Ruan et al. BMC Pediatrics(2024) 24:530
https://doi.org/10.1186/s12887-024-04996-5

BMC Pediatrics

RESEARCH

Open Access

The effect of iron supplementation in preterm
infants at different gestational ages
Sufeng Ruan1, Jinrong Li1, Fei Xiong1, Di Qie1, You Lu1, Sufei Yang1, Zhanghui Tang1 and Fan Yang1*
Abstract
Background Iron deficiency (ID) is the most prevalent nutritional deficiency disease in preterm infants, significantly
affecting their growth and development. For preterm infants to flourish physically and neurologically, timely
iron supplementation is essential. The main goals of this study were to determine whether the present ironsupplementation regimen results in iron overload in late preterm infants and whether it can meet the growthrequirements of early preterm infants for catch-up.
Methods We conducted a prospective follow-up study on preterm infants at the Department of Child Health, West
China Second University Hospital, Sichuan University, from January 1, 2020, to August 31, 2020. In this study, 177preterm infants were divided into two groups based on gestational age—early preterm infants (gestational age < 34weeks) and late preterm infants (gestational age ≥ 34 weeks and < 37 weeks)—to compare the incidence of irondeficiency, iron status, and physical growth of preterm infants receiving iron supplements (2–4 mg/kg/d).
Results Iron supplementation considerably reduced the incidence of iron deficiency in preterm infants. The
prevalence of iron deficiency in early preterm infants and late preterm infants was 11.3% and 5.1%, respectively, at thecorrected gestational age of 3 months; at the corrected gestational age of 6 months, the prevalence was 5.3% and6.3%, respectively. No preterm infants with iron deficiency were detected in either group at the corrected gestationalage of 12 months. Ferritin was substantially lower in early preterm infants (36.87 ± 31.57 ng/ml) than in late preterm
infants (65.78 ± 75.76 ng/ml) at the corrected gestational age of 3 months (p < 0.05). A multifactorial regression
analysis of factors influencing iron metabolism levels in preterm infants revealed a positive relationship between
log10hepcidin, birth weight, and ferritin, with higher birth weights resulting in higher ferritin levels.
Conclusions Postnatal iron supplementation at 2–4 mg/kg/d in preterm infants significantly decreases the incidenceof ID. There were substantial differences in iron levels across preterm infants of varying gestational ages. A tailorediron supplementation plan based on growth, birth weight, and gestational age may be a more suitable route for ironsupplementation. Although the current study found that the postnatal iron status of early preterm infants differedfrom that of late preterm infants, the actual mechanism of action remains unknown, and large-sample, multicenter
clinical studies are required to investigate this further.
Keywords Preterm infant, Iron supplementation, Iron metabolism, Iron overload, Hepcidin*Correspondence:
Fan Yang
506742295@qq.com
1
Department of Pediatrics, Key Laboratory of Birth Defects and RelatedDiseases of Women and Children (Sichuan University), Ministry of
Education, West China Second University Hospital, Sichuan University,
Chengdu, Sichuan Province 610041, China© The Author(s) 2024. Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0
International License, which permits any non-commercial use, sharing, 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 you modified thelicensed material. You do not have permission under this licence to share adapted material derived from this article or parts of it. The images orother third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to thematerial. 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 http://
creativecommons.org/licenses/by-nc-nd/4.0/.

Ruan et al. BMC Pediatrics(2024) 24:530

Background
Iron is an essential micronutrient for cellular activities
such as hemoglobin synthesis, DNA synthesis, energy
metabolism, electron transfer, and cellular differentiation [1]. It is also important for infant and child neurodevelopment, physical growth, and immune function [2,3]. Preterm infants have a much greater risk of iron deficiency and iron deficiency anemia [4] than term infants
due to inadequate iron stores, a relatively short red blood
cell lifespan, postnatal catch-up growth, and medical
blood loss [5, 6]. The prevalence of iron deficiency (ID)and iron deficiency anemia (IDA) in preterm infants varies by country, geography, and age and can reach 48% and26.5%, respectively [5]. Prior to the development of IDA,
ID can cause irreparable damage to the nervous system[32]. It is especially crucial to identify and treat iron deficiency early in preterm infants.
Preterm infants require substantially more iron thanterm infants since they need to catch up on growth. In
the absence of exogenous iron supplementation, preterm infants’ innate iron stores can only meet the iron
requirements for catch-up and double birth weight [7].
Food sources of iron are inadequate to meet the irondemands of preterm infants, and oral iron supplements
are frequently required [8]. There are no defined guidelines for the starting time, dose, or finishing time of oraliron intake for preterm infants. According to China’s
Recommendations for the Prevention and Control ofIron Deficiency and Iron Deficiency Anemia in Children,
preterm infants should begin receiving iron supplements(2–4 mg/kg/d) two weeks after delivery [9].
Hepcidin is one of the main regulators of iron metabolism. Hepcidin binds to ferroportin, internalizing anddegrading it. This reduces the release of iron from hepatocytes, macrophages, and intestinal cells into the bloodstream, resulting in decreased dietary iron absorption
and serum iron, and increased intracellular iron [10].
Compared to healthy newborns, infants with ID exhibited significantly decreased hepcidin levels at 1.5 monthsof age [11]. This shows that hepcidin can be an early indicator of newborn iron status. Little research has been
conducted on the hepcidin level in preterm infants, andfurther research is still needed to determine the specificity and sensitivity of hepcidin for the diagnosis of ID and
IDA in preterm infants.
Neonates mostly acquire iron from their mothers
throughout late gestation [12]. Early preterm infants may
have significantly lower body iron levels than late preterm
infants. Current guidelines for preventing iron deficiency
in preterm infants are based on studies of iron status inearly preterm infants (gestational age < 34 weeks) [13],with few studies on iron status in late preterm infants anda lack of studies on iron status in preterm infants of various gestational ages. This study classified preterm infants

Page 2 of 10

into two groups based on gestational age: early preterm
infants and late preterm infants. By comparing their ironmetabolism and physical growth indicators, we evaluatedwhether the present iron supplementation regimen could
meet the iron demand of early preterm infants and if it
resulted in iron overload in late preterm infants.

Methods
Study population

This prospective cohort study was carried out at the
Department of Child Health, West China Second University Hospital, Sichuan University. A total of 180 preterm infants were recruited at the West China Second
University Hospital’s Child Health Unit between January
1, 2020, and August 31, 2020. This study included infants
with a gestational age less than 37 weeks and appropriate
for gestational age. The study excluded preterm infants
with gastrointestinal disorders, active bleeding, serious
infections, severe cardiovascular problems, cerebrovascular diseases, liver diseases, renal diseases, neuropsychiatric diseases, and hereditary metabolic diseases.
The parents of the research subjects consented to theirinfant’s involvement in the study, signed an informed
consent form, and demonstrated good compliancethroughout the whole study process. We gathered baseline information from research participants at the corrected gestational age of 40 weeks of preterm, and those
who did not fit the requirements for inclusion were notincluded.
The corrected gestational age of preterm infants
was calculated by subtracting the remaining weeks toreach 400/7 weeks. The infants were required to visit
the study center three days before or three days after
the corrected gestational age of 3, 6 and 12 months.
The data of those who followed up in accordance withthe agreed-upon time were recorded and included in
the analysis of the results. If the subjects did not follow up during the specified period, they were added to
the list of missed visits, and the data for these cases
were recorded but excluded from the results analysis.
The records were adjusted for follow-up at the Child
Health Unit at the corrected gestational age of 3, 6,
and 12 months, and each of the three follow-up visits
included a complete record of the infants’ hemoglobin
levels and at least two iron metabolism indices.
Data collection

An electronic medical information system was used
to obtain information on the participants’ mothers’age, gestational comorbidities, iron supplementation
throughout pregnancy, maternal ferritin levels, education level, maternal history, fetal sex, birth weight, andgestational age. At the follow-up appointments for preterm correction at 3, 6, and 12 months of gestational

Ruan et al. BMC Pediatrics(2024) 24:530

ages, the following data were collected: feeding status, blood counts, iron metabolism markers, and bodymeasurements. The period for gathering the data was
January 1, 2020, through February 28, 2022.
Case definitions

The study divided preterm infants into two groups
based on gestational age: the early preterm group (gestational age < 34 weeks) and the late preterm group(gestational age ≥ 34 weeks and < 37 weeks). According
to the Chinese Pediatric Association’s Recommendations for Prevention and Control of Iron Deficiency
and Iron Deficiency Anemia in Children, preterm
infants were given iron supplements (2–4 mg/kg/d)
between two weeks and twelve months of age after
delivery.
Blood was obtained in the morning while the patient
was fasting. Hemoglobin (HGB) was identified using
sulfated hemoglobin spectrophotometry, as well asmean corpuscular volume (MCV), mean corpuscularhemoglobin (MCH), and mean corpuscular hemoglobin concentration (MCHC). Serum iron (SI) was
identified using the ferrozine method (ADVIA XPT
automatic biochemical analysis, Germany). Chemiluminescence was used to detect ferritin (SF) (CENTAUR XPT, Germany). Total iron binding capacity(TIBC, ADVIA XPT fully automated biochemical
analyzer, Germany) was measured using an immunoturbidimetric technique. Transferrin saturation (TS)was computed using SI and TIBC. The enzyme-linked
immunosorbent test (ELISA) was used to quantify ferritin (RD, USA).
The ID diagnostic criteria were as follows:
SF < 12 µg/L and CRP < 5 mg/L or SF < 30 µg/L andCRP ≥ 5 mg/L [14, 15]; anemia was defined as hemoglobin levels below 90 g/L at 1–4 months, < 100 g/L
at 4–6 months, and < 110 g/L between 6 months and6 years of age [16]. Iron deficiency anemia is characterized by the presence of both iron deficiency andanemia.
Statistical analysis

SPSS 23.0 was used to analyze the data. The fundamental features of the study population were analyzed
using descriptive statistics; the measurement data were
described using the mean ± standard deviation (𝒙̅ ± 𝒔),
and the count data were characterized using the constructed ratio. The distribution of basic features acrossgroups was examined for differences. Measurement
data was analyzed using the t-test, and count data wereanalyzed using the 𝜒 2 test or Fisher’s exact probability test. Multivariate logistic regression models were
utilized to examine the correlations among hepcidin,

Page 3 of 10

ferritin, and markers associated with iron metabolismthat impact preterm infants.
Ethical statement

The Ethics Review Committee of West China Second
Hospital of Sichuan University approved this study (Medical Research 2019 On Approval No. (086)). Informed
consent to participate in the study was obtained from theparticipants’ parents or legal guardians.

Results
A total of 180 preterm infants were included in the regular follow-up. There were 177, 177, and 173 preterm
infants followed up to the corrected gestational age of
3, 6, and 12 months, respectively, of whom 162, 169, and
154 underwent blood sampling (Fig. 1).
Univariate analyses of clinical characteristics

Compared to the late preterm group, the early pretermgroup had a lower gestational age, birth weight, and birthlength (P = 0.00, Table 1). Early preterm infants were more
likely than late preterm infants to require blood transfusion and NICU hospitalization for treatment (P < 0.001,
Table 1). Our comparison of iron metabolism indicators
in early preterm infants of corrected gestational age of
3 months in the transfused and non-transfused groups
revealed no significant differences (Table 2).
Iron metabolism index analysis in two groups of preterm
infants

Compared with late preterm infants, early preterm
infants demonstrated significantly lower MCHC
(335.41 ± 13.41 g/L vs. 340.22 ± 14.56 g/L, P < 0.05) andSF (36.87 ± 31.57 ng/mL vs. 65.78 ± 75.76 ng/mL, P < 0.05;
Table 3) at the corrected gestational age of 3 months.
There was no statistically significant difference in ironmetabolism indicators between the two groups of preterm infants at the corrected gestational age of 6 and12 months. The prevalence of iron deficiency in earlypreterm infants and late preterm infants was 11.3% and5.1%, respectively, at the corrected gestational age of 3
months; at the corrected gestational age of 6 months, theprevalence was 5.3% and 6.3%, respectively. This difference was not statistically significant. Iron deficiency was
not detected in any preterm infants at the corrected gestational age of 12 months.
Based on ferritin levels, the corrected 6-month-old
preterm infants were categorized into two groups: thegroup without ID and the group with ID. Table 4 displays
a comparison of the iron metabolism parameters of preterm infants in the groups with ID and those without ID.
Preterm infants in the ID group had considerably lower
Hb, MCV, SI, SF, TIBC, and hepcidin levels than those in
the non-ID group, and the differences were statisticallyRuan et al. BMC Pediatrics(2024) 24:530

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Fig. 1 Flowchart of the study. Early-preterm infant: gestational age < 34 weeks; late-preterm infant: gestational age ≥ 34 weeks and < 37 weeks

significant (P < 0.05). Preterm infants in the ID group had
greater TS than those in the non-ID group, and this difference was statistically significant (P < 0.05).
Multifactorial regression analysis was used to determine iron metabolism and physical growth indices inpreterm infants to investigate the factors influencing ferritin and hepcidin in preterm infants. Two significant factors influencing hepcidin in preterm infants are weightgain and ferritin. Preterm infants with faster weight gain
exhibited greater levels of hepcidin (OR= -3.531, 95%
CI: 227.83 to 5.73, p = 0.04), as did preterm infants withhigher ferritin levels (OR = 0.699, 95% CI: 0.36 to 1.01,
p = 0). Ferritin in preterm infants was significantly influenced by birth weight, and hepcidin: Ferritin levels weregreater in preterm infants with heavier birth weights(OR = 0.335, 95% CI: 0.01 to 0.03, p = 0.01; Table 5).
Analysis of physical growth indices in two groups ofpreterm infants

Table 6 presents a comparison of physical growth indices between the two groups of preterm infants. The
heights of the early preterm infants at 3, 6, and 12
months were substantially shorter than those of the late
preterm infants (60.59 ± 2.45 cm vs. 61.52 ± 2.13 cm,
66.63 ± 2.95 cm vs. 67.54 ± 2.19 cm, and 75.03 ± 3.34 cm
vs. 76.15 ± 2.80 cm, respectively, P < 0.05). The body massindex at corrected 3 months of age was greater in earlypreterm infants (18.08 ± 1.87 kg/m2 vs. 17.51 ± 1.45 kg/
m2, P < 0.05) than in late preterm infants. There wereno statistically significant differences in weight, sittingheight, head circumference, weight gain, or height gain(p > 0.05) between the two groups of preterm infants at
3, 6, or 12 months. There was no significant difference inthe physical growth indices of preterm infants between
the ID group and the non-ID group.

Discussion
The current study offers an important update on theeffects of long-term, frequent iron supplementation
on iron metabolism, growth, and development in earlyand late preterm infants. We discovered that consistentlong-term iron supplementation up to 1 year of preterm
corrected age greatly reduced the frequency of iron deficiency in preterm infants while not producing iron overload in late preterm infants or impairing preterm growthand development.
In our research, the prevalence of ID in preterm infants
declined gradually with age and continued iron supplementation; at one year of age, no preterm infants with
ID were detected in either group. This finding implies
that daily iron supplementation at 2–4 mg/kg considerably lowers the incidence of ID compared to that inpreterm infants who do not receive oral iron supplementation. This finding is consistent with the results ofthe majority of related studies, which have shown thatiron supplementation improves hematological indicators or iron status in preterm infants and reduces theincidence of anemia or iron deficiency [17]. In a trial of
100 term newborns, Bora R et al. reported that infants
at risk of iron deficiency with daily supplementation

Ruan et al. BMC Pediatrics(2024) 24:530

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Table 1 Comparison of baseline characteristics between the two groups
Variables
Neonatal(N)Female,%Birth weight, gBirth length, cm
Gestational age, wk
Transfuse blood
No, n(%)
Yes, n(%)
Mode of delivery, vaginal delivery, n(%)
Regular iron supplementation, yes, n(%)
Admission to NICU after birth, yes, n(%)
Feeding Methods
Exclusive breast feeding, n(%)
Mixed feeding, n(%)
Artificial feeding, n(%)
Timing of complementary food introduction, months
Mother(N)Maternal age
Mother’s attained education, n(%)
Primary and below, n(%)
Junior high school (secondary school), n(%)
High school (technical secondary school), n(%)
University and above, n(%)
Ferritin, ng/ml
Gestational obesity, n(%)
IVF, n(%)
Anemia, n(%)
Premature rupture of membranes, n(%)
Gestational diabetes, n(%)
Hypothyroidism, n(%)

Early-preterm infant
70
38(54.30%)
1643.01 ± 422.34
40.44 ± 3.69
31.78 ± 2.06

Late-preterm infant
107
53(49.50%)
2295.88 ± 392.39
45.03 ± 2.50
35.26 ± 0.94t, z or x2

P value

0.383
10.5
9.12
13.23

0.536
0.00
0.00
0.00

50
11
18(25.7%)60(85.7%)
61(100%)90
014(13.1%)
93(86.9%)41(45.6%)

12.04

0.001

4.55
0.052
4.55

0.033
0.82
0.033

21(30.0%)17(24.3%)
32(45.7%)
6.18 ± 0.71
70
32.06 ± 4.91

40(37.4%)
36(33.6%)
31(29.0%)
5.97 ± 0.65
107
32.48 ± 4.14

5.24

0.073

2.02

0.045

0.543

0.588

13(19.4%)
22(32.8%)21(31.3%)
11(16.4%)
57.195 ± 73.76
4(5.7%)
23(32.9%)6(9.8%)
17(27.9%)20(32.8%)
07(9.1%)
26(33.8%)35(45.5)9(11.7%)38.25 ± 52.78
2(1.9%)
38(35.5%)6(7.1%)
22(25.9%)19(22.4%)6(7.1%)

5.164

0.16

1.535
1.03
0.132
0.363
0.07
1.97
2.87

0.129
0.31
0.716
0.547
0.79
0.16
0.09

Data are presented as mean ± SD, range, or number of patients (percentages). IVF, in vitro fertilization. A total of 14.6% of the infants had missing data for transfused
blood (Early-preterm infant, 5%; Late-preterm infant, 9.6%). A total of 18.6% of mothers had missing data for mother’s attained education (Early-preterm infant, 1.7%;
Late-preterm infant, 16.9%). A total of 14.7% of the infants had missing data for admission to NICU after birth (Early-preterm infant, 5.1%; Late-preterm infant,9.6%).
A total of 17.5% of mothers had missing data for anemia, premature rupture of membranes, gestational diabetes and hypothyroidism (Early-preterm infant, 5.1%;
Late-preterm infant,12.4%). Missing data were not included in the study. *P value < 0.05 was considered to indicate statistical significanceof 2 mg/kg iron had higher Hb (103.7 ± 9.3 g/L vs.
97.0 ± 9.4 g/L, P < 0.0001) and serum ferritin concentrations (133.93 ± 52.41 ng/mL vs. 78.09 ± 42.03 ng/mL,
P < 0.001) [18]. In a trial involving 285 low birth weight
infants, Berglund S et al. reported a noteworthy decreasein the risk of ID in preterm infants supplemented with
2 mg/kg iron daily [19].
To further study whether blood transfusion in the
neonatal period affects the efficiency of normal ironsupplementation in preterm infants, we compared theiron status parameters of early preterm infants to thoseof infants who did not receive blood transfusions. At thecorrected age of 3 months, we analyzed the iron samples of early preterm infants in the transfused and nontransfused groups and observed no significant differencein ferritin between the two groups. This is because preterm infants receiving blood transfusions receive comparatively less iron than those receiving long-term ironsupplements. Heme iron makes up about 190–210 milligrams per unit of blood [20]. About 140 mL of red blood
cells make up one unit of blood. Preterm babies usually
receive a 15 mL/kg red blood cell transfusion [21]. For
preterm infants, each transfusion provides 20-22.5 mg/
kg of elemental iron. China’s Recommendations for thePrevention and Control of Iron Deficiency and Iron Deficiency Anemia in Children state that two weeks after
delivery, preterm newborns should start taking ironsupplements (2–4 mg/kg/d) [9]. Therefore, regardless ofwhether the neonate receives a transfusion or not, we
believe that early preterm infants should receive an ironsupplementation of 2–4 mg/kg/day.
Ferritin levels in late preterm infants in our study
decreased significantly between 3 months and 6 monthsof age and slowed down between 6 and 12 months of age.
In contrast, early preterm infants showed no discernible
changes in ferritin levels between 3 and 12 months ofRuan et al. BMC Pediatrics(2024) 24:530

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Table 2 Comparison of iron metabolism levels between
transfusion early-preterm infant and non-transfusion earlypreterm infant of the corrected gestational age of 3 months
Transfusion
early-preterm
infantchronologicalages, days
Hb, g/L
MCV, fl.
MCH, pg
MCHC, g/L
SI, umol/L
SF, ng/ml
TIBC, umol/LTS, %t, z or x2

151.92 ± 20.05

Nontransfusion
early-preterm
infant126.0 ± 11.38

P
value

9.82

0.001

116.25 ± 7.40
80.05 ± 4.94
27.18 ± 1.56
339.50 ± 16.81
11.53 ± 2.05
37.98 ± 40.01
20.19 ± 6.95
60.03 ± 11.89

118.55 ± 11.37
77.83 ± 4.78
26.16 ± 2.13
335.81 ± 12.03
10.99 ± 3.38
37.80 ± 31.02
17.74 ± 5.99
63.59 ± 10.91

-0.55
1.20
1.27
0.75
0.59
0.01
1.03
-0.83

0.59
0.24
0.21
0.46
0.56
0.99
0.31
0.41

Data are presented as mean ± SD. Hb, hemoglobin; MCV, mean corpuscularvolume; MCH, mean corpuscular hemoglobin; MCHC, mean corpuscularhemoglobin concentration; SI, serum iron; SF, ferritin; TS, transferrin saturation;
TIBC, total iron binding capacity. *P value < 0.05 was considered to indicate
statistical significance

Table 4 Comparison of physical development indicators
between the two groups

3 months’corrected Age
Weight, kg
Height, cm
Head circumference, cm
Crown-rump length, cm
Body mass index, kg/m2
Gain weight, g/dGain height, cm/d
6 months’ corrected Age
Weight, kg
Height, cm
Head circumference, cm
Crown-rump length, cm
Body mass index, kg/m2
Gain weight, g/dGain height, cm/d
12 months’corrected Age
Weight, kg
Height, cm
Head circumference, cm
Crown-rump length, cm
Body mass index, kg/m2
Gain weight, g/dGain height, cm/d

Earlypreterm
infantLatepreterm
infantt, z or P
x2
value

6.64 ± 0.91
60.59 ± 2.45
40.21 ± 1.28
40.25 ± 1.72
18.08 ± 1.87
33.40 ± 6.08
0.13 ± 0.02

6.63 ± 0.79
61.52 ± 2.13
40.62 ± 1.70
40.43 ± 1.95
17.51 ± 1.45
34.65 ± 6.50
0.13 ± 0.02

0.061
2.651
1.747
0.641
2.28
-1.28
0.68

0.951
0.009
0.082
0.523
0.02
0.20
0.50

8.04 ± 1.01
66.63 ± 2.95
43.18 ± 1.63
43.10 ± 1.59
17.94 ± 1.91
15.57 ± 5.71
0.07 ± 0.02

8.08 ± 0.88
67.54 ± 2.19
43.64 ± 1.27
43.42 ± 1.77
17.71 ± 1.36
16.40 ± 4.54
0.07 ± 0.01

0.306
2.195
1.997
1.207
0.88
-1.07
-0.16

0.76
0.03
0.048
0.229
0.38
0.29
0.87

9.53 ± 1.20
75.03 ± 3.34
47.17 ± 2.09
45.80 ± 1.48
16.89 ± 1.53
8.19 ± 4.33
0.05 ± 0.01

9.77 ± 1.07
76.15 ± 2.80
47.68 ± 1.86
46.12 ± 1.25
16.82 ± 1.26
8.93 ± 2.98
0.05 ± 0.01

1.341
2.349
1.656
1.525
0.30
-1.35
-0.27

0.182
0.02
0.1
0.129
0.75
0.18
0.79

Table 3 Comparison of iron metabolism levels between the twogroups3
months’correctedAgechronologicalages, days
Hb, g/L
MCV, fl.
MCH, pg
MCHC, g/L
SI, umol/L
SF, ng/ml
TIBC, umol/L
TS, %
ID
6
months’correctedAgechronologicalages, days
Hb, g/L
MCV, fl.
MCH, pg
MCHC, g/L
SI, umol/L
SF, ng/ml
TIBC, umol/LTS, %log10 hepcidin,
ng/ml
ID
12
months’correctedAgechronologicalages, days
Hb, g/L
MCV, fl.
MCH, pg
MCHC, g/L
SI, umol/L
SF, ng/ml
TIBC, umol/LTS, %Early-preterm
infantLate-preterm
infantt, z or P
x2
value

151.92 ± 20.05

126.0 ± 11.38

9.82

0.001

117.11 ± 11.57
77.42 ± 5.53
26.00 ± 2.44
335.41 ± 13.41
11.36 ± 3.50
36.87 ± 31.57
18.68 ± 6.93
63.04 ± 11.77
6(11.3%)118.13 ± 10.42
77.70 ± 4.09
26.56 ± 1.64
340.22 ± 14.56
10.90 ± 3.36
65.78 ± 75.76
18.74 ± 7.26
60.49 ± 11.99
5(5.3%)

0.57
0.344
1.57
2.08
0.805
3.38
1.29
0.051
1.04

0.567
0.732
0.119
0.039
0.422
0.001
0.197
0.959
0.308

244.73 ± 23.41

215.11 ± 11.28

9.82

0.001

124.75 ± 9.10
76.75 ± 5.28
25.75 ± 2.90
332.75 ± 38.85
12.75 ± 4.13
32.75 ± 19.08
21.75 ± 8.36
59.75 ± 10.68
26.72(16.28,39.61)

122.75 ± 9.49
76.75 ± 3.98
28.75 ± 31.86
332.75 ± 34.96
12.75 ± 3.67
38.75 ± 34.52
21.75 ± 7.34
59.75 ± 10.72
28.29(16.58,54.04)

1.69
0.31
0.82
0.12
0.10
1.40
0.07
0.12
0.839

0.09
0.75
0.41
0.89
0.91
0.16
0.94
0.90
0.402

3(5.1%)

6(6.3%)

0.00

0.99

429.17 ± 26.71

401.62 ± 11.62

9.82

0.001

124.65 ± 17.01
78.80 ± 5.39
26.47 ± 2.31
330.32 ± 42.03
12.02 ± 4.04
38.87 ± 45.78
21.70 ± 7.57
55.74 ± 6.34

127.20 ± 8.17
78.64 ± 3.96
26.64 ± 1.84
338.60 ± 12.54
12.07 ± 3.88
33.63 ± 14.95
25.37 ± 24.14
54.59 ± 9.67

1.24
0.21
0.49
1.77
0.07
0.94
1.08
0.77

0.21
0.83
0.62
0.07
0.94
0.34
0.27
0.44

Data are presented as mean ± SD, range or number of patients (percentages).
Hb, hemoglobin; MCV, mean corpuscular volume; MCH, mean corpuscularhemoglobin; MCHC, mean corpuscular hemoglobin concentration; SI, serumiron; SF, ferritin; TS, transferrin saturation; TIBC, total iron binding capacity.
Hepcidin did not follow a normal distribution and was log-transformed. *Pvalue < 0.05 was considered to indicate statistical significance* Data are presented as mean ± SD. P value < 0.05 was considered to indicate
statistical significanceage. This could be linked to significant variations in ironreserves between preterm infants born at various gestational ages. Siddappa AM et al. synthesized and analyzed
multiple previous studies to plot ferritin concentration

Ruan et al. BMC Pediatrics(2024) 24:530

Page 7 of 10

Table 5 Comparison of iron metabolism and physical growthlevels between preterm infants in the ID and non-ID groups at 6
months of corrected age

Hb, g/L
MCV, fl.
MCH, pg
MCH, g/L
S, umol/LSF, ng/ml
TIB, umol/L
TS,%
Weight, kg
Height, cm
Head circumference,cm
Crown-rumplength, cm
log10
hepcidinGain weight,
g/d

ID

no ID

T/X2

116.56 ± 9.19
73.33 ± 3.01
23.18 ± 1.53
315.89 ± 11.17
7.58 ± 2.49
8.01 ± 4.48
11.02 ± 5.10
73.07 ± 15.27
8.54 ± 1.42
67.14 ± 3.45
43.31 ± 1.71

123.70 ± 9.19
76.55 ± 4.58
27.97 ± 26.78
332.88 ± 38.84
12.57 ± 3.77
37.71 ± 29.69
21.78 ± 7.51
58.26 ± 9.82
8.05 ± 0.87
67.23 ± 2.41
43.48 ± 1.39

2.26
2.08
0.54
1.31
3.92
10.31
4.23
4.23
1.04
0.10
0.34

P
value
0.03
0.04
0.59
0.19
0.00
0.00
0.00
0.00
0.33
0.92
0.73

43.84 ± 1.71

43.27 ± 1.66

1.01

0.31

6.36(3.15,30.69)

27.92(18.49,48.85)

2.43

0.025

16.20 ± 3.09

16.09 ± 5.20

0.05

0.95

* Data are presented as mean ± S. Hb, hemoglobin; MCV, mean corpuscularvolume; MCH, mean corpuscular hemoglobin; MCHC, mean corpuscularhemoglobin concentration; SI, serum iron; SF, ferritin; TS, transferrin saturation;
TIBC, total iron binding capacity. Hepcidin did not follow a normal distribution
and was log-transformed. *P value < 0.05 was considered to indicate statisticalsignificancecurves in babies aged 23 to 41 weeks and discoveredthat total body iron levels differed substantially among
preterm infants of different gestational ages [22]. Thisis because preterm infants’ postnatal iron storage is primarily provided by their mothers. 80% of the iron given
from mother to baby during pregnancy happens in
the late stages of pregnancy (25–37 gestational weeks)[23], and the older the preterm infant, the more iron isstored in the body. Consequently, compared to late preterm infants, early preterm infants have much less ironstored within their bodies. This is in line with our findings. We discovered that ferritin levels were considerably
lower in early preterm infants at corrected 3 months ofage compared to late preterm infants. This implies that,even with iron supplementation of 2–4 mg/kg/d, earlypreterm infants do not have as much iron as late preterm
infants by the time of the corrected gestational age of 3
months. According to the research conducted by Reinbott et al., ferritin levels in healthy Cambodian newborns
decreased from 50 µg/L to 15 µg/L between the ages of 3
and 6 months and then decreased at a slower rate (from15 µg to 14 µg/L) between the ages of 6 and 24 months[24]. This pattern is similar to the ferritin changes inlate preterm infants in our study. We know that in term
infants, iron stores fulfill metabolic requirements until
approximately 4–6 months of age. Therefore, ferritin
is significantly depleted and gradually declines in term
infants between 3 and 6 months. This change in ferritin
was not observed in early preterm infants, which may

Table 6 Correction of cross-sectional correlations of hepcidin, ferritin, and explanatory variables in preterm infants at 6 months ofcorrected age

Maternal age
IVF
Iron supplementation during pregnancy > 90 days
Anemicnumber of pregnancies
time of birth
Birth weight
Birth length
Timing of complementary food introduction
Feeding Methods
Regular iron supplementation
Transfuse blood
WeightHeightGain weight
Gain heightHbSF
log10 hepcidin

Hepcidin
P value
0.93
0.60
0.64
0.51
0.40
0.72
0.28
0.83
0.48
0.96
0.73
0.64
0.08
0.05
0.46
0.03
0.53
0.004

β
0.10
6.31
-5.88
-7.69
-3.48
3.22
-0.02
1.10
-5.89
0.36
-4.92
12.23
17.34
6.86
1.15
-795.900.45
0.63

ferritin
95% CIb
P value(-2.41, 2.62)
0.45(-18.31, 30.92)
0.74(-31.30, 19.55)
0.32(-31.25, 15.88)
0.10(-11.90, 4.94)
0.56(-15.20, 21.65)
0.28(-0.06, 0.02)
0.04(-9.19, 11.39)
0.90(-22.92, 11.14)
0.37(-13.00, 13.73)
0.26(-33.78, 23.93)
0.87(-40.74, 65.19)
0.78(-36.67, 1.99)
0.90(-0.04, 13.75)
0.17(-2.03, 4.33)
0.74(-1,501.80, -89.82)
0.14(-0.99, 1.88)
0.20(-0.21, 1.04)
0.004

β
0.77
-3.31
10.34
-0.03
2.03
-8.06
0.03
-0.52
6.22
-6.08
-2.02
-5.96
1.50
-4.07
-0.43
461.470.75

95% CIb(-1.31, 2.85)(-23.93, 17.32)(-10.58, 31.25)(-19.88, 19.81)(-5.06, 9.11)(-23.14, 7.02)(-0.00, 0.06)(-9.12, 8.08)(-7.92, 20.36)(-16.97, 4.81)(-26.16, 22.12)(-50.32, 38.39)(-15.66, 18.65)(-10.05, 1.92)(-3.11, 2.25)(-158.82, 1,081.77)(-0.42, 1.92)

0.44(-0.15, 0.72)*P value < 0.05 was considered to indicate statistical significance. Hepcidin did not follow a normal distribution and was log-transformed

Ruan et al. BMC Pediatrics(2024) 24:530

be because preterm infants have insufficient stored ironfrom birth, the ferritin in the body continues to be at alower level and there is not enough stored iron to be consumed. Therefore, we hypothesized that the iron status oflate preterm infants may be closer to that of term infants
and quite different from that of early preterm infants.
Our multifactorial regression analysis of the relationship between indicators of iron metabolism affecting preterm infants and ferritin revealed that birth weight had
a positive correlation with ferritin; the greater the birthweight was, the greater the ferritin level. This shows that
total body iron levels can change dramatically for preterm infants of different gestational ages and weights. The
World Health Organization recommends using a threshold of 12 µg/L to diagnose SF in infants under 2 years old[25]. However, these data are extrapolated from the dataof older children and have limited relevance for infants.
In a study of 285 low-birth-weight infants, Berglund S
et al. reported that ferritin concentrations in preterm
infants supplemented with 2 mg/kg/day of iron were significantly greater than those in preterm infants supplemented with 1 mg/kg/day of iron and no iron [19]. There
was also a positive correlation between ferritin concentration and iron supplementation dose (p < 0.001) [19].
Uniformly supplementing preterm infants with 2–4 mg/
kg/day iron may not be able to meet the early catch-up
growth needs of very early preterm infants, especiallyultra early preterm infants. This may also result in ironoverload and related side effects in late preterm infants
with heavier birth weights and older gestational ages
who are not at high risk for other iron deficiency factors. These issues stem from a lack of thorough researchon iron status in preterm infants, the numerous variables that affect iron stores in preterm infants, and thewide variation in total iron content in preterm infants.
This can cause iron overload and related adverse effects
in late preterm infants. Consequently, a more appropriate approach to iron supplementation in preterm infants
may be to evaluate an individualized iron supplementation regimen based on gestational age, birth weight, andferritin and Hb levels.
Hepcidin is a 25-amino-acid peptide produced byhepatocytes that is a critical regulator of iron release andstorage in tissues. It is primarily utilized to maintain astable supply of iron to erythrocytes and other tissues, aswell as to avoid the negative effects of iron excess [26].
At 4 months, infants with ID exhibited significantly lower
hepcidin levels at 1.5 months than infants without ID[11]. This finding implies that ID may serve as a marker
for the early identification of infants. In the diagnosis of
ID, a hepcidin concentration ≤ 6.895 ng/mL exhibited
79.2% sensitivity and 82.8% specificity. In line with previous research, we discovered that at corrected 6 monthsof age, hepcidin levels in preterm infants with ID were

Page 8 of 10

considerably lower than those in preterm infants without
ID (P = 0.025). Furthermore, hepcidin and ferritin showed
a positive correlation in our multifactorial regression
study of the relationship between iron metabolism indicators affecting preterm infants, suggesting that hepcidin
may also respond to stored iron. This finding is consistent with the results of Berglund S et al [27]. In 285 low
birthweight infants, Berglund S et al. reported that hepcidin levels were considerably lower in iron-deficient
infants and that hepcidin and ferritin were positively correlated [27]. In an investigation of 111 preterm infants
with gestational ages ranging from 32 to 36 + 6 weeks,
Uijterschout L et al. reported that the hepcidin levelsof preterm infants seemed to decrease in a pattern thatwas consistent with their ferritin levels [11]. Preterm
infants can acquire the iron they require to catch up withgrowth due to rapid variations in body iron status, highiron demand in the first year of life, and lower ferritin,which mobilizes stored iron and enhances dietary ironabsorption.
Our study revealed that early preterm infants were significantly shorter than late preterm infants at a correctedgestational age of 12 months. We found a negative correlation between hepcidin and height growth in preterm
infants based on our multivariate regression analysis of
the associations between hepcidin and indicators of ironmetabolism in preterm infants. The increase in heightmight be connected to the regulatory mechanism ofhepcidin. Hepcidin fluctuates throughout the day, peaking in the early morning and then gradually increasing
throughout the day, and hepcidin decreases following
growth hormone injection [28]. Growth factors restrict
hepcidin production in the liver by boosting erythropoiesis, which promotes increased intestinal iron absorption
and iron release from the reticuloendothelial system tomeet erythropoietic iron requirements [29]. Goodnough
JB et al. discovered that hepatic epidermal growth factor decreased hepcidin messenger RNA synthesis in part
via the PI3 kinase MEK/ERK kinase pathway, resulting in reduced hepcidin concentrations [30]. During thefirst four months of life, the concentration of hepcidin in
infants decreases in an attempt to fulfill the high growthrequirements of infants by increasing the amount ofavailable iron. Armitage AE et al. reported a significant
association between an increase in infant weight and adecrease in infant hepcidin [31]. Hepcidin inhibition andiron deficiency may be linked to the catch-up growthobserved in preterm infants during the first year of life, asiron is preferentially supplied for the synthesis of hemoglobin in the body rather than to other tissues, such asmuscle and brain. However, more research is necessary
to determine the exact relationship and mechanism ofaction involved.

Ruan et al. BMC Pediatrics(2024) 24:530

Our research has several limitations. First, as a medical
center in Southwest China, we have a greater proportionof preterm infants, particularly very preterm infants, andas a single-center study, we cannot completely exclude
bias caused by geography, medical procedures, and theenvironment. Furthermore, while we supplemented each
preterm infant with 2–4 mg/kg/day iron as directed by
the guidelines, it remains unclear whether the youngage, challenges with feeding, and family compliance ofpreterm infants will enable them to follow the doctor’sorders for iron supplementation. Third, we lack an accurate way to determine the birth reserves of iron in preterm infants of varying gestational ages, and to create atailored iron supplementation regimen for them.

Conclusion
There were substantial differences in iron levels acrosspreterm infants of varying gestational ages. Specifically,late preterm infants had an iron status that was significantly different from that of early preterm infants andcloser to that of term newborns. Postnatal iron supplementation at 2–4 mg/kg/d in preterm infants significantly decreases the incidence of ID; however, ironsupplementation based on a single standard may not fully
satisfy the need for early preterm infant catch-up growth
because of the multitude of factors affecting preterm
iron stores and the wide variation in total iron content
in preterm infants. For preterm infants, a tailored ironsupplementation plan based on growth, birth weight,
and gestational age may be a more suitable route for ironsupplementation. Although the current study revealed
that the postnatal iron status of early preterm infants differed from that of late preterm infants, our study was asingle-center clinical investigation, and current evidence
does not fully explain the pattern of iron metabolism inpreterm newborns. A large-sample, multicenter clinicalstudy is required to elucidate the causes of this variationand the precise mechanism of action.
Abbreviations
ID	Iron deficiency
IDA	Iron deficiency anemia
Hb	Hemoglobin
MCV	Mean corpuscular volume
MCH	Mean corpuscular hemoglobin
MCHC	Mean corpuscular hemoglobin concentration
SI	Serum ironSF	FerritinTS	Transferrin saturation
TIBC	Total iron binding capacity
Acknowledgements
The authors express their gratitude to everyone who contributed to this study.
Author contributions
RSF and YF: concept development, formal analysis, and first draft writing. LZY,
XF, and QD: investigation. LY and YSF: investigation and formal analysis. TZH:
data collection. All of the authors had full access to all of the data in the study
and were ultimately responsible for the decision to submit for publication.

Page 9 of 10

Funding
This article was funded by a grant from the 19H0546 A Real World Study of the
Efficacy and Safety of Iron Dextrose Oral Solution for the Prevention of Iron
Deficiency Anemia in Preterm Infants project (00402155A1033).
Data availability
Access to the study’s data will be granted to qualified investigators uponrequest by contacting the corresponding authors and receiving approval from
Sichuan University’s West China Second Hospital Ethics Committee.

Declarations
Ethics approval and consent to participate
The Ethics Review Committee of West China Second Hospital of Sichuan
University approved this study (Medical Research 2019 On Approval No.
(086)). Informed consent to participate in the study was obtained from theparticipants’ parents or legal guardians.
Consent for publication
Not applicable.
Competing interests
The authors declare no competing interests.
Received: 19 May 2024 / Accepted: 7 August 2024

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