Enteral L Citrulline Supplementation in Preterm Infants – Safety, Efficacy and Dosing

Background: Citrulline (CIT) is a semi-essential amino acid in preterm infants (PI). Through enzymatic reactions in the urea cycle, CIT is converted to arginine (ARG) and serves as a precursor for endogenous synthesis of nitric oxide (NO), a known potent vasodilator that regulates blood supply to the lungs, intestines and other organs. Lower levels of ARG and CIT have been reported in infants with NEC, bronchopulmonary dysplasia associated pulmonary hypertension (BPD-PH) and sepsis. We hypothesized that enteral CIT supplementation is safe in PI and increases plasma CIT and ARG levels in PI. We speculate that the higher plasma ARG levels in turn will result in increased endogenous NO production, conferring potential benefits in BPD-PH and NEC.

Methods: A pilot, double blinded, randomized controlled trial (NCT03649932) of 42 PI ( < 33 weeks) was conducted. Subjects were randomized to receive low, medium and high dose CIT (100, 200 and 300 mg/kg/day divided Q12H) when they were on full feeds. The study drug was administered twice daily as a 10% solution via gavage feeding. Adverse events such as hypotension were monitored for all subjects. Blood samples were collected prior to starting enteral CIT (Day 0) and after completing 7 days (Day 7). Plasma CIT and ARG levels were measured by liquid chromatography mass spectrometry. Demographic and clinical data was collected from the electronic medical record. Data analysis was performed with SPSS 24.0. Paired t-test was used to compare pre- and post-supplementation plasma CIT and ARG levels.

Results: 42 PI were recruited with a mean +- SD gestational age of 29.4 + 2 weeks and birth weight 1275 +- 388g, of which 88% were delivered via C-section and 64% were female. All subjects tolerated enteral supplementation of CIT without side effects. Enteral CIT supplementation for 7 days significantly increased plasma CIT levels in all groups with a mean increase of 88 micromol/L (95% CI 17-158, p= 0.022) in low dose, a mean increase of 117 micromol/L (95% CI 60-175, p= 0.001) in medium dose and a mean increase of 144 micromol/L (95% CI 51-238, p= 0.006) in the high dose group (Figure 1). Similarly, ARG levels increased in the medium and high dose groups with a mean increase of 77 micromol/L (95% CI 2-153, p = 0.046) in medium dose group and mean increase of 103 (95% CI 24-181, p = 0.020) in the high dose group (Figure 2).

Conclusion: Enteral CIT supplementation in PI is feasible, safe, well-tolerated and effective in increasing plasma CIT and ARG levels at medium or high doses. The authors plan to conduct a randomized placebo-controlled trial using the medium dose (200 mg/kg/day) to evaluate the role of enteral CIT supplementation in BPD-PH.