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Archives of Disease in Childhood
Arch Dis Child. 2011 September 1; 96(9): 817–822.
Published online 2011 June 14. doi:  10.1136/adc.2010.185637
PMCID: PMC3155119

Increased protein-energy intake promotes anabolism in critically ill infants with viral bronchiolitis: a double-blind randomised controlled trial



The preservation of nutritional status and growth is an important aim in critically ill infants, but difficult to achieve due to the metabolic stress response and inadequate nutritional intake, leading to negative protein balance. This study investigated whether increasing protein and energy intakes can promote anabolism. The primary outcome was whole body protein balance, and the secondary outcome was first pass splanchnic phenylalanine extraction (SPEPhe).


This was a double-blind randomised controlled trial. Infants (n=18) admitted to the paediatric intensive care unit with respiratory failure due to viral bronchiolitis were randomised to continuous enteral feeding with protein and energy enriched formula (PE-formula) (n=8; 3.1±0.3 g protein/kg/24 h, 119±25 kcal/kg/24 h) or standard formula (S-formula) (n=10; 1.7±0.2 g protein/kg/24 h, 84±15 kcal/kg/24 h; equivalent to recommended intakes for healthy infants <6 months). A combined intravenous-enteral phenylalanine stable isotope protocol was used on day 5 after admission to determine whole body protein metabolism and SPEPhe.


Protein balance was significantly higher with PE-formula than with S-formula (PE-formula: 0.73±0.5 vs S-formula: 0.02±0.6 g/kg/24 h) resulting from significantly increased protein synthesis (PE-formula: 9.6±4.4, S-formula: 5.2±2.3 g/kg/24 h), despite significantly increased protein breakdown (PE-formula: 8.9±4.3, S-formula: 5.2±2.6 g/kg/24 h). SPEPhe was not statistically different between the two groups (PE-formula: 39.8±18.3%, S-formula: 52.4±13.6%).


Increasing protein and energy intakes promotes protein anabolism in critically ill infants in the first days after admission. Since this is an important target of nutritional support, increased protein and energy intakes should be preferred above standard intakes in these infants.

Dutch Trial Register number: NTR 515.


The preservation of nutritional status and growth is a specific aim in critically ill children, but difficult to achieve. This is due to a metabolic stress response with profound changes in protein metabolism leading to a negative protein balance and loss of lean body mass. Inadequate nutritional intake in the paediatric intensive care unit (PICU), often due to fluid restriction, further leads to protein and energy deficits, especially early after admission.1 Other factors that hinder adequate nutrition are impaired intracellular insulin signalling,2 impaired glucose uptake3 and reduced mitochondrial capacity during critical illness.4 These factors are probably the reason why protein-energy malnutrition is observed in 16–24% of critically ill children5 6 and is associated with adverse clinical outcome.79

What is already known on this topic

  • [triangle] Critical illness in children is associated with increased protein breakdown, negative protein balance and adverse clinical outcome.
  • [triangle] Inadequate nutritional support further leads to protein-energy malnutrition during admission to the paediatric intensive care unit.

What this study adds

  • [triangle] Protein anabolism in critically ill infants can be achieved in the first days after admission by increasing protein and energy intakes above reference levels.
  • [triangle] The higher protein balance resulted from stimulated protein synthesis exceeding the rate of concomitant stimulated protein breakdown.
  • [triangle] Increased protein and energy intakes are recommended in critically ill infants with viral bronchiolitis.

A common but threatening disease in infants is viral bronchiolitis, which in severe cases leads to respiratory failure with need for ventilatory support and PICU admission. Adequate nutritional support in these critically ill infants is important, with protein anabolism as goal. However, up to now common practice has been to use standard infant formulas to provide approximately 1.5 g protein/kg/day and 100 kcal/kg/day.

Increased protein intake with adequate energy provision promotes anabolism in preterm infants,1012 in neonates undergoing surgery13 and in children with burns14 and cystic fibrosis.15 In relation to these observations, it is important to note that protein synthesis is a high-energy consuming process16 and energy deficiency worsens nitrogen balance.17 18 Hence, to induce net protein anabolism, it is essential to provide an adequate energy intake. We therefore hypothesised that increasing protein and energy intakes would induce net protein anabolism in critically ill infants.

Stable isotope amino acid methods are used to determine net protein balance.19 During feeding, amino acids appearing in the circulation originate from protein breakdown and from the fraction of meal-derived amino acids that are not retained in the splanchnic area. Protein synthesis during feeding can be calculated from the disappearance of essential amino acids (EAAs) such as phenylalanine from the circulation, corrected for non-protein synthesis related disposal (eg, oxidation, hydroxylation). Therefore, all these factors need to be considered if whole body net protein anabolism during feeding is to be calculated.20 21 Splanchnic extraction (SPE) of meal-derived amino acids has not been reported before in critically ill children.

The present study was part of a larger study on the nutritional and metabolic effects of increased protein and energy intakes using a protein and energy enriched formula (PE-formula) compared with a standard infant formula (S-formula).22 In the present study we studied the efficacy of increased protein and energy intakes to promote protein anabolism and the underlying mechanisms by using intravenous-enteral phenylalanine/tyrosine stable isotope method protocol. The primary outcome measure was whole body protein balance (WbPBal) at day 5 after admission. SPE of phenylalanine was a secondary outcome measure. The 24 h nitrogen balance was used as alternative method to assess protein balance. To gain more insight into the role of separate amino acids in protein kinetics, correlations between plasma amino acid concentrations and protein metabolism were assessed.


Setting and patients

Infants admitted to the PICU of Maastricht University Medical Center (MUMC) or ErasmusMC-Sophia Children's Hospital (ErasmusMC) meeting the following inclusion criteria were enrolled: (1) respiratory failure due to viral bronchiolitis; (2) age 4 weeks to 12 months; (3) >40 weeks postmenstrual age; (4) ability to start enteral feeding <24 h after admission; (5) expected length of stay >96 h; and (6) venous and arterial catheters present. Exclusion criteria were as follows: (1) gastrointestinal, metabolic or chromosomal disorder; (2) parenteral nutrition other than intravenous dextrose; and (3) breast feeding. The inclusion and exclusion criteria were chosen to create a homogenous population of infants. Inclusion criteria 4, 5 and 6 were necessary for performance of the study protocol.

The Central Committee on Research Involving Human Subjects (CCMO, The Hague, The Netherlands) and local ethics committees approved this study. Written informed consent was obtained from parents or caregivers.

Anthropometric characteristics and severity of illness (Paediatric Risk of Mortality II)23 were assessed at inclusion. Duration of mechanical ventilation and length of PICU stay were noted. To determine the metabolic state of the patients, plasma amino acid concentrations were determined in arterial blood collected in the fed state at the start of the stable isotope protocol on day 5 using fully automated high-performance liquid chromatography as described before.24 The roles of specific amino acids were identified through correlation with whole body protein metabolism (WbPM).


Patients were randomised (randomisation and blinding as described before22) within 24 h after admission to receive continuous enteral feeding with PE-formula (Infatrini: 2.6 g protein/100 ml, 100 kcal/100 ml) or with S-formula (Nutrilon 1: 1.4 g protein/100 ml, 67 kcal/100 ml) both from Nutricia Advanced Medical Nutrition, Zoetermeer, The Netherlands. Compositions are summarised in appendix 1. Formulas were administered as previously described, starting 25.3±5.6 versus 23.4±5.4 h after PICU-admission in the PE-group and S-group, respectively.22 The ranges of protein and energy intakes on day 5 in the S-group (1.7±0.2 g protein/kg/24 h, 84±15 kcal/kg/24 h) covered recommended intakes for healthy infants <6 months (1.14–1.77 g protein/kg/24 h, 81–113 kcal/kg/24 h, depending on age in months).16 25 The ranges were significantly higher in the PE-group (3.1±0.3 g protein/kg/24 h, p<0.001; 119±25 kcal/kg/24 h, p<0.001) and were 175–272% and 105–147% of recommended intakes for protein and energy, respectively. Intake by volume was not significantly different between groups; 120.6±13.4 ml/kg/24 h in the PE-group versus 118.5±13.4 ml/kg/24 h in the S-group. As the target volume was 130 ml/kg/day, this was the maximum achievable intake for both groups for medical reasons (eg, fluid restriction) as decided by the treating physician. Details of nutritional intake are summarised in appendix 2.

Main outcome measures

WbPM and splanchnic phenylalanine extraction

On day 5 WbPM and splanchnic phenylalanine extraction (SPEPhe) were assessed by using a stable isotope protocol in the fed state. Several methods can be used to determine protein metabolism. We used the phenylalanine/tyrosine method because of the advantage that only blood samples are needed instead of both blood and breath samples as for methods based on leucine isotopes.26 In order to attain steady state, the infusion rate of enteral nutrition was not changed in the 6 h before the start of or during the stable isotope protocol. The stable isotope protocol was conducted by a research physician or research nurse. Intravenous amino acid tracers were administered continuously for 2 h with calibrated syringe pumps after a priming dose, using the following tracers, priming doses (µmol/kg) and infusion rates (µmol/kg/h), respectively: L-[ring-2H5]phenylalanine, 4.4 µmol/kg, 4.5 µmol/kg/h; L-[ring-2H2]tyrosine, 1.9 µmol/kg, 1.5 µmol/kg/h; L-[ring-2H4]tyrosine, 0.63 µmol/kg. For assessment of SPEPhe, L-[1-13C]-phenylalanine was administered as a primed-continuous enteral infusion (4.4 µmol/kg, 9.0 µmol/kg/h, respectively). Stable isotope tracers (>98% enriched) were purchased from Cambridge Isotope Laboratories (Woburn, Massachusetts, USA). Infusates were prepared by the centres' clinical pharmacists. Arterial blood was sampled (500 μl) before isotope infusion to determine background enrichment and at 60, 90 and 120 min of infusion to determine isotopic enrichment. Samples were put on ice and centrifuged (3500×g) for 10 min at 4°C. Plasma was deproteinised with 5% sulfosalicylic acid, frozen in liquid nitrogen and stored at −80°C until analysis. Tracer-to-tracee ratios (TTRs) were analysed using a liquid chromatography–mass spectrometry system as described before.27 TTRs were corrected for background enrichment and contribution to the measured TTRs of isotopomers with lower masses as described before.28 Isotopic enrichment reached a steady state after 1 h infusion, as shown by the lack of a statistically significant slope of calculated TTRs at 60, 90 and 120 min (data not shown). The mean enrichment was used for further calculations as described before.19 These calculations are explained in detail in appendix 3.

Nitrogen balance

The 24 h nitrogen balance on day 5 was assessed as described before, with urinary urea being converted to total urinary nitrogen (TUN) excretion.22

Statistical analysis

Power analysis was based on protein metabolism parameters in infants in earlier reports.29 To detect a 20% difference in protein balance between groups with 0.05 two-sided significance and 0.80 sensitivity, eight patients per group were required. Data were analysed on an intention-to-treat basis with the SPSS statistical software package (v 12.0; SPSS, Chicago, Illinois, USA). Differences between groups were assessed with Mann–Whitney U analysis. Correlations among parameters were tested with Spearman correlation coefficients. Statistical significance was defined as two-tailed p<0.05. Data are presented as mean±SD.



Twenty infants with respiratory failure due to viral bronchiolitis were enrolled (MUMC: n=10; Erasmus MC: n=10; December 2003 to February 2006). Ten patients were randomised and allocated to receive PE-formula and 10 to receive S-formula. All patients received the allocated formula. Two patients in the PE-group were lost to follow-up because vascular catheters were removed after extubation before day 5, and hence WbPM could not be measured. Patient characteristics are shown in table 1. Gestational age was significantly lower in PE-infants, but other parameters did not differ significantly. There were no significant differences in characteristics between patients enrolled in MUMC and in Erasmus MC (data not shown).

Table 1
Patient characteristics of the study population

Main outcome measures

WbPM and SPEPhe

The rates of phenylalanine kinetics on day 5 are shown in table 2. These values are directly derived from the phenylalanine and tyrosine stable isotope tracer results and subsequently used to calculate whole body protein kinetics as shown in figure 1. Whole body phenylalanine kinetics were significantly higher in the PE-group than in the S-group, apart from phenylalanine hydroxylation, which was higher in the PE-group but did not reach significance. Although SPEPhe (%) tended to be higher in the S-group than in the PE-group (p=0.08), absolute SPE was highest in the PE-group, but did not reach significance in either group.

Figure 1
Rates of protein kinetics (g/kg/24 h) in both study groups on day 5. Data are presented as mean±SD. *p<0.05. PE-group, protein and energy enriched formula fed group; S-group, standard formula fed group; WbPB, whole body protein breakdown; ...
Table 2
Whole body and splanchnic phenylalanine kinetics on day 5

Figure 1 depicts the rates of whole body protein synthesis (WbPS), whole body protein balance breakdown (WbPBal) and WbPBal in g/kg/24 h. It shows that WbPBal on day 5 was positive in the PE-group, while in the S-group it did not differ significantly from zero (0.73±0.5 vs 0.02±0.6 g/kg/24 h, p=0.026). The higher WbPBal was achieved through higher WbPS in the PE-group (9.6±4.4 vs 5.2±2.3 g/kg/24 h, p=0.019), despite concomitant higher WbPB (8.9±4.3 vs 5.2±2.6 g/kg/24 h, p=0.046). Negative WbPBal, reflecting catabolism, was found in one subject (13%) in the PE-group, but in four infants in the S-group (40%).

Whole body protein turnover in the PE-group was higher than in the S-group (10.7±4.3 vs 5.8±2.6 g/kg/24 h, p=0.012). Whole body protein oxidation, calculated from hydroxylation of phenylalanine to tyrosine, was higher with the PE-formula than with the S-formula, but not significantly so (1.2±0.8 vs 0.7±0.4 g/kg/24 h, p=0.25).

Plasma amino acid concentrations on day 5 are shown in appendix 4. The concentrations of five EAAs (methionine, histidine, phenylalanine, lysine and valine) and ornithine were significantly higher in the PE-group. The sums of branched chain amino acids (BCAAs) and EAAs were also significantly higher. WbPS was positively correlated with concentrations of the EAAs histidine (r=0.46, p<0.05), methionine (r=0.64, p<0.01), tryptophan (r=0.51, p<0.05), leucine (r=0.56, p<0.05) and isoleucine (r=0.47, p<0.05) and with sums of BCAAs (r=0.51, p<0.05) and EAAs (r=0.51, p<0.05). WbPBal was positively correlated with isoleucine (r=0.52, p<0.05), valine (r=0.46, p<0.05) and the sum of BCAA (r=0.53, p<0.05).

Nitrogen balance

The 24 h nitrogen balance on day 5 was significantly higher in PE-infants (274±127 vs 137±53 mg/kg/24 h, p<0.05). Multiplication of the results by 6.25 (the average amount of nitrogen in protein) resulted in protein balances of 1.71 vs 0.85 g/kg/24 h for the PE-group and S-group, respectively. TUN excretion on day 5, as a measure of amino acid oxidation, was higher in PE-infants, but not significantly so (171±81 vs 103±54 mg/kg/24 h, respectively, p=0.37).


The present study is the first to show that protein anabolism, an important target of nutritional support in critically ill infants, can be achieved within the first days after admission to the PICU by increasing enteral protein and energy intakes above dietary reference levels using a protein-energy enriched formula. This target was not achieved with a standard infant formula. The higher protein balance resulted from stimulated protein synthesis exceeding the rate of concomitant stimulated protein breakdown. Nitrogen balance data confirmed our phenylalanine results.

Our findings of increased protein synthesis and protein balance are in agreement with several studies in premature and term neonates evaluating the effects of amino acid supplementation.1013 2933 This is also true for protein breakdown which was either increased33 or not affected by amino acid supplementation.11 13 29 31 Although Poindexter30 has also reported suppression of proteolysis, this was in healthy instead of critically ill infants, receiving short term supplementation. Our finding of both increased protein synthesis and protein breakdown with higher protein and energy intakes is probably due to overall stimulation of protein turnover, as shown by the increased whole body protein turnover rate in the PE-group.34

Increased protein intake promotes protein anabolism, but may lead to increased amino acid oxidation with urea formation as seen in neonates with increasing amino acid supplementation,11 13 31 when exceeding needs. However, in the present study, neither phenylalanine hydroxylation nor TUN excretion (both reflecting amino acid oxidation), nor plasma urea concentrations (as described in our previous report)22 differed significantly between groups, suggesting that protein intake up to, and probably above, 3.1 g/kg/day does not exceed these infants' needs.

We are aware that using a PE-formula makes it difficult to discern the influences of separate macronutrients on protein metabolism. However, studies in adults and children have shown that protein is the major dietary determinant of WbPM as long as energy intake is sufficient.35 Additionally, supporting this hypothesis, the finding of a positive relationship between plasma EAAs and protein synthesis and balance suggests that EAA availability plays a crucial role in increasing protein synthesis and protein balance. It also agrees with previous observations in healthy adults indicating that (essential) amino acids are the primary stimulus for (muscle) protein synthesis.36

In these critically ill infants, receiving large amounts of intravenous fluids and medications, 120 ml/kg/day was the maximum achievable nutritional volume intake. Despite these fluid restrictions, an anabolic state was obtained within 5 days after admission using a protein-energy enriched formula, thereby limiting delay of growth and neurodevelopment during critical illness as much as possible. We have previously reported that the PE-formula is safe, well tolerated and improves nitrogen and energy balance at days 1–5 after admission.22 This type of formula is thus preferable to standard formulas to achieve adequate nutrition in comparable clinical settings. Since the subjects were a typical sample of infants with respiratory insufficiency due to viral bronchiolitis, we suggest that the results apply to the general population of these critically ill infants.

Our study is also the first to report values of first pass SPEPhe in continuously enterally fed critically ill infants. In this population, first pass SPEPhe did not differ between groups with an average of 46.8%. Comparable values have been described in healthy adults after a meal21 and in enterally fed piglets.37 There is discussion about correcting protein intake for SPE in calculations of WbPBal, since these retained amino acids are used for constitutive or secreted (glyco-)proteins in the gut,38 39 which is then considered part of WbPS. We have therefore also calculated the data without correction for SPE (not shown) and found that protein breakdown was 15–19% lower and protein balance 2.7–3.9 times higher. Only the absolute values are affected by this calculation, and the main conclusion of the study is not affected.

There are several limitations to this study. Despite using a randomised design, gestational age was significantly lower in the group receiving protein-energy enriched formula. This might have biased our results of protein metabolism as protein turnover decreases with increased (post-)conceptional age.40

Furthermore, the proportion of female subjects was relatively high. Protein deposition has been shown to be similar for healthy male and female children prior to adolescence and it is recommended that estimates of protein requirements for healthy children are calculated for both sexes combined.25 However, in children with burns (8 years of age on average), females had a less negative net muscle protein balance compared to males, and females gained lean body mass whereas males lost lean body mass. These differences were possibly due to the observed attenuated hypermetabolic response in females.41 Assuming that the same differences are true for critically ill infants, this would mean that the achievement of protein anabolism in the first days after admission in our study population could have been biased by the high proportion of females. However, gender differences in protein kinetics have not been described for critically ill infants. Moreover, our study population of infants with a viral infection is distinctly different from children with burns, who are subject to an extended hypermetabolic stress response with high inflammation.41 Also, when comparing the female with the male subjects within the PE- and S-groups of our study, the only notable difference was a non-significant trend towards higher protein turnover, synthesis and breakdown in the females compared to the males within the PE-group, but resulting in similar protein balances in both sexes. Therefore it seems unlikely that our results were affected by gender differences, despite the high proportion of females. Since the female subjects were equally distributed among both groups in our study, neither did it influence the comparison of groups.

Another limitation is that protein synthesis and protein breakdown were derived by extrapolating phenylalanine metabolism, which in fact only reflects the effects on the kinetics of this particular EAA. Other amino acid tracers may have shown different patterns, although the phenylalanine/tyrosine and leucine methods are considered to be reference methods to obtain reliable estimates of whole-body protein metabolism in most physiological conditions.26 The present study was not designed to establish exact protein and energy needs in critically ill infants. Neither was it adequately powered to detect clinical effects. Dose–response studies and studies into the clinical effects of improved protein balance in larger groups of critically ill infants are therefore necessary.

In conclusion, protein anabolism in critically ill infants with viral bronchiolitis can be achieved in the first days after admission by increasing protein and energy intakes above reference levels. Since protein anabolism is an important goal of nutritional support in these infants, increased protein and energy intakes should be preferred over standard intakes.


The authors would like to thank the participating children and their parents. They also thank Marianne Maliepaard for patient enrolment and data collection, and the nursing and medical staff of the paediatric intensive care units of Maastricht University Medical Center and ErasmusMC–Sophia Children's Hospital for their support.


Appendix 1

Macronutrient composition of the infant formulas used in this study

Average content per 100 mlPE-formulaS-formula
Energy (kcal)10067
Protein (g)2.61.4
 En% (protein-energy ratio)108
 Casein/whey (g)1.0/1.60.6/0.8
Amino acids
 L-alanine (mg)11755
 L-arginine (mg)8346
 L-aspartic acid/L-asparagine (mg)225120
 L-cystine (mg)3922
 L-glutamic acid/L-glutamine (mg)556260
 Glycine (mg)5227
 L-histidine (mg)6035
 L-isoleucine (mg)15974
 L-leucine (mg)278130
 L-lysine (mg)257120
 L-methionine (mg)6834
 L-phenylalanine (mg)11255
 L-proline (mg)198110
 L-serine (mg)15669
 L-threonine (mg)16973
 L-tryptophan (mg)3921
 L-tyrosine (mg)10744
 L-valine (mg)16982
Carbohydrates (g)10.37.5
Fat (g)5.43.5

PE-formula and S-formula from Nutricia Advanced Medical Nutrition, Zoetermeer, The Netherlands.

En%, energy percentage; PE-formula, protein and energy enriched formula; S-formula, standard infant formula.

Appendix 2

Rate of nutrient intake (per kg/24 h) during the stable isotope infusion protocol on day 5

PE-group (n=8)S-group (n=10)p Value
Volume (ml)120.6±13.4118.5±13.4NS
Energy (kcal)119±2584±15<0.001
 Energy as % of recommended intakes (81–113 kcal/kg/day)16105–14774–104
Protein (g)3.1±0.31.7±0.2<0.001
 Protein as % of recommended intakes (1.14–1.77 g/kg/day)25175–27296–149
 Intravenous (g)1.3±1.31.7±1.7NS
 Enteral (g)11.4±2.89.0±1.9<0.001
Fat (g)6.5±0.74.1±0.5<0.001

Data are presented as mean±SD. Intake per kg/24 h was calculated from the rate per kg/h during the stable isotope infusion protocol on day 5.

PE-group, protein and energy enriched formula fed group; S-group, standard formula fed group.

Appendix 3

Calculations of whole body protein metabolism with a combined intravenous-enteral phenylalanine stable isotope protocol

During steady state, the appearance (rate of appearance) of amino acids in the active metabolic amino acid pool is balanced by their disappearance. Appearance in fed subjects results from protein breakdown (PB) and partially from protein intake. Disappearance (rate of disappearance, Rd) results partially from the use of amino acids for protein synthesis (PS) and non-PS related disposal, eg, oxidation or hydroxylation. In the case of phenylalanine (Phe), non-PS related disposal resembles Phe hydroxylation (OH) to tyrosine (Tyr). So that protein breakdown during feeding can be calculated, the amount of amino acids originating from the meal and entering the circulation needs to be known. This is calculated from the fraction of amino acids that is extracted from the meal-derived amino acids in the splanchnic area (splanchnic extraction (SPE)).
Summarised for Phe (μmol/kg/h):
Whole body rate of appearance (WbRa) of Phe=Phe from PB+Phe intake×(1−SPEphe)=Phe for PS+Phe-OH.42
WbRa of the traced amino acid is calculated:
1. WbRa=tracer infusion rate/tracer-to-tracee ratio (TTR) in arterial plasma.
SPE and ASPE of phenylalanine (%) are calculated43:
2. SPEPhe=[1−(WbRa[2H5]Phe/WbRa[13C]Phe)]×100%where WbRa[2H5]Phe and WbRa[13C]Phe represent WbRa of Phe, calculated with intravenous [2H5]Phe or enteral [13C]Phe.
3. ASPEPhe=dietary intake of Phe×SPEPhe.
Further equations used (μmol/kg/h):
4. Phe OH to Tyr (OHPhe→Tyr)=WbRa[2H2]Tyr×(TTR[2H4]Tyr/TTR[2H5]Phe).
5. Phe coming from WbPB (Phe from PB)=WbRa[2H5]Phe−(Phe intake×(1−SPEPhe)).
6. Phe utilised for WbPS (Phe for PS)=WbRaPhe−OHPhe→Tyr.
7. Whole body Phe balance=Phe for PS−Phe from PB.

Whole body protein kinetics in g/kg/24 h (WbPB, WbPS and WbPBal) can be calculated by multiplying Phe kinetics (μmol/kg/h) using the results of calculations 5, 6 and 7, respectively, with the average Phe content in human protein, then multiplying by 24 h. We used 165.2 g/mol as the molecular weight of Phe and 280 μmol/g protein of Phe content in human protein.44

ASPE, absolute splanchnic extraction; ASPEPhe, absolute splanchnic phenylalanine extraction; SPEPhe, splanchnic phenylalanine extraction; WbPB, whole body protein breakdown; WbPBal, whole body protein balance; WbPS, whole body protein synthesis.

Appendix 4

Amino acid profile on day 5

Amino acid (mmol/l)PE-group (n=8)S-group (n=10)p ValueReference values (mmol/l)
Glutamine acid198±21191±20NS181–379
Sum of all AA3026±2262648±98NS
Essential AA1047±80817±42<0.05
Branched chain AA305±17252±9<0.001

Data are presented as mean±SEM.

*Essential AAs.
Branched chain AAs.

AA, amino acids; PE-group, protein and energy enriched formula fed group; S-group, standard formula fed group; Sum of all AA, sum of all measured AA.

Reference values are derived from Scott et al. Clin Chem 1990;36:1922–7.


Funding This study was financially supported by a grant from Nutricia Advanced Medical Nutrition, Zoetermeer, The Netherlands. Nutricia was not involved in the study design, in the collection, analysis and interpretation of data or in the decision to submit the paper.

Competing interests None.

Ethics approval This study was conducted with the approval of the Central Committee on Research Involving Human Subjects (CCMO, The Hague, The Netherlands) and the local ethics committees of Maastricht University Medical Center, Maastricht, The Netherlands and Erasmus Medical Center–Sophia Children's Hospital, Rotterdam, The Netherlands.

Provenance and peer review Not commissioned; externally peer reviewed.


1. Hulst JM, van Goudoever JB, Zimmermann LJ, et al. The effect of cumulative energy and protein deficiency on anthropometric parameters in a pediatric ICU population. Clin Nutr 2004;23:1381–9 [PubMed]
2. Sugita H, Kaneki M, Sugita M, et al. Burn injury impairs insulin-stimulated Akt/PKB activation in skeletal muscle. Am J Physiol Endocrinol Metab 2005;288:E585–91 [PubMed]
3. Vanhorebeek I, Langouche L. Molecular mechanisms behind clinical benefits of intensive insulin therapy during critical illness: glucose versus insulin. Best Pract Res Clin Anaesthesiol 2009;23:449–59 [PubMed]
4. Brealey D, Brand M, Hargreaves I, et al. Association between mitochondrial dysfunction and severity and outcome of septic shock. Lancet 2002;360:219–23 [PubMed]
5. Hulst J, Joosten K, Zimmermann L, et al. Malnutrition in critically ill children: from admission to 6 months after discharge. Clin Nutr 2004;23:223–32 [PubMed]
6. Pollack M. Nutritional support in the intensive care unit. : Suskind R, Suskind L, eds . Textbook of Pediatric Nutrition. Philadelphia, PA: WB Saunders; 1989:1118–25
7. Pollack M. Nutritional support of children in the intensive care unit. : Suskind R, Lewinter-Suskind L, eds. Textbook of Pediatric Nutrition. 2nd edition New York, NY: Raven Press; 1993:207–16
8. Pollack MM, Ruttimann UE, Wiley JS. Nutritional depletions in critically ill children: associations with physiologic instability and increased quantity of care. JPEN J Parenter Enteral Nutr 1985;9:309–13 [PubMed]
9. Briassoulis G, Zavras N, Hatzis T. Malnutrition, nutritional indices, and early enteral feeding in critically ill children. Nutrition 2001;17:548–57 [PubMed]
10. van Lingen RA, van Goudoever JB, Luijendijk IH, et al. Effects of early amino acid administration during total parenteral nutrition on protein metabolism in pre-term infants. Clin Sci 1992;82:199–203 [PubMed]
11. Rivera A, Jr, Bell EF, Bier DM. Effect of intravenous amino acids on protein metabolism of preterm infants during the first three days of life. Pediatr Res 1993;33:106–11 [PubMed]
12. te Braake FW, van den Akker CH, Wattimena DJ, et al. Amino acid administration to premature infants directly after birth. J Pediatr 2005;147:457–61 [PubMed]
13. Reynolds RM, Bass KD, Thureen PJ. Achieving positive protein balance in the immediate postoperative period in neonates undergoing abdominal surgery. J Pediatr 2008;152:63–7 [PubMed]
14. Alexander JW, MacMillan BG, Stinnett JD, et al. Beneficial effects of aggressive protein feeding in severely burned children. Ann Surg 1980;192:505–17 [PubMed]
15. Geukers VG, Oudshoorn JH, Taminiau JA, et al. Short-term protein intake and stimulation of protein synthesis in stunted children with cystic fibrosis. Am J Clin Nutr 2005;81:605–10 [PubMed]
16. Butte NF. Energy requirements of infants. Public Health Nutr 2005;8:953–67 [PubMed]
17. Calloway DH, Spector H. Nitrogen balance as related to caloric and protein intake in active young men. Am J Clin Nutr 1954;2:405–12 [PubMed]
18. Millward DJ. Macronutrient intakes as determinants of dietary protein and amino acid adequacy. J Nutr 2004;134(6 Suppl):1588S–96S [PubMed]
19. Engelen MP, Rutten EP, De Castro CL, et al. Altered interorgan response to feeding in patients with chronic obstructive pulmonary disease. Am J Clin Nutr 2005;82:366–72 [PubMed]
20. Soeters PB, de Blaauw I, van Acker BA, et al. In vivo inter-organ protein metabolism of the splanchnic region and muscle during trauma, cancer and enteral nutrition. Baillieres Clin Endocrinol Metab 1997;11:659–77 [PubMed]
21. Biolo G, Tessari P, Inchiostro S, et al. Leucine and phenylalanine kinetics during mixed meal ingestion: a multiple tracer approach. Am J Physiol 1992;262:E455–63 [PubMed]
22. van Waardenburg DA, de Betue CT, Goudoever JB, et al. Critically ill infants benefit from early administration of protein and energy-enriched formula: a randomized controlled trial. Clin Nutr 2009;28:249–55 [PubMed]
23. Pollack MM, Ruttimann UE, Getson PR. Pediatric risk of mortality (PRISM) score. Crit Care Med 1988;16:1110–16 [PubMed]
24. van Eijk HM, Rooyakkers DR, Deutz NE. Rapid routine determination of amino acids in plasma by high-performance liquid chromatography with a 2–3 microns Spherisorb ODS II column. J Chromatogr 1993;620:143–8 [PubMed]
25. WHO Technical Report Series Protein and Amino Acid Requirements in Human Nutrition – Protein and Amino Acid Requirements of Infants and Children. Geneva: WHO, 2002:161–84
26. Wagenmakers AJ. Tracers to investigate protein and amino acid metabolism in human subjects. Proc Nutr Soc 1999;58:987–1000 [PubMed]
27. van Eijk HM, Suylen DP, Dejong CH, et al. Measurement of amino acid isotope enrichment by liquid chromatography mass spectroscopy after derivatization with 9-fluorenylmethylchloroformate. J Chromatogr B Analyt Technol Biomed Life Sci 2007;856:48–56 [PubMed]
28. Vogt JA, Chapman TE, Wagner DA, et al. Determination of the isotope enrichment of one or a mixture of two stable labelled tracers of the same compound using the complete isotopomer distribution of an ion fragment; theory and application to in vivo human tracer studies. Biol Mass Spectrom 1993;22:600–12 [PubMed]
29. Poindexter BB, Karn CA, Leitch CA, et al. Amino acids do not suppress proteolysis in premature neonates. Am J Physiol Endocrinol Metab 2001;281:E472–8 [PubMed]
30. Poindexter BB, Karn CA, Ahlrichs JA, et al. Amino acids suppress proteolysis independent of insulin throughout the neonatal period. Am J Physiol 1997;272:E592–9 [PubMed]
31. van den Akker CH, te Braake FW, Wattimena DJ, et al. Effects of early amino acid administration on leucine and glucose kinetics in premature infants. Pediatr Res 2006;59:732–5 [PubMed]
32. Denne SC, Rossi EM, Kalhan SC. Leucine kinetics during feeding in normal newborns. Pediatr Res 1991;30:23–7 [PubMed]
33. Mitton SG, Garlick PJ. Changes in protein turnover after the introduction of parenteral nutrition in premature infants: comparison of breast milk and egg protein-based amino acid solutions. Pediatr Res 1992;32:447–54 [PubMed]
34. Wolfe RR, Goodenough RD, Burke JF, et al. Response of protein and urea kinetics in burn patients to different levels of protein intake. Ann Surg 1983;197:163–71 [PubMed]
35. Garlick PJ, McNurlan MA, Ballmer PE. Influence of dietary protein intake on whole-body protein turnover in humans. Diabetes Care 1991;14:1189–98 [PubMed]
36. Volpi E, Mittendorfer B, Rasmussen BB, et al. The response of muscle protein anabolism to combined hyperaminoacidemia and glucose-induced hyperinsulinemia is impaired in the elderly. J Clin Endocrinol Metab 2000;85:4481–90 [PMC free article] [PubMed]
37. Van Der Schoor SR, Reeds PJ, Stoll B, et al. The high metabolic cost of a functional gut. Gastroenterology 2002;123:1931–40 [PubMed]
38. van der Schoor SR, Wattimena DL, Huijmans J, et al. The gut takes nearly all: threonine kinetics in infants. Am J Clin Nutr 2007;86:1132–8 [PubMed]
39. van der Schoor SR, Schierbeek H, Bet PM, et al. Majority of dietary glutamine is utilized in first pass in preterm infants. Pediatr Res 2010;67:194–9 [PubMed]
40. Nissim I, Yudkoff M, Pereira G, et al. Effects of conceptual age and dietary intake on protein metabolism in premature infants. J Pediatr Gastroenterol Nutr 1983;2:507–16 [PubMed]
41. Jeschke MG, Mlcak RP, Finnerty CC, et al. Gender differences in pediatric burn patients: does it make a difference? Ann Surg 2008;248:126–36 [PubMed]
42. Tessari P, Barazzoni R, Zanetti M, et al. The role of substrates in the regulation of protein metabolism. Baillieres Clin Endocrinol Metab 1996;10:511–32 [PubMed]
43. Matthews DE, Marano MA, Campbell RG. Splanchnic bed utilization of leucine and phenylalanine in humans. Am J Physiol 1993;264:E109–18 [PubMed]
44. Castillo L, Yu YM, Marchini JS, et al. Phenylalanine and tyrosine kinetics in critically ill children with sepsis. Pediatr Res 1994;35:580–8 [PubMed]
45. Scott PH, Sandham S, Balmer SE, et al. Diet-related reference values for plasma amino acids in newborns measured by reversed-phase HPLC. Clin Chem 1990;36:1922–7 [PubMed]

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