Receiving alimentation with cow milk–based formulas premature infants run a considerable risk of incipient late metabolic acidosis (ILMA) developing (1). This obligatory early stage during the development of manifest late metabolic acidosis is characterized by persistent maximum renal acid excretion (urine pH < 5.4), but normal or almost normal systemic acid–base status. Incipient late metabolic acidosis is accompanied by impaired weight gain, decreased nitrogen assimilation, and adaptive hormonal reactions (2,3). We investigated whether bone metabolism is involved in pathophysiologic mechanisms in premature infants with ILMA.
MATERIALS AND METHODS
Urinary ionograms were compared between 10 premature infants with a body weight of 1.0 kg to 1.9 kg on day 1 with ILMA (group IO) and 10 pair-matched (gestational age, actual body weight) premature infants without ILMA (group NO) participating in another study (4), both receiving full oral alimentation with the standard cow milk formula F. Moreover, in a prospective randomized study, alkali therapy was compared with sodium chloride (NaCl, equivalent molar load) supplement for 7 days each in 37 premature infants (body weight, 1.5–1.9 kg) with spontaneous development of ILMA while receiving formula F or receiving oral/parenteral nutrition (but without parenteral intake of calcium or phosphorus) (5). Patients with severe clinical disorders, such as cardiac defects, or those receiving medication that could influence acid–base homeostasis were excluded. Urine specimens were analyzed for calcium (Ca) and phosphorus (P) on day 1 before treatment and on day 7 of the treatment period in 20 patients (23 treatment periods) receiving alkali therapy (2 mmol · kg −1 · d −1 NaHCO 3 ; group IA) and 17 patients receiving NaCl supplement (2 mmol · kg −1 · d −1 NaCl; group IC).
The composition of the standard commercial formula F (Prematil®, Milupa, Friedrichsdorf, Germany) was energy, 75 kcal/dL (3.140 kJ/L); protein, 1.8 g/dL; Na, 31 mg/dL (13.5 mmol/L); K, 80 mg/dL (20.5 mmol/L); Cl, 53 mg/dL (14.9 mmol/L); Ca, 62 mg/dL (15.5 mmol/L); P, 36 mg/dL (11.6 mmol/L); and vitamin D 3 , 2μg/dL. To ensure an early start and a reasonable duration of Ca–P supplementation in the body weight class 1.5 kg to 1.9 kg (6), all premature infants with a birth weight less than 1.5 kg, irrespective of acid base metabolism, were given formula F with Ca–P supplement, and premature infants with a birth weight of 1.5 kg or more were randomly assigned to either formula F with or without Ca–P supplement, respectively (formula with Ca–P supplement [+Ca–P]: Ca, 87 mg/dL [21.8 mmol/L]; P, 43 mg/dL [13.9 mmol/L]). In all patients, vitamin D supplementation (25 μg/d) was started at the age of 10 days.
The acid–base status of the blood, serum parameters, and urinary concentrations of creatinine, Na, K, Mg, and Cl were assayed using standard methods. Ca in urine was determined by flame atomic absorption spectrometry. Ca in serum and P in urine and serum were measured photometrically. Urinary nitrogen (N) content was measured using a modification of the Kjeldahl method. The glomerular filtration rate (GFR) was determined as the clearance of endogenous creatinine. Intake of N, Ca, and P was calculated by use of the analytical data given by the manufacturer. Urinary concentrations of titratable acidity, ammonium (NH 4 ), and bicarbonate (HCO 3 ) were determined by the method of Lüthy et al. (7); organic acids were measured by titration, and sulfate was determined using a Dionex 10 Ion Chromatograph (Dionex GmbH, Idstein, Germany). Renal net acid excretion (NAE) corresponds to TA + NH 4 − HCO 3 .
Statistical evaluations were performed using nonparametric tests (one-way analysis of variance [Kruskal-Wallis] or Mann-Whitney U tests) (8), P values less than 0.05 were accepted as significant.
General care of the premature infants, nutrition, and sampling of blood (venous and capillary) and urine (spontaneous voiding), was conducted, according to the standard regimen of the ward. Informed parenteral consent was obtained in all patients with timed urine sampling and who were receiving therapy with either NaHCO 3 or NaCl.
Patients receiving full oral alimentation with the standard formula F and spontaneous development of ILMA (group IO) had a higher birth weight and were younger than the premature infants without ILMA (group NO), pair matched for actual body weight (Table 1). Patients with ILMA showed higher values of serum creatinine and NAE and a higher ratio of urinary N excretion to N intake. In patients with incomplete oral alimentation on the first day, ILMA spontaneously developed at about 2 weeks' actual age, and these patients showed no differences in birth weight; gestational age; actual body weight; intake volume; intake of Na, Ca, and P; serum parameters; and NAE at the time of random allocation to one of the treatment regimens (groups IA and IC). All three groups with ILMA (groups IO, IA, and IC) showed similar high values of serum creatinine and the same relation between urinary N excretion and N intake. The GFR indicated by clearance of endogenous creatinine showed a tendency to increase with postnatal age.
Analysis of urinary ionography from the two groups of premature infants (n = 10 each) receiving full oral nutrition with the same standard formula F (Table 2) shows that on the first day of spontaneously developed ILMA (group IO), infants demonstrated higher urinary values of P, sodium, titratable acidity, and a tendency to higher values of Ca and chloride compared with 10 pair-matched premature infants without ILMA (group NO). Despite maximum stimulation of renal acidifying mechanisms in patients of group IO, ammoniuria was not higher compared with patients of the NO group without ILMA.
On day 7 of the treatment period, patients of group IA receiving alkali therapy had lower urinary excretion of Na and Ca compared with patients of group IC receiving NaCl supplement (Table 3). Premature infants of group IC with NaCl supplement showed an increase in urinary excretion of Na and Ca from day 1 to day 7 of the treatment period.
Incipient late metabolic acidosis is characterized by an excess of nonvolatile acid load just surpassing the maximum capacity of renal acid excretion (1). Compensating mechanisms, such as volume contraction, hyperventilation, and chronic shifting of the renal bicarbonate threshold (the plasma bicarbonate level at which urine pH equals 6.1) to a higher plasma bicarbonate level (9), may mask ILMA at normal values of systemic blood acid–base status (1). Two groups of premature infants with ILMA showed an increase of either renal Ca or P excretion, respectively. We hypothetize that these data point to an impairment of bone mineralization even in this early stage of retention acidosis without an obvious shift in the systemic acid–base status.
Evidence suggests that bone is a major site of acid buffering during acute (6) and chronic (10) metabolic acidosis (11). Bone mineral provides an alkali pool protecting against acidemia by exchanging protons for sodium on the bone surface and by dissolving bone and thus releasing hydrogen buffers, such as hydroxyl ions, carbonate, and phosphate (12). During established metabolic acidosis, urinary Ca excretion is known to be increased, which is reversed during correction of the acidosis by alkali administration (13). In adults, increasing urinary net acid excretion was shown to be accompanied by an increased urinary Ca excretion (14). Moreover, in chronic renal failure, attrition of skeletal net (nonmetabolizable) base stores reduces actual renal net acid load (15).
In premature infants, mineral retention is improved by supplying sufficient mineral content and a well-balanced Ca/P ratio of the formula, thus avoiding side effects such as hypercalciuria or high renal NAE (16). The overall range of renal excretion of Ca and P was in agreement with earlier observations in premature infants (17) and indicates a sufficient, well-balanced nutritional supply of these minerals in our study group (18). In this study, the dietary regimens and mean oral and mean parenteral intake (and therfore rates of intake of protein, Ca, and P) were the same for the premature infants in groups IO and NO and premature infants in groups IA and IC. Premature patients with ILMA, however, showed a higher urinary excretion of Ca and P than did premature infants without ILMA.
In adults, oral ingestion of NH 4 Cl leads to a marked increase in urinary Ca excretion, whereas NaHCO 3 leads to a decrease in urinary Ca excretion (19), suggesting that bone is a sink for additional hydrogen ions (helping to maintain the extracellular fluid–bicarbonate content) and the source of additional urinary Ca. Expansion and contraction of the extracellular fluid volume are known to increase or decrease, respectively, urinary excretion of Ca and Na (20). In patients in group IC with NaCl supplement, urinary excretion of chloride and Ca was higher on day 7 of the treatment period compared with patients in group IA receiving alkali therapy, probably because of a trend toward metabolic acidosis indicated by decreased base excess values. In addition, volume contraction, known to be associated with ILMA, could be balanced by volume expansion caused by higher chloride intake. In the stage of ILMA, increased urinary Ca excretion may be caused by physicochemical dissolution of bone (21,22) and by a preponderance of the activity of osteoclasts (23,24) but could also be enhanced by the impairment of renal distal nephron Ca reabsorption in acidosis (20). The limited capacity for renal acid excretion in small premature infants and their high risk of spontaneous development of ILMA (3) may therefore also contribute to the high risk of nephrocalcinosis in very-low-birth-weight infants (25).
An increase or decrease in P intake induces a respective increase or decrease in urinary P excretion (20). Urinary excretion of P is increased by metabolic acidosis (26); moreover, chronic metabolic acidosis can impair renal phosphate reabsorption independently of parathyroid hormone (27). An increase in the alimentary supply of P may be attributed to the trend toward increased urinary P excretion from day 1 to day 7 of the study period in premature infants of the study groups IA and IC (Table 3). However, there were no significant differences on day 1 and day 7 of the treatment period in alimentary or parenteral intake of Ca or P between the groups IA and IC. Therefore, the tentatively lower increase of urinary P excretion in group IA patients on day 7 may be caused by increased Ca and P retention during alkali therapy.
Our data for GFR are consistent with data from the literature concerning the wide range of rates and slow postnatal increase of GFR values as well (28). Because GFR was not different between premature infants of groups IA versus IC and groups IO versus NO, the higher relation of urinary nitrogen excretion to nitrogen intake in patients with ILMA cannot solely be attributed to differences in dynamic development of renal function but may be assumed to indicate a lower rate of nitrogen assimilation in patients with ILMA (1).
Premature infants of group IA and IC with ILMA and NO without ILMA showed the same values of renal NAE (Table 1). The premature infants of group IA and IC therefore represent those premature infants with an especially low maximum renal capacity for renal hydrogen excretion. The premature infants with ILMA of the IO group were older than the premature infants of groups IA and IC; the higher renal NAE in the IO group thus points to an age-related increase of maximum renal acid excretion capacity. In premature infants of group IO with ILMA, ammoniuria was on the same level as in pair-matched premature patients of group NO without ILMA (Table 2). Thus, related to urine pH values, stimulation of ammoniuria seemed to be impaired in patients in group IO with spontaneous development of ILMA.
Alkali therapy on an individual basis was shown to be an effective therapy in premature infants with ILMA (29). In this study alkali therapy not only abolished ILMA in all treated premature infants (group IA) but also prevented the increase of urinary Ca and P excretion as indicated in patients with ILMA receiving NaCl supplement (group IC). Alkali therapy on a general preventive level was shown to be highly effective in preventing spontaneous development of ILMA in small premature infants with low GFR using a modified standard formula (30). Efforts to reduce the “potential renal acid load” of diets (9,31) may also be helpful in elderly patients with reduced GFR and mild chronic acidosis (32).
In summary, our data on urinary Ca and P excretion in premature infants with ILMA support the hypothesis that net bone alkali release is increased in these patients, probably preserving acid–base balance at the expense of bone mineral metabolism.
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