signifi-
cant changes in bone mineral density were detected in any of
the 4 treatment groups. Finally, considering the cohort as a
whole, bone loss was significantly greater in the placebo group
and in the group with only TMIN supplementation compared
to the Ca +TMIN group. The merit of this study was to extend
from the animal model to humans the evidence of an essential
role of Cu, Mn and Zn in the maintenance of BMD, even if
each element has not been investigated individually.25
A study very similar to the one just described was conducted
by the same authors on 59 post- menopausal women over 50
years of age (average age 66 ± 7 years), not on estrogen ther-
apy, also divided in this case into 4 groups: placebo (n = 18);
only supplemented with 1000 mg of calcium citrate malate/
day (n = 13); only integration with TMIN, comprising 15.0 mg
Zn, 2.5 mg Cu and 5.0 mg Mn/day (n = 14); Ca + TMIN (n =
14). After a period of 2 years, the loss of BMD at the L2- L4
level (assessed by DXA) was greater in the placebo group, with
a significant difference compared to the baseline only in that
group. By comparing the 4 groups with each other, Ca +TMIN
proved to be the only treatment to differ significantly from
placebo in the change in lumbar bone density. These results
highlight how the lumbar bone loss observed in post-
menopausal women receiving calcium supplementation can be
further slowed through the simultaneous increase in trace min-
eral intake.26
That serum Mn levels may be related to bone quality and
density in postmenopausal women was also suggested by a
prospective cross- sectional pilot study of 41 postmenopausal
women who received no treatment, 21 of whom had osteopo-
rosis (lumbar and/or femoral T- score ≤−2.5), in which the
blood concentrations of some trace elements were investi-
gated, including Mn, Cu, magnesium (Mg), Zn and cadmium
(CD). The analysis of the collected data revealed a significant
positive correlation between serum Mn levels and lumbar or
femoral BMD and a negative correlation between Cu levels and
BMD at both measurement sites. On the contrary, a respec-
tively negative and positive correlation was found between the
serum levels of Mn and Cu and the number of fractures, but
no significant relationship between the concentrations of Mg,
Zn and Cd and bone parameters. This study has therefore
shown how the concentrations of manganese and copper in
serum have a certain predictive value toward lumbar and fem-
oral BMD, as well as bone quality; but considering the small
number of participants, these results should be confirmed by
further investigations.5,27
In a study of 100 subjects between 30 and 60 years of age
(average age 41.2 ± 4.9 years, 47 women and 53 men), of which
30 normal controls (T- score between 0 and −0.99), 40 osteope-
nic (T- score between - 1 and −2.49) and 30 osteoporotic
(T- score ≤−2.5), in addition, significantly lower serum levels
of calcium, Mn, Cu, and Zn were found in osteopenic and
osteoporotic subjects compared to controls. Furthermore, fol-
lowing a 12 week period of moderate aerobic physical activity
(1 hour/day for 3 days/week), a significant increase in serum
Ca and Mn concentrations and a decrease in serum concentra-
tions of Cu and Zn were observed in all 3 groups. These
changes were significantly correlated with the increase in serum
bone alkaline phosphatase (sBAP) levels and with the improve-
ment of BMD values (detected by DEXA) at both femoral and
vertebral levels. These observations seem to confirm the essen-
tial role of some trace elements, including manganese, in the
synthesis of cartilage and bone collagen, as well as in bone
mineralization. Furthermore, moderate aerobic exercise could
be protective for bone and cartilage by regulating the body lev-
els of these trace elements, involved in the biosynthesis of
matrix structures and in the inhibition of bone resorption
processes.28
In the study by Friedman and colleagues in 1987, the man-
ganese balance and clinical effects of a diet poor in this ele-
ment had been investigated in a group of 7 male subjects aged
between 19 and 22 years. The study protocol included an initial
21- day period characterized by a baseline diet, with a daily
intake of 2.59 mg of Mn, followed by 39 days of a “purified”
Figure 1. Flow chart of literature research

Rondanelli etal. 5
Table 1. Studies Regarding Manganese and Bone Health in Humans.
First author,
year Study design Setting Inclusion criteria Exclusion criteria Intervention Parallel treatments
Number of
subjects (M- F)
Duration of
the interventionPrimary outcomes
Secondary
outcomes Results
Friedman
B.J.,
198729
Balance study Metabolic unit of
the University
of Texas at
Austin
Informed consent // Purified formula diet containing 0.11 mg
Mn/d (depletion period)
Before depletion period:
baseline diet
(2.59 mg Mn/d)
prepared in the
kitchens of the
Department of
Home Economics
(base- line period);
after depletion
period: depletion
diet with Mn
supplementation
for two 5- days
periods (total Mn
1.53 and 2.55
mg/d: repletion
periods)
Seven male
subjects,
aged 19‐22
years.
Base- line
period =
21 days;
depletion
period
= 39
days; two
5- days
repletion
periods
= 10
days
Effects of
manganese-
deficient and
-adequate diets
on manganese
balance.
Physiological
effects of
manganese
depletion.
Slightly negative manganese
balance for most of
the depletion period
and positive balance
with manganese
supplementation.
Calcium, phosphorus and
alkaline phosphatase
serum concentrations
increased significantly
during manganese
depletion
Saltman
P.D.,
199325
Prospective, double-
blind, placebo-
controlled study
Free living subjects Caucasian,
postmenopausal
women, >50
years, in good
general health
as judged by
medical history
and routine
clinical blood
analysis; written
informed
consent
Subjects with <60%
compliance,
surgical
complications
or required
medications after
completion of the
study
Supplement with 250 mg of Ca as its citrate-
malate- complex, 4 times per day, with
a combination of Cu (5.0 mg), Mn (2.5
mg), and Zn (15 mg) (TMIN)
Placebo; supplement, 4
times per day, with
250 mg of Ca as
its citrate- malate-
complex alone;
combination of
Cu (5.0 mg), Mn
(2.5 mg), and Zn
(15 mg) (TMIN)
alone
137 women,
median age
64.6 years
(SD 7)
2 years Lumbar (L2- L4)
vertebral area,
bone mineral
content and
BMD
// In subjects without estrogen
therapy (n = 76), loss
of BMD was greatest
in the placebo group
and significantly
different compared
to baseline, with the
only significant group
difference between
placebo and Ca
+TMIN. For the whole
study cohort, bone
loss was significantly
greater in the placebo
group and TMIN alone
group compared to the
Ca +TMIN group
Staruse L.,
199426
Double- blind,
placebo-
controlled
prospective trial
Free- living subjects Post- menopausal
women > 50
y old and in
good general
health as judged
by medical
history and
routine clinical
blood analyses
(complete
blood count
and differential
count); written
informed
consent
A positive Pap smear
or mammogram
during the
previous year;
any disease
or condition
known to affect
bone or calcium
metabolism; a
history of chronic
renal, hepatic or
gastrointestinal
disease; evidence
of collapsed or
focal vertebral
sclerosis. To be a
dedicated athlete;
use of estrogen,
calcitonin, vitamin
D, fluoride,
bisphosphonates,
thyroid hormone
or thiazides
Active calcium (1000 mg elemental calcium/d
in the form of calcium citrate malate)
+active trace minerals (15.0 mg of zinc
as sulfate salt, 2.5 mg of copper, and 5.0
mg of manganese as gluconate salts/d)
Placebo calcium
+placebo trace
minerals; placebo
calcium +active
trace minerals;
active calcium
+placebo trace
minerals
59 Caucasian
post-
menopausal
women,
mean age
66 ± 7
years
2 years Lumbar (L2- L4)
bone mineral
density (BMD)
measured
by dual
energy X- ray
absorptiometry
// Loss of bone density
was greatest in the
placebo group and was
significantly different
compared with the
base- line value; bone
loss compared with the
base- line value was not
significant for women
given calcium alone,
trace minerals alone
or calcium plus trace
minerals. In comparing
the 4 treatment groups,
calcium plus trace
minerals was the only
treatment that differed
significantly from
placebo
(Continued)

Natural Product Communications
6
diet containing 0.11 mg Mn/day (depletion period) and subse-
quently by 2 periods of 5 days each with a progressively higher
daily intake of Mn (1.53 mg the first 5 days and 2.55 mg/day
the last 5; repletion period). The participants showed a slightly
negative Mn balance during the depletion period, which then
turned positive in the 10 following days of Mn reintegration.
Furthermore, a significant increase in serum concentrations of
calcium, phosphorus and alkaline phosphatase (ALP) had been
found in the 39 days of the Mn- poor diet, suggesting that the
deficiency of Mn could influence bone remodeling.25,29
Finally, a clinical trial conducted in Mexico in 2001 demon-
strated how a multiple supplement with vitamins and microele-
ments, including Mn, {administration of a drink containing the
recommended daily dose -RDA- of vitamins D3, E, K1, B6,
niacin, thiamine, biotin, folic acid, pantothenic acid and miner-
als (iodine, Cu, Mn and selenium); 1.2 times the RDA of vita-
min A and 1.5 times the RDA of ascorbic acid, riboflavin,
vitamin B12, iron and Zn)} given 6 days/week for an average
period of 12.2 months, is able to improve growth in length
(with a gain of almost 5 mm) in children aged between 8 and
14 months compared to the placebo group . In particular, the
effect of supplementation was greater in children less than 12
months of age at the start of the study (average increase in
length of 8.3 mm) compared to children ≥ 12 months (average
increase of 2.0 mm).30
Unfortunately, to date, the available data are insufficient to
establish an AR and a PRI for Mn; for European countries,
however, the EFSA (European Food Safety Agency) has pro-
posed an AI for adults equal to 3 mg/day.17 Similarly, the SINU,
in the IV revision of the LARN (Reference Assumption Levels
of Nutrients and Energy for the Italian population), has estab-
lished, for the adult Italian population, an AI equal to 2.7 mg/
day for males and 2.3 mg/day for females.20 Furthermore, cur-
rently, again due to the scarcity and inadequacy of the available
data, a tolerable Upper intake Level of Mn has not yet been
established.2,20
Research conducted in several European countries has
shown an estimated average daily intake of 2‐6 mg, with most
values around 3 mg/day,17 values which are in line with what is
also estimated in a recent Italian study (average adult intake of
2.34 mg Mn/ day).21
All the literature published to date is in agreement in show-
ing that osteoporotic women have lower serum Mn levels than
women with normal bone mineral density),25 thus confirming
the essential role of manganese in the synthesis of cartilage
and bone collagen, as well as in bone mineralization5,27 and
confirming the studies on the animal model.
Conclusion
Considering the human studies that evaluated the effectiveness
of an oral manganese supplement for a long period (2 years)
on the bone mineral density of menopausal women (in total
196 women), both of the double blind clinical trials that evalu-
ated this topic showed that bone loss was significantly greater
First author,
year Study design Setting Inclusion criteria Exclusion criteria Intervention Parallel treatments
Number of
subjects (M- F)
Duration of
the interventionPrimary outcomes
Secondary
outcomes Results
Rivera J.A.,
200130
Double- blind,
placebo-
controlled,
supplementation
trial
Mexican subjects
living in a
setting where
the diet is
poor in several
micronutrients
Written consent
obtained from
parents
// 30 ml of a beverage containing multiple
micronutrients (the RDA for children
aged 1‐3 y of the vitamins D3 (as
cholecalciferol), E (as tocopheryl
acetate), K1, niacin (as nicotinamide),
thiamine (as thiamine mononitrate),
B-6 (as pyridoxine hydrochloride),
D- biotin, folic acid, and pantothenic
acid (as D- calcium pantothenate) and
of the minerals iodine (as potassium
iodide), copper (as copper gluconate),
manganese (as manganese sulfate), and
selenium (as selenium sulfate); 1.2 times
the RDA for children aged 1‐3 y of
vitamin A (as retinyl palmitate) and 1.5
times the RDA of ascorbic acid (extra
fine), riboflavin, vitamin B-12, iron (as
ferric orthophosphate), and zinc (as zinc
sulfate)), 6d/wk
30 ml of placebo 319 children
aged 8‐14
mo
Average of
12.2 ±
1.8 mo
Linear growth
of children
evaluated by
measurement
of body length
// Supplemented infants
initially aged <12 mo
had significantly greater
length gains than did
the placebo group,
with a difference of
8.2 mm at the end of
supplementation. In
contrast, differences in
length gains between
the supplemented and
placebo groups initially
aged ≥12 mo were not
significant
Table 1. Continued

Rondanelli etal. 7
in the placebo group than in the group taking supplementation,
equal to 5.0 mg Mn/day for the Straus study,26 and 2.5 mg Mn/
day for the study of Saltman et al.,25 but considering, however,
that supplementation was represented by a set of microele-
ments (Mn, Cu and Zn) and by calcium.
Material and Methods
The present narrative review was performed following the
steps by Egger et al.31 as follows:
1. Configuration of a working group: 3 operators skilled in
clinical nutrition (one acting as a methodological opera-
tor and 2 participating as clinical operators).
2. Formulation of the revision question on the basis of
considerations made in the abstract: “the state of the art
on the relation between Mn and bone health in humans
and the effectiveness of manganese supplementation
(alone or with other micronutrients) on bone mineral-
ization”
3. Identification of relevant studies: a research strategy was
planned on PubMed (Public MedIine run by the Nation-
al Center of Biotechnology Information (NCBI) of the
National Library of Medicine of Bethesda (USA)) as fol-
lows: (a) Definition of the keywords (manganese, bone
health, humans, supplementation, bone mineral densi-
ty), allowing the definition of the interest field of the
documents to be searched, grouped in quotation marks
(“…”) and used separately or in combination; (b) use of:
the Boolean (a data type with only 2 possible values: true
or false) AND operator, that allows the establishments
of logical relations among concepts; (c) Research mo-
dalities: advanced search; (d) Limits: time limits: papers
published in the last 20 years; humans; adults; languag-
es: English; (e) Manual search performed by the senior
researchers experienced in clinical nutrition through the
revision of articles on the state of the art on the relation
between manganese and bone health in humans and the
effectiveness of manganese supplementation (alone or
with other micronutrients) on bone mineralization
4. Published in journals qualified in the Index Medicus.
5. Analysis and presentation of the outcomes: we create
paragraphs about the state of the art on the relation be-
tween manganese and bone health in humans and the
effectiveness of manganese supplementation (alone or
with other micronutrients) on bone mineralization and
the data extrapolated from the “revised studies” were
collocated in tables; in particular, for each study we spec-
ified the author and year of publication and study char-
acteristics.
6. The analysis was carried out in the form of a narrative
review of the reports. At the beginning of each section,
the keywords considered and the type of studies chosen
are reported. We evaluated, as is suitable for the narrative
review, studies of any design which considered the state
of the art on the relation between manganese and bone
health in humans and the effectiveness of manganese
supplementation (alone or with other micronutrients) on
bone mineralization
Table1 shows studies regarding manganese and bone health
in humans.
Figure1 shows flow chart of literature research.
Declaration of Conflicting Interests
The author(s) declared no potential conflicts of interest with respect
to the research, authorship, and/or publication of this article.
Funding
The author(s) received no financial support for the research,
authorship, and/or publication of this article.
ORCID ID
Gabriella Peroni https:// orcid. org/ 0000- 0002- 1632- 1787
References
1. Expert Group on Vitamins and Minerals. Safe Upper Levels for
Vitamins and Minerals; 2003.
2. European Food Safety Authority. Tolerable Upper Intake Levels for
Vitamins and Minerals. Scientific Committee on Food; Scientific Panel on
Dietetic Products, Nutrition and Allergies; 2006.
3. Food and Nutrition Board. Dietary reference intakes for vitamin A,
vitamin K, arsenic, boron, chromium, copper, iodine, iron, manganese, molyb-
denum, nickel, silicon, vanadium. National Academies Press; 2001.
4. O'Neal SL, Hong L, Fu S, etal. Manganese accumulation in bone
following chronic exposure in rats: steady- state concentration
and half- life in bone. Toxicol Lett. 2014;229(1):93-100. doi: 10.
1016/ j. toxlet. 2014. 06. 019
5. Zofková I, Nemcikova P, Matucha P. Trace elements and
bone health. Clin Chem Lab Med. 2013;51(8):1-7. doi: 10. 1515/
cclm- 2012- 0868
6. Strause LG, Hegenauer J, Saltman P, Cone R, Resnick D. Effects
of long- term dietary manganese and copper deficiency on rat
skeleton. JNutr. 1986;116(1):135-141. doi: 10. 1093/ jn/ 116. 1. 135
7. Liu R, Jin C, Wang Z, Wang Z, Wang J, Wang L. Effects of
manganese deficiency on the microstructure of proximal tibia
and OPG/RANKL gene expression in chicks. Vet Res Commun.
2015;39(1):31-37. doi: 10. 1007/ s11259- 015- 9626-5
8. Wang J, Wang ZY, Wang ZJ, Liu R, Liu SQ, Wang L. Effects
of manganese deficiency on chondrocyte development in
tibia growth plate of Arbor Acres chicks. J Bone Miner Metab.
2015;33(1):23-29. doi: 10. 1007/ s00774- 014- 0563-0
9. Clegg MS, Donovan SM, Monaco MH, Baly DL, Ensunsa JL,
Keen CL. The influence of manganese deficiency on serum
IGF-1 and IGF binding proteins in the male rat. Proc Soc Exp Biol
Med. 1998;219(1):41-47. doi: 10. 3181/ 00379727- 219- 44314
10. Rico H, Gómez- Raso N, Revilla M, etal. Effects on bone loss
of manganese alone or with copper supplement in ovariecto-
mized rats. A morphometric and densitomeric study. Eur J Obstet

Natural Product Communications
8
Gynecol Reprod Biol. 2000;90(1):97-101. doi: 10. 1016/ s0301- 2115(
99) 00223-7
11. Rahnama M, Błoniarz J, Zareba S, Swiatkowski W. Study of estro-
gen deficiency impact on manganese levels in teeth and mandible
of rats after ovariectomy. Rocz Panstw Zakl Hig. 2003;54(1):33-38.
12. Bae Y- J, Kim M- H. Manganese supplementation improves
mineral density of the spine and femur and serum osteocal-
cin in rats. Biol Trace Elem Res. 2008;124(1):28-34. doi: 10. 1007/
s12011- 008- 8119-6
13. Miola M, Brovarone CV, Maina G, et al. In vitro study of
manganese- doped bioactive glasses for bone regeneration. Mater
Sci Eng C Mater Biol Appl. 2014;38(1):107-118. doi: 10. 1016/ j.
msec. 2014. 01. 045
14. Barrioni BR, Norris E, Li S, Naruphontjirakul P, Jones JR,
Pereira MdeM. Osteogenic potential of sol- gel bioactive glasses
containing manganese. JMater Sci Mater Med. 2019;30(7):86. doi:
10. 1007/ s10856- 019- 6288-9
15. Biego GH, Joyeux M, Hartemann P, Debry G. Daily intake of
essential minerals and metallic micropollutants from foods in
France. Sci Total Environ. 1998;217(1-2):27-36. doi: 10. 1016/
S0048- 9697( 98) 00160-0
16. World Health Organization. Guidelines for Drinking- Water Quality.
4th ed; 2011.
17. European Food Safety Authority. Scientific opinion on dietary
reference values for manganese. Efsa J. 2013;11(11):3419.
18. National Research Council. Subcommittee on the Tenth Edition of
the Recommended Dietary Allowances. Recommended Dietary Allowances.
10th ed. National Academies Press; 1989.
19. Greger JL. Nutrition versus toxicology of manganese in
humans: evaluation of potential biomarkers. Neurotoxicology.
1999;20(2-3):205-212.
20. SINU. Tabelle Larn. http://www. sinu. it/ html/ pag/ tabelle_ larn_
2014_ rev. asp
21. Filippini T, Cilloni S, Malavolti M, etal. Dietary intake of cad-
mium, chromium, copper, manganese, selenium and zinc in a
northern Italy community. JTrace Elem Med Biol. 2018;50:508-517.
doi: 10. 1016/ j. jtemb. 2018. 03. 001
22. Pennington JA, Young BE, Wilson DB. Nutritional elements in
U.S. diets: results from the total diet study, 1982 to 1986. J Am
Diet Assoc. 1989;89(5):659-664.
23. Buchet JP, Lauwerys R, Vandevoorde A, Pycke JM. Oral daily
intake of cadmium, lead, manganese, copper, chromium, mer-
cury, calcium, zinc and arsenic in Belgium: a duplicate meal study.
Food Chem Toxicol. 1983;21(1):19-24. doi: 10. 1016/ 0278- 6915( 83)
90263-6
24. Wenlock RW, Buss DH, Dixon EJ. Trace nutrients. Br J Nutr.
1979;41(2):253-261. doi: 10. 1079/ BJN1 9790034
25. Saltman PD, Strause LG. The role of trace minerals in osteopo-
rosis. JAm Coll Nutr. 1993;12(4):384-389. doi: 10. 1080/ 07315724.
1993. 10718327
26. Strause L, Saltman P, Smith KT, Bracker M, Andon MB. Spinal
bone loss in postmenopausal women supplemented with calcium
and trace minerals. JNutr. 1994;124(7):1060-1064. doi: 10. 1093/
jn/ 124. 7. 1060
27. Nemcikova P, Spevackova V, Cejchanova M, Hill M, Zofkova I.
Relationship of serum manganese and copper levels to bone den-
sity and quality in postmenopausal women. Apilot study. Osteol
Bull. 2009;14:97-100.
28. Alghadir AH, Gabr SA, Al- Eisa ES, Alghadir MH. Correla-
tion between bone mineral density and serum trace elements in
response to supervised aerobic training in older adults. Clin Interv
Aging. 2016;11:265-273. doi: 10. 2147/ CIA. S100566
29. Friedman BJ, Freeland- Graves JH, Bales CW, et al. Manganese
balance and clinical observations in young men fed a manganese-
deficient diet. JNutr. 1987;117(1):133-143. doi: 10. 1093/ jn/ 117. 1.
133
30. Rivera JA, González- Cossío T, Flores M, etal. Multiple micro-
nutrient supplementation increases the growth of Mexican
infants. AmJ Clin Nutr. 2001;74(5):657-663. doi: 10. 1093/ ajcn/
74. 5. 657
31. Egger M, Dickersin K, Smith GD. Problems and Limitations in
Conducting Systematic Reviews. In: Systematic Reviews in Health
Care. BMJ Publishing Group; 2008:43-68.