Journal Pre-proof

Early vs late initiation of sodium glycerophosphate: Impact on hypophosphatemia in

preterm infants <32 weeks

Buse Ozer Bekmez, Serife Suna Oguz

PII: S0261-5614(21)00561-6
DOI: https://doi.org/10.1016/j.clnu.2021.12.011
Reference: YCLNU 5072

To appear in: Clinical Nutrition

Received Date: 21 September 2021

Revised Date: 27 November 2021
Accepted Date: 7 December 2021

Please cite this article as: Bekmez BO, Oguz SS, Early vs late initiation of sodium glycerophosphate:
Impact on hypophosphatemia in preterm infants <32 weeks, Clinical Nutrition, https://doi.org/10.1016/
j.clnu.2021.12.011.

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© 2021 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.
Early vs late initiation of sodium glycerophosphate: Impact on hypophosphatemia in

preterm infants <32 weeks

Buse Ozer Bekmez1, Serife Suna Oguz2

1 Sariyer Hamidiye Etfal Education and Research Hospital, Istanbul, Turkey
2 Ankara City Hospital, Ankara, Turkey

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Corresponding author: Buse Ozer Bekmez

Email address: ozerbuse@hotmail.com

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Phone number: +902123386300

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Kazim Karabekir Pasa, Bahcekoy Street No:62 34453 Sariyer/Istanbul

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Abstract
Background&Aim: Early electrolyte and mineral imbalances have emerged as a conspicuous
problem in very preterm babies since the revision of nutrition guidelines and the eventual
implementation of early aggressive parenteral nutrition (PN). We opted to carry out a study with
the introduction of phosphorus as sodium glycerophosphate in PN from the first day onward to
reveal the impact on serum phosphorus and calcium levels following the surge in the incidence
of hypercalcemia and hypophosphatemia.
Methods: In this single-center, prospective, observational cohort study, inborn babies <32
gestational weeks and <1500 g between August 2017 and July 2018 were enrolled consecutively.

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Infants born in the first 6-month of this period were initiated PN (Early phosphorus group)
containing phosphorus (1 mmol P as sodium glycerophosphate/100 ml PN) immediately after

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birth, and in the latter six-months, mineral-free standard PN (Control group) was commenced up

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until 48 hours of life. Parenteral nutritional prescriptions of both groups were similar in terms of
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macro and micronutrient intakes except for early phosphorus, calcium, and sodium. Serum
mineral and electrolyte levels were measured on Days 1-3-7 and compared between the groups.
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The primary outcome was the presence of hypophosphatemia in the first week of life. The
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secondary outcome was hypercalcemia, preterm morbidity, and mortality.

Results: A total of 261 infants were included in this study. There were 130 babies in Early
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phosphorus group and 131 in control group. Gestational ages (28.79±2.1 vs 28.46±2.2 weeks,
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respectively) and birth weights (1138±273 vs 1090±274 g, respectively) were similar in the
groups. Mean serum phosphorus levels were higher on all days in Early phosphorus group
(p<0.001). Early phosphorus group had a lower incidence of hypophosphatemia on days 1-3 and
7 (p<0.001). The percentage of hypercalcemic infants was significantly lower in Early
phosphorus group on day 3 (p<0.001). No difference was noted in terms of hypernatremia in the
groups.
Conclusions: Adding phosphorus to PN in the first hours of life reduced the frequency of
hypophosphatemia and hypercalcemia without any surge in hypernatremia or morbidity.
Nutrition guidelines need to be revised accordingly in terms of early mineral/electrolyte
supplementation.
Key words: Prematurity, hypophosphatemia, parenteral nutrition, sodium
glycerophosphate

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Introduction

Early parenteral nutrition (PN) is critical for very preterm infants to sustain a growth rate
similar to in utero while ensuring energy balance with prompt and sufficient macro and
micronutrient introduction (1). Extremely preterm babies are at particular risk of substantial
growth restriction in early life due to abrupt cessation of abundant intrauterine supply (2).
Continuation of the satisfactory catering of calcium and phosphorus via PN may ease meeting
the demands for optimal physiological functions and bone mineralization (3,4). Nevertheless, a

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decent amount of phosphorus supply during the early days seems unlikely given the limited
solubility and drawbacks of early electrolyte usage.

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High protein and glucose supply are implemented immediately after birth all over the

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world based on the recent nutrition guidelines since the beginning of the 20th century (5). This
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amelioration is known to maintain early anabolism while avoiding ‘metabolic shock’. Current
guidelines propose protein supply as 2.5-3.5 g/kg in very preterm babies (5). Phosphorus, an
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outstanding intracellular mineral, has a fundamental role in energy metabolism. The deficiency
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might end up with detrimental consequences namely respiratory and neuromuscular failure,
infections, hyperglycemia, and even death (6). To date, a bunch of studies has proclaimed early
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hypercalcemia and hypophosphatemia that was attributed to early aggressive PN in very preterm
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babies as opposed to the era of moderate protein provision (1,2,6,7-11). Insufficient phosphorus
supply not meeting the high anabolism triggered by aggressive PN, utilization of phosphorus in
intracellular processes only, and release of bone calcium to the circulation are hypothesized to
end up with hypercalcemia and hypophosphatemia (12). This was described as “refeeding
syndrome” by some researchers (6,9). Decent phosphorus supply is essential for growth and
bone mineralization as delineated in the phosphorus requirement calculation formula (9). Recent
ESPHGAN guidelines suggested 1.0-2.0 mmol/kg of phosphorus and 0.8-2.0 mmol/kg of
calcium supplementation in preterm infants during the first few days of life (4). Besides, the
authors proposed a Ca:P molar ratio below 1 (0.8-1.0) to reduce the incidence of neonatal
refeeding syndrome (4). We were not used to commencing phosphorus in our clinic up until 48
hours of life. However, we noticed a remarkable rise in the incidence of early hypercalcemia and
hypophosphatemia in our clinic similar to concurrent studies. Thus, we designed a prospective

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observational study with two consecutive epochs with or without early phosphorus and calcium
including PN initiated once the baby lost 2% of the birth weight and/or urine output was
0.5ml/kg/h in the last 6 hours to put forth the impact on early mineral imbalances.

Methods

Study center and population

This single-center prospective observational study was carried out in Zekai Tahir Burak

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Women’s Health and Education Research Hospital between August 2017-July 2018. Our clinic
is a fetal-neonatal intensive care center with 130 NICU incubator capacity and 18000 annual

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deliveries in Ankara, Turkey. This study was approved by the hospital’s ethics committee.

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Written or verbal informed consent was obtained from all parents.
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Infants with ≤32 gestational weeks and birth weight ≤1500 g consecutively enrolled in this
study. Exclusion criteria were <24 weeks of gestation, major chromosomal and congenital
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anomalies, death in the first 48 hours of life, prescription of individualized PN solutions, and
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lack of serum mineral/electrolyte values.

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Study design
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In the 1st six-month, a PN including sodium glycerophosphate in addition to calcium was

initiated in the shortest time following birth via an umbilical vein catheter once the baby lost 2%
of the birth weight and/or urine output was 0.5ml/kg/h in the last 6 hours and continued up until
48 hours (Early phosphorus group). Preterm babies were followed up closely regarding urine
output and postnatal weight loss. In the 2nd six-month, babies were started on mineral-free PN
(PN1A/1B) right after birth based on our unit’s policy and continued for 48 hours (Control
group). Ready-to-Use PN solutions have been implemented in our unit since 2010. We amend
and optimize our PN solutions regularly in the light of up-to-date ASPEN, ESPGHAN, and
Turkish Neonatology Society guidelines. Nine different standard PN solutions are available in
our pharmacy that are administered based on the postnatal age (PN 1 (0-48 hours)/PN 2 (48
hours -7 days)/PN 3 (>7 days)) and birth weight (PN 1A <1000 g/ PN 1B 1000-1500 g/ PN 1C
>1500g). These standardized PN solutions are prepared with either sodium glycerophosphate or

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potassium phosphate based upon the availability in the pharmacy unit. Ready to use PN solutions
were labeled in pharmacy revealing the type of the solution, however concealing the ingredients
of the PN during the study period. No mineral or electrolyte addition was performed except the
pharmacy unit.
In the 1st six-month, 100 ml PN 1a (80-100 ml/kg/d) comprised of 2.45 g amino acid
(Primene [BAXTER, Plattling, Germany]), 7.5 g dextrose, 0.5 g lipid (SMOFlipid [FRESENIUS
KABI AB. Uppsala, Sweden] 20% 2 g), 0 potassium, 1 mmol phosphorus (Glycophos®
[FRESENIUS KABI AB. Uppsala, Sweden]) calcium 0.8 mmol (calcium gluconate), sodium 2
mEq, and vitamin as Vitalipid (FRESENIUS KABI AB, Uppsala, Sweden). The only

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discrepancy of TPN 1b was 11 g dextrose. A molar Ca:P ratio of 0.8 was adjusted.
In the 2nd six-month; 2.45 g amino acid, 7.5 g dextrose, 0.5 g lipid and vitamin as Vitalipid

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were added to 100 ml TPN 1a. 100 ml TPN 1b was prepared with 2.45 g amino acid, 10 g
dextrose, 1 g lipid, and vitamins.
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The babies in both groups received TPN 2a (120 ml/kg/d) that was prepared with 2.82 g
amino acid, 7.5 g dextrose, 1.5 g lipid, 2 meq potassium, 4 meq sodium, 2 mmol calcium, 2
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mmol phosphate, vitamins. Ca:P molar ratio was adjusted as 1. The only adjustment in PN 2b
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was 11 g dextrose per 100 ml. Increments of PN were performed daily with dextrose in water
solution once required at the discretion of the neonatologist.
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All very preterm babies were initiated on 10-20 ml/kg minimal trophic enteral nutrition at
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the earliest as long as the mother’s milk was available with no contraindication. Trophic feeds
were initiated and sustained for at least 72-hours; 20-30 ml/kg/d increments were fulfilled
thereafter at the discretion of the attending neonatologist.

Study outcomes
The primary outcome was hypophosphatemia in the first week of life. Secondary outcomes
were hypercalcemia, electrolyte abnormalities, and preterm morbidity. Serum mineral and
electrolyte levels were checked at least once daily on postnatal days 1,3 and 7 in all preterm
infants who were on PN as per the unit’s protocol. Ionized calcium, sodium, and potassium
levels were monitored via daily/twice daily blood gases based upon the unit’s policy. In the final
analysis, serum electrolyte and mineral values in the first week of life were compared between
Group 1 and 2.

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 Hypophosphatemia was described as serum P concentration < 5 mg/dl (4,10,13),
hypocalcemia as total serum Ca < 7mg/dl and/or ionized Ca <1.1 mmol/L, and hypercalcemia as
total serum Ca >11 mg/dl and/or ionized Ca >1.4 mmol/L. On the other side, serum K < 3.5
mmol/L was defined as hypokalemia and >6.5 mmol/L as hyperkalemia. The definition of
hypernatremia and hyponatremia was >145 mmol/L and <135 mmol/L respectively.
Gestational age, birth weight, sex, delivery method, advanced resuscitation in the delivery
room, presence of intrauterine growth restriction (IUGR), and/or small for gestational age (SGA)
were recorded in all babies. Preterm morbidities included neonatal morbidities, namely
respiratory distress syndrome, duration of invasive and noninvasive ventilation,

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hemodynamically significant patent ductus arteriosus, grade 3-4 intraventricular hemorrhage,
pulmonary hemorrhage, necrotizing enterocolitis (>grade 2), and bronchopulmonary dysplasia

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(+glucocorticoid treatment). RDS was defined as the presence of respiratory distress and

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requirement for supplemental oxygen along with pathognomonic chest X-ray findings. A
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diagnosis of Patent Ductus Arteriosus (PDA) was assigned in the presence of a characteristic
heart murmur or clinical signs (hyperdynamic precordial impulse, full pulses, widened pulse
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pressure, and/or worsening of the respiratory status) with a ductal right-to-left shunt in the
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echocardiography. Papile classification was used for the diagnosis and grading of intraventricular
hemorrhage (14). Bell classification was used for the diagnosis and grading of NEC (15). The
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diagnosis of bronchopulmonary dysplasia was put based on the definition from the National
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Institutes of Child Health and Human Development Neonatal Research Network (16).

Statistics
Discrete variables were described as proportions and continuous variables as the mean or
median with their corresponding standard deviation or extreme values, respectively. We used
Shapiro-Wilk test to assess the normal distribution of the variables. While mean and standard
deviation was used to compare normally distributed continuous variables, median (min-max) was
used for the comparison of non-normal data. Percentages were preferred for summarizing

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categorical data. Independent Samples t Test was used for the comparison of the normally
distributed continuous variables. Continuous, not normally distributed variables were compared
via Mann-Whitney U test.
IBM SPSS Statistics 20 program was used. All tests were 2-tailed with statistical significance
defined as a P-value of <.05. Intention-to-treat analysis was used to manage the negative effects
of the dropout and missing data to be experienced in the study. Repeated measures ANOVA was
used to compare serum electrolyte and mineral values in the groups.
Sample size calculation was performed using GPower 3.1.9.2 program. The rate of
hypophosphatemia (defined as <5 mg/dl) was calculated as 28% in the first six-month epoch of

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our study. The estimated sample size would be at least 101 patients in each group (a total of 202)
with a two-sided alpha error of 0.05 and beta error of 0.2 (80% power) given the

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hypothesis/assumption of reduction of hypophosphatemia by 20%.

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Results
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482 inborn babies were admitted to the NICU in this period. 261 of them were
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consecutively enrolled in the final analysis (Figure 1). While Early phosphorus group included
130 babies, the remaining 131 were recruited in the control group. Demographic features are
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designated in Table 1. Mean gestational age (28.79±2.1 vs 28.46±2.2 weeks, respectively) and
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birth weight (1138±273 vs 1090±274 g, respectively) were similar in the groups. Mean time of
initiation of phosphorus comprising PN was 8±3.4 hours of life in Early phosphorus group.
Serum mineral and electrolyte levels are shown in Table 2. While mean serum phosphorus
levels were higher in Early phosphorus group on all days, the control group had significantly
lower serum calcium levels on 3 and 7 compared to Group 2 (p<0.001) (Figure 2-3). However,
no significant difference was noted in terms of mean serum sodium or potassium levels (p>0.05).
Table 3 is showing the percentage of mineral and electrolyte imbalances of the groups.
Early phosphorus group had a lower incidence of hypophosphatemia on days 1-3 and 7
(p<0.001). Despite the early provision of phosphorus, the high incidence of hypophosphatemia
and absence of hyperphosphatemia in Early phosphorus group was remarkable. The percentage
of hypercalcemic infants was significantly lower in Early phosphorus group on day 3 (p<0.001).
Due to the lack of clear recommendations for normal serum sodium range in preterm babies, we

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performed two separate statistical analyses for hypo- and hypernatremia. Whatever cut-off we
take into account, the incidence of hyponatremia was noted to be lower in Early phosphorus
group on day 3.
Figure 4 delineates the course of mean phosphorus levels of the groups regarding days.
The variation in serum phosphorus level based on time was not similar in the groups. Hence,
interaction term was found to be significant (G; F(1,228)= 152.894, <0.001). When serum
phosphorus levels were analyzed separately in the groups, a significant difference was present
between days 1, 3, and 7. Not only the drop of serum phosphorus level on day 3 was more
pronounced in the control group, but also the difference was noted to be statistically significant

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in both groups.
The percentage of serum sodium abnormalities of the whole cohort and the groups based

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on the days is represented in Figures 5 and 6. The high incidence of hyponatremia on day 3 in

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both groups and the whole cohort was notable. However, it was more pronounced in the control
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group.
The morbidity and mortality of the groups are displayed in Table 4. Duration of
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noninvasive ventilation, invasive ventilation requirement on day 28, corticosteroid treatment for
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bronchopulmonary dysplasia, and length of hospital stay were revealed to be lower in Early
phosphorus group (p<0.05). The mortality of the groups was similar.
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A negative correlation was noted between the duration of invasive ventilation and serum
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phosphorus levels on days 1, 3, and 7 (p<0.001). While serum phosphorus levels went down, the
duration of mechanical ventilation increased. In addition, length of hospital stay and serum
phosphorus level on day 7 were noted to have an inverse relationship (p<0.001).

Table 1. Demographic characteristics of the groups

Early phosphorus Control p
group group

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 (n=130) (n=131)

Gestational age 28.79±2.28 28.48±2.11 0.165

(weeks)a
Birth weight (g)a 1136.89±274.46 1092.73±273.79 0.148

Apgar 1a b 6 (0-7) 6 (0-7) 0.213

Apgar 5a b 8 (3-9) 8 (2-9) 0.130
Male, n % 64 (49.6) 66 (50.8) 0.892

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SGA, n % 17 (13.2) 28 (21.5) 0.173

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IUGR, n % 14(10.9) 18 (13.8) 0.464

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C/S, n % 105 (81.4) 105 (80.8) 0.898
Advanced 31(24) 36 (27.7) 0.501
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resuscitation in the
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delivery room, n %
Preeclampsia, n % 25 (19.4) 21 (16.2) 0.497
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ACS, n% 108 (83.7) 101 (77.7) 0.219

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PPROM, n % 24 (18.6) 25 (19.2) 0.898

Multiple pregnancy, n 29 (22.5) 42 (32.3) 0.076
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%
Chorioamnionitis, n % 16 (10.1) 19 (14.6) 0.267
a
Mean±SD
b
Median (min-max)
SGA:Small for gestational age IUGR:Intrauterine growth restriction C/S:Cesarean section ACS:Antenatal
corticosteroid PPROM:Preterm premature rupture of membranes

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Table 2. Serum mineral and electrolyte levels of the groups
Early phosphorus Control
group group p
(n=130) (n=131)
P, PD1 5.26±0.84 4.99±0.82 0.009
mg/dl PD3 4.75±0.83 3.18±0.82 0.000
PD7 5.57±0.67 4.64±0.64 0.000
Ca, PD1 8.05±0.49 8.14±0.43 0.121
mg/dl PD3 9.07±0.85 10.57±0.63 0.000

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PD7 10.06±0.62 10.60±0.35 0.000

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Na, PD1 136.98±2.54 137.32±2.77 0.296
mEq/L PD3 138.23±3.96 137.24±6.06 0.131
PD7 137.69±2.56
-p 138.28±2.27 0.073
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K, PD1 4.09±0.55 4.15±0.50 0.397
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mEq/L PD3 3.85±0.59 3.98±0.63 0.068

PD7 4.39±0.46 4.46±0.38 0.245
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P:Phosphorus. Na:Sodium. K:Potassium. Ca:Calcium PD:Postnatal day

All values are given as mean±SD
Independent Samples t test was used to obtain the P-value.
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 Table 3. Electrolyte and mineral imbalances of the groups

Postnatal period of transition

Day 1 Day 3

Early Control Early Control

phosphorus group p phosphorus group p

group group

Na 12.4 10 0.637 18.6* 38.5* <0.001

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<130mEq/L

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Na 26.4 27.7 0.412 21.2* 39.4* <0.001
<135mEq/L
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Na 0 0.4 0.576 11 12.7 0.312
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>145mEq/L
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K 8.5 5.3 0.572 20.1 12.3 0.854

<3.0mEq/L
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Ca 0 0 0.720 3.9* 33.3* <0.001

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>11mg/dl

P <5 mg/dl 28.7* 49.2* 0.006 52.7* 93.1* <0.001

P:Phosphorus. Na:Sodium. K:Potassium. Ca:Calcium

All values are given as percentage
Significant p values are highlighted
Statistically significant differences in the variables are highlighted and labelled as *.
Chi-square and Fisher exact test were used for the comparison of the categorical variables.

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Table 4. Morbidity and mortality of the groups
Early phosphorus Control p
group group
(n=130) (n=131)
RDS (surfactant 64 (49.6) 71 (54.6) 0.420
treatment), n %
Invasive ventilation in 53 (40.7) 66 (50.3) 0.131
the first 72 hours, n

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%

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IVH (grade 3-4), n % 17 (13.2) 21 (16.2) 0.767
PDA, n % 27 (20) 37 (28) 0.355
SIP, n % 8 (6.2)
-p 4 (3.1) 0.232
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NEC, n % 7 (5.3) 15 (11.5) 0.100
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Total NIV duration a b 4.99±7.59 8.64±11.10 0.015

3 (0-55) 4 (0-54)
Total MV duration a b
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4.84±10.53 8.86±17.11 0.051

1 (0-86) 2 (0-102)
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O2 requirement on PN 22 (16.9) 21 (16.1) 0.620

day 28, n %
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NIV on PN day 28, n 5 (3.8) 8 (6.1) 0.486

%
Intubation on PN day 4 (3) 16 (12.3) 0.009
28, n %
Corticosteroid for 11 (8.5) 22 (16.9) 0.043
BPD, n %
Length of hospital 33.18±26.58 46.01±36.94 0.004
28 (1-132) 36 (2-227)
stay a b, n %
Mortality, n % 32 (24.8) 27 (20.7) 0.439
Mean±SD
b
Median (min-max)

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RDS:Respiratory distress syndrome IVH:Intraventricular hemorrhage PDA:Patent ductus arteriosus
SIP:Spontaneous intestinal perforation NEC:Necrotising enterocolitis NIV:Noninvasive ventilatilation
MV:Mechanical ventilation PN:Postnatal BPD:Bronchopulmonary dysplasia

Discussion
This study proposes that early sodium glycerophosphate with a Ca:P molar ratio <1 during
the first 48 hours of life caters a way better mineral support by at least diminution of the rates of
hypercalcemia and hypophosphatemia. Besides, we reaffirmed the safety and requisite of sodium
supplementation in the first days of life without leading neither to an increment in the incidence

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of hypernatremia nor morbidity. Even, early sodium glycerophosphate provision culminated in

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less invasive ventilation on day 28 and eventual corticosteroid treatment for BPD.

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A series of cohort studies and case reports published significant electrolyte and mineral
imbalances, namely hypercalcemia and hypophosphatemia/hypokalemia, in preterm intrauterine
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growth restricted (IUGR) babies in early postnatal life after the implementation of high protein
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and glucose including PN at the beginning of the 2010s (2,6,10-11). Latter studies highlighted
this entity, seen not only in preterm IUGR but also in a notable proportion of the very preterm
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babies (1,7-9,12,17). Bonsante named this condition as Placental Incompletely Restored Feeding
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(PI-ReFeeding) Syndrome, which takes place due to inadequate supply of minerals and
electrolytes for the high anabolic rate that is led off by decent nitrogen and calorie introduction
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(9). Insulin secretion orients phosphorus into the cells and simultaneous scanty supply via PN
ends up with eventual hypophosphatemia which in turn triggers release from the bone (9).
Concomitant liberation of calcium with phosphorus culminates in hypercalcemia (9). Hence,
optimal and balanced provision of the minerals is highly critical for very preterm babies to
maintain mineral balance.
Based on the theoretical model developed by Bonsante for the estimation of phosphorus
need, a Ca:P molar ratio <1 (0.8-1.0) was suggested in the recent ESPGHAN nutrition guideline
to ameliorate the serum phosphorus levels and avoid the occurrence of hypercalcemia from the
first day on (4,9). The suggested parenteral calcium and phosphorus intake was revealed as 0.8-
2.0 and 1.0-2.0 mmol/kg/d during the first days of life (4). However, the optimal provision of
phosphorus in early days is challenging and limited due to the requirement for transport

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electrolytes, namely sodium or potassium, that is not favored by neonatologists given the
dilemma for early initiation. Nevertheless, emerging evidence has reported a considerably high
incidence of early hypokalemia and hyponatremia verifying the insufficient catering of
electrolytes as well during the early neonatal period (13-20).
There is a scant number of studies that were designed by early commencement of calcium
and phosphorus in preterm babies. In the first one, Lozano et al implemented an observational
study including 54 neonates <1250 g who received a 0.7 mmol phosphorus and 1.4 mmol sodium
as sodium glycerophosphate, 0.75 mmol Ca, and 0 potassium in 100 ml standard or tailored PN
immediately after birth (21). This study unveiled a noteworthy decline in the incidence of

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hypercalcemia and hypophosphatemia in the first week of life in comparison to a retrospective
cohort control. Besides, the safety of early sodium provision namely no surge in hypernatremia

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was remarkably similar to our study. However, the cut-off for hypophosphatemia was < 4 mg/dL

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in that study contrary to ESPHGAN suggestions that might have underestimated the actual
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incidence.
Hair et al reported a decrement in the incidence of both hypercalcemia and peak ionized
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calcium (mean iCa 1.64 0.27 to 1.50 0.23) via the commencement of calcium, and phosphorus in
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custom PN solutions with a molar ratio of 1/1 within the first day of life (1). However, the
precise amount of mineral supply was not noted in the study. Moreover, sodium
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glycerophosphate was utilized for a while in case of a shortage of potassium phosphate.

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Mulla et al designed a study with two epochs by different Ca/P molar ratios initiated in the
first day of life. When this ratio drops to 1:1 from 1.3-1.5, a decline in hypercalcemia and
hypophosphatemia was noted (7).
Senterre et al conducted a prospective cohort study with the implementation of electrolytes
and minerals from the first day onward via a standardized PN solution comprising 1.6 mmol
sodium, 1.5 mmol potassium, 1.8 mmol calcium, and 1.8 mmol phosphorus (Ca:P molar ratio of
1) (13). Despite this optimization accomplished, approximately one-third of the study group was
noted to have hypophosphatemia with a 12.7 % of hypercalcemia. Besides a considerable
amount of additional mineral and electrolyte intake, especially sodium, was provided. Of note,
hyponatremia incidence was substantially high by 30.4% notwithstanding the early introduction
of sodium in PN. This implies the safety of sodium and potassium intake by 1 mmol/kg/d in
extremely preterm babies on aggressive PN from the first day on. The limitation of this study

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was the absence of a control group and the plentiful amount of additional mineral/electrolyte
intake given during the study.
Australasian neonatal parenteral nutrition consensus recently updated their
recommendations. Bolisetty et al suggested the early introduction of calcium and phosphorus as
sodium glycerophosphate by 2.5 mmol of calcium, 1.5 mmol of phosphorus, 3 mmol sodium,
and 0 potassium in 100 ml standard PN (22). However, this infers a Ca:P molar ratio of 1.6
contrary to the suggestions of ESPGHAN and our study.
There are some theoretical models for the calculation of actual phosphorus needs. It has
been estimated that 1 g of protein accretion needs 0.3 mmol of phosphorus. Besides, phosphorus

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is required for bone mineralization that is calculated as calcium intake (mmol/kg)/1.67.
However, ideal adjustment of the precise need for the minerals is challenging due to an uncertain

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protein accretion rate. We preferred a ratio below 1 in the light of the above-mentioned studies in

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which the extremely preterm babies experienced a remarkable rate of hypercalcemia and
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hypophosphatemia despite a molar ratio of 1:1 to 1.5:1. Our results seem a way better but still
cannot meet the demands.
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Emerging evidence has been referring to earlier and a higher need for sodium intake than
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currently supplied, albeit the current ESPGHAN guideline suggesting the allowance for an early
negative sodium balance in the first couple of days due to the risk of increased morbidity
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(4,13,20). The lack of a calculation method for precise need, the discrepancy of the thresholds
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between the centers and even neonatologists, dilemma regarding the timing of sodium
introduction with the contradiction of hypernatremia and triggering morbidity via expansion of
the extracellular volume take part in the challenges of sodium supplementation. Hypernatremia
mostly originates from high water loss, not often due to excessive intake. A considerable number
of centers and physicians underestimate the actual rate of hyponatremia with the usage of cut-off
130. Birschoff et al highlighted the necessity of keeping sodium levels at 135-145 range in
preterm babies due to lack of evidence prompting a deviation from the standard range for the
adult population (23). Besides, the hyponatremia rate was considerably high in Group 1 despite
early catering of sodium indicating the higher requirement than supplied. A bunch of crucial
points should prompt current nutrition suggestions to be revised. Firstly, the high incidence of
hyponatremia in preterm infants in early life and the association of hyponatremia with adverse
neurodevelopmental outcomes are remarkable (24). Secondly, insufficient intake accounts for

14
growth restriction in preterm babies (23). Last but not least, delay of sodium/potassium
introduction defers phosphorus provision that ends up with hypophosphatemia and associated
morbidity.
The rate of invasive ventilation on day 28 was significantly lower in Group 1. In addition,
we revealed an inverse relationship between the duration of invasive ventilation and serum
phosphorus levels. Severe hypophosphatemia is well-known to end up with respiratory muscle
weakness leading to a higher need for mechanical ventilation and eventual chronic lung disease
(6). So, this finding is not surprising from the phosphorus point of view. Nevertheless, early
sodium supply has been a dilemma for the likelihood of triggering preterm morbidity including

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BPD (19). On the other hand, optimal growth has been attributed to adequate sodium intake and
some researchers have given a heads up for the treatments, e.g diuretics that reduce extracellular

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sodium (25). Thus, we speculated that early sodium and phosphorus provision might have taken

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a part in reducing the incidence of BPD even if it is hard to ensure.
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There are some limitations of this study. Firstly, it is a single-center observational study.
Secondly, we didn’t consider the sodium that a significant proportion of the babies received via
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antibiotics or other fluid management. Babies should have received more sodium than calculated
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in PN. Moreover, no calculation was performed for additional calcium or phosphorus intake.
However, this study is the first in terms of early introduced sodium glycerophosphate within the
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first day of life with a Ca:P molar ratio <1 via a ready-to-use PN with a prospective control
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group who received minerals and electrolytes after 48 hours of life. The provision of
micronutrients via a standardized PN at the same rate enabled a thorough and reliable
comparison of the groups. Our prospective control group received the same nutrient intake other
than calcium and phosphorus, sodium as well, during the first 48 hours. The lack of any surge in
hypernatremia or morbidity, even a drop in invasive ventilation rate on day 28, is an indicator of
the requirement for earlier supply. Even if we designed this study with 0.8 mmol phosphorus, 1.6
mmol sodium, and 1 mmol calcium in 100 ml PN, low blood values were substantially high
indicating still insufficient provision. One would expect superiority of tailored PN solutions
based on the specific needs of the baby, but recent studies have claimed the opposite. Moreover,
we consider the utilization of ready-to-use PN solutions as a superiority of this study. A
systematic review highlighted several advantages of standardized PN in neonates including

15
fewer bloodstream infections and prescription errors, longer shelf life, and last but not the least,
higher nutrient intake and weight gain (26).

Conclusion
Sodium glycerophosphate introduction in very preterm babies on the first day of life seems
effective in diminishing the incidence of hypercalcemia and hypophosphatemia as well as safe
and non-inferior to mineral/electrolyte-free PN in terms of electrolyte imbalances. The secondary
outcomes of this study are as substantial as the primary ones. We validated the favorable effect
of early phosphorus provision on reducing energy requiring morbidity such as the length of

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artificial ventilation and additional oxygen supply in very preterm infants. Randomized
controlled studies are required to reveal the optimal time and amount of mineral supply and Ca:P

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molar ratio that minimizes the risk and severity of hypophosphatemia and hypercalcemia.

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Funding statement:
The authors did not receive any specific grant from funding agencies in the public, commercial,
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or not-for-profit sectors.
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Conflict of interest:
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The authors have no potential conflict of interest.

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Acknowledgements:
The authors would like to thank Mehmet Buyuktiryaki and Nilufer Okur for their support to
patient collection.

Supporting information:
S1 STROBE Checklist. STROBE Statement—Checklist of items that should be included in
reports of cohort studies. STROBE, Strengthening the Reporting of Observational Studies in
Epidemiology.

16
Author statement:
Buse Ozer Bekmez: Conceptualization, Software, Investigation, Writing-Original draft
preparation
Serife Suna Oguz: Methodology, Formal Analysis, Supervision, Validation, Writing-Reviewing
and Editing

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Figure 1. STROBE flow chart. STROBE, Strengthening the Reporting of Observational Studies
in Epidemiology.

Assessed for eligibility

(n=482)

Excluded due to exclusion criteria (n=183)

-Gestational age <24 weeks
-Major chromosomal or congenital anomalies
-Severe perinatal asphyxia

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Fulfilled inclusion
criteria (n=299)

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Excluded due to missing values (n=38)
-Missing serum mineral/electrolyte levels

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Analyzed (n=261)
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Early phosphorus group Control group

(n=130) (n=131)
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Figure 2. Serum phosphorus levels of the groups according to days

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DOL: Day of life

Mean ± SD
Serum phosphorus level was higher in Group 1 on Day 1-3 and 7 (p<0.001). Independent samples t test was used.
Figure 3. Serum calcium levels of the groups according to days

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Serum Ca levels were similar on Day 1.

However, Group 1 had a significantly lower serum Ca levels on Day 3 and 7 (p<0.001). Repeated measures ANOVA
was used.
Figure 4. The course of serum phosphorus levels of the groups

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Mean ± SD
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DOL:Day of life
The variation in serum phosphorus levels based on days was not similar in the groups.
Not only the drop in serum phosphorus level on day 3 was more pronounced in the control group, but also the
difference was noted to be statistically significant in both groups. Repeated measures ANOVA was used.
Figure 5. The percentage of serum hypo-, normo- and hypernatremia in the whole cohort based
on the days

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DOL:Day of life
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Blue <130
Red 131-134
Green 135-145
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Black >145
The high incidence of hyponatremia on day 3 in the whole cohort was notable.
Chi-squared test was used.
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Figure 6. The percentage of serum hypo-, normo- and hypernatremia in the groups based on the
days

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Green- Early phosphorus group

Blue- Control group
Hyponatremia incidence was remarkably high on day 3. However, it was more pronounced in the control group.
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Chi-squared and Fisher exact test were used.

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