Inhalation is the preferred route for treatment of respiratory disorders, also in infants, as it enables high drug concentrations to be achieved at the target organ with a minimum risk of systemic toxicity. However, when nebulisation is used, only a small proportion of the total dose reaches the lungs, while the remainder is swallowed, wasted into the atmosphere, or remains in the nebuliser device. In particular, in neonates and infants age-related characteristics must be considered which, compared with adults, entail a major reduction of lung deposition: (a) their breathing parameters reduce the amount of the nebulised dose that could be inhaled1 2; (b) in distressed and crying infants, the inhaled dose diminishes in addition and a great proportion of the inhaled dose is deposited in the upper airways3 4; (c) special anatomical and physiological considerations such as nasal breathing, a smaller pharynx, and a smaller airway calibre reduce the amount of aerosol reaching the lower airways and favour a more central pattern of deposition. Therefore, fine aerosol droplets (<2 μm in diameter) are more likely to reach the lower airways of young infants.5

Only a few studies have been reported in which the deposition of aerosol in young infants was measured in vivo. The most direct way of measurement is to quantify the dose and distribution of an inhaled drug using radiolabelled aerosols. However, γ-scintigraphic methods, particularly for premature infants, can hardly be justified from an ethical viewpoint.6 7 The filter technique, which is also employed to measure inhaled mass, only estimates the total dose delivered to patients, and allows no conclusion to be reached about the amount that reaches the lower airways.8 9 Measurement of response to β-agonist inhalation (lung function, heart rate) has often been used in infants and young children to compare the clinical efficacy of delivery systems.10^–12 However, the drug effect measured did not represent the deposition in the lower respiratory tract only. Yet, β-agonists deposited on the upper airways or swallowed are almost completely absorbed into systemic circulation; hence, this effect too needs to be taken into account.12 13

An established method adapted to evaluate relative lung deposition (relative bioavailability) involves inhalation of sodium cromoglycate (SCG) as a marker substance.3 14^–17 SCG, a drug that has a wide margin of safety, is used for prophylactic management of asthma. Even in premature newborns after long-term prophylactic administration of SCG no adverse effects were noted.18 SCG is a lipid-insoluble substance, which is well absorbed by the lungs, but the fraction impacted on the upper airways is swallowed and poorly absorbed (<1%) by the gastrointestinal tract.19 SCG is not metabolised and is rapidly eliminated in both urine and bile in approximately equal proportions. Urinary excretion can be used to indicate the amount of drug reaching the respiratory tract and, thus, determine the relative bioavailability to the lungs after inhalation.19^–21 It was the purpose of the present investigation to compare the relative lung deposition in premature infants after inhalation with three different nebuliser devices, under conditions relevant to clinical practice, by measuring urinary excretion of the marker substance, SCG.

PATIENTS AND METHODS

Patients

Spontaneously breathing, oxygen-independent preterm infants were consecutively recruited at the intensive care unit of the Magdeburg University Children’s Hospital. Of 17 preterm infants enrolled in the study, 13 had required mechanical ventilation or continuous positive airway pressure ventilation, or both. None of these infants had developed chronic lung disease of prematurity.22 Boys only were chosen for the study because it was easier to collect urine samples from them. Table 1 presents the main characteristics of the study group. Assuming a tidal volume of 6 ml/kg23 and a respiratory rate of 50/min,24 the mean minute ventilation calculated with reference to the body weight (table 1) was 660 (55) ml/min.

Devices

The test inhalations were carried out with the following devices: LC Star jet nebuliser and PARI Master compressor (Pari, Starnberg, Germany), LS 290 ultrasonic nebuliser (System, Villeneuve sur Lot, France), and Projet ultrasonic nebuliser (Artsana, Grandate, Italy). On the LC Star device, the aerosol is moved out of the nebuliser by the compressor-induced jet flow. The two ultrasonic nebulisers have additional blower units to generate the flow required to discharge the produced aerosol from the devices. On the Projet device, the blower flow rate is adjustable, and for the test inhalations it was set to the minimum. On the LS 290, by contrast, the aerosol output rate is adjustable, and it was set to level 1 (minimum). As all test inhalations were performed at the bedside, a 0.6 m corrugated tube was connected between the relevant nebuliser and the face mask (Baby mask, size 0; Pari). Table 2 lists the aerosol characteristics of the nebulisers as measured under conditions of test inhalations. Mass median diameters of aerosols were measured at the “far” end of the corrugated tube, using a laser diffraction particle size analyser (Malvern MastersizerX; Malvern Instruments, Herrenberg, Germany).

Study design

The inhalation study was designed as a randomised three-period, crossover trial. Within a period of 1 week the preterm infants inhaled in a random order using the three different nebulisers, including a 1-day washout period between the test inhalations.

The solution used for inhalation contained 20 mg SCG (2 ml of 1% Intal nebuliser solution; Aventis Pharma, Bad Soden, Germany).

The test inhalations were only started while the infants were in supine position and settled or sleeping. The face mask was held to the infant’s face, covering both mouth and nose. Inhalation was completed as soon as the nebulised mist was no longer visible to the naked eye. The duration of inhalation was recorded and the SCG residue in the nebuliser was measured.

Distress scoring

Levels of infant distress during inhalation were scored as follows: 1, settled (regular tidal breathing pattern, regular respiratory rate); 2, slightly unsettled (irregular breathing pattern, raised respiratory rate); and 3, moderately unsettled (slight defensive movements and/or mask lost contact with infant’s face for <10 seconds).

Collection of urine

Serial urine samples (4–7 fractions) were collected into urine bags (Urinocol; B. Braun Medicare, Melsungen, Germany) until 11–14 hours after inhalations. This period proved to be sufficient for determining the total amount of SCG excreted.14 15 25

The collection bags were checked at frequent intervals and drained as soon as nurses noted that urine had passed. Time and urine volumes were recorded. The particular times of collection bag draining were set as the end of the successive collection periods since the actual moment of last urination could not be accurately determined. Leakage from the collection bag was measured by recording the change in weight of the preweighed nappy. Data of test inhalations were excluded from evaluation if leakage occurred (LC Star, n = 2; LS 290, n = 3; and Projet, n = 4). Completeness of collected urine was additionally checked by measuring the urinary creatinine excretion during total collection periods.

Measurement of SCG

Aliquots of urine samples were frozen at −20°C until SCG analysis by high-performance liquid chromatography, as described elsewhere.14 The portion of the SCG dose retained in the nebuliser after inhalation was rinsed out with water and analysed after dilution, as appropriate.

Statistical analysis

Statistical evaluation was made by using SPSS for MS Windows, release 13.0. Normal distribution of the variables was proved using the Kolmogorov–Smirnov test. Data from inhalation with differing nebulisers (amount of SCG and its influencing factors) were compared using repeated measures analysis of variance followed by Bonferroni test for pairwise comparisons. For the distress score, the Friedman test was used instead.

Correlation between two variables was determined by the Pearson coefficient of correlation. The effect of the distress score on the amount of SCG was tested by one-way analysis of variance. For all evaluations, the level of significance accepted was p = 0.05. Results are given as mean (SD). An additional post hoc power calculation was conducted to test the differences of effect between nebulisers using the t-test with Bonferroni-adjusted α.

Ethical approval

The study protocol was approved by the human ethics committee of the Otto von Guericke University’s Medical Faculty, and written informed consent was obtained from all parents.

RESULTS

The mean (SD) percentage of cumulative urinary excretion of SCG (referred to the total amount measured) 4–6 hours after inhalation with LC Star, LS 290 and Projet was as high as 66 (13)%, 69 (14)% and 67 (13)%, respectively. After 10–12 hours, excretion of SCG was almost complete, being respectively 93 (6)%, 90 (6)% and 89 (8)% of the total amount measured. This confirms that the total collection period of 11–14 hours for the infants investigated was sufficient to record completely the excreted SCG in urine after inhalation.

The highest total amounts of SCG excreted in urine were measured after inhalation with the LC Star jet nebuliser (0.089 (0.036) mg, n = 15). Using LS 290 and Projet ultrasonic nebulisers we found 0.055 (0.019) mg (n = 14) and 0.046 (0.025) mg (n = 13), respectively (considering all inhalations with each nebuliser). The overall test of complete data (n = 10) showed a significant difference (p<0.001). Pairwise comparisons demonstrated that the amount of SCG using the LC Star nebuliser was significantly higher than after using the LS 290 (p<0.001) or the Projet nebuliser (p = 0.001) for the test inhalation (fig 1).

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Figure 1 Individual amounts of sodium cromoglycate (SCG) recovered in urine of 10 premature infants during the total collection periods (about 12 hours after inhalation) after inhalation of 20 mg SCG via LC Star (0.087 (0.028) mg), LS 290 (0.056 (0.020) mg) and Projet (0.042 (0.021) mg), overall p<0.001. Each line shows the values for a single patient. Data from infants with one (x) or two (+) missing values are additionally shown, identified by patient No (see table 1).

The infants tolerated the bedside inhalations with all nebulisers well. Their behaviour was rated in 35 (83%) of the 42 inhalations with distress score 1 (settled), in 10% with distress score 2 (slightly unsettled), and in 7% with distress score 3 (moderately unsettled). No statistically significant difference in distress scores was noted between the various nebulisers and no significant difference of SCG excreted in urine was seen as a function of the distress scores.

The study group was homogeneous with infants of similar post-conceptional age, weight and length at the time of test inhalations (table 1). No statistically significant differences between nebulisers were observed and no significant correlations were found between these parameters and the amount of SCG excreted in urine.

The duration of inhalation was significantly different between all nebulisers, being the longest after inhalation with the LS 290 (12.4 (2.7) minutes) and becoming shorter with the LC Star (9.4 (1.2) minutes) and the Projet (7.0 (1.8) minutes) (table 3). The amount of SCG retained in each nebuliser after test inhalation was very similar in LC Star (9.1 (0.8) mg) and LS 290 (9.2 (0.8) mg), but significantly higher in Projet (13.1 (1.3) mg; p<0.001) (table 3).

DISCUSSION

This study was conducted to compare the relative lung deposition after inhalation with three nebuliser systems in premature infants by measuring the cumulative urine excretion of the marker substance, SCG. Under conditions that are relevant to clinical practice the highest lung deposition was found after inhalation with the LC Star jet nebuliser.

When employing aerosol therapy in young infants fine aerosol droplets are more likely to reach the lower airways.5 Since the LC Star produces a considerably greater proportion of droplets <2 μm than the two other nebulisers, this may explain why in our study the highest lung deposition was measured after inhalation with the LC Star (table 2). We noticed that (a) the highest SCG residue remained in the device (table 3) and (b) the highest aerosol deposition in the corrugated tube while using the Projet nebuliser (table 2). This great loss of nominal dose obviously leads to poor lung deposition after inhalation with the Projet nebuliser.

The low tidal inspiratory flow rates in young infants compared with nebuliser flow rates indicate that a major part of the aerosol delivered to the face mask is lost into the environment.1 At a minute ventilation of 0.66 l/min as calculated for this study group and the flow rates of the LC Star and LS 290 of 7.55 l/min and 5.04 l/min, respectively, the respective average inhaled part could be at most 9% and 13% of the effective output of the nebulisers. For the Projet only, provision is made to permit the flow rate of the ventilator to be reduced. At the minimum setting (1.40 l/min), the proportion that can be theoretically inhaled is 47% of the effective outputs, thus being markedly higher than observed for the other two nebulisers. On the other hand, this low flow rate seems to account for the high dose loss caused by deposition of the aerosol in the corrugated tube (53% of the total output rate).

In several investigations, it was found that in distressed and crying infants delivery of drug to the lungs is greatly reduced.3 4 8 Therefore, in our study, we tried to eliminate this confounding factor for intraindividual comparison of nebulisers as far as possible. In this study only a few infants were slightly to moderately unsettled during bedside inhalation and the distress scores were not significantly different between inhalations with the various nebulisers tested.

The relative lung deposition after inhalation can be clearly evaluated from the SCG urine excretion in an intraindividual comparison. An accurate calculation of the absolute value of lung deposition would require additional measurement after intravenous SCG administration. In healthy adults it was determined that about 50% of the SCG dose reaching the systemic circulation is excreted in urine.26 27 Assuming, like other workers,3 16 that the data between adults and infants do not differ, one can roughly estimate the lung deposition. So, the average pulmonary deposition after inhalation via LC Star, LS 290 and Projet nebulisers was respectively 0.89%, 0.55%, and 0.46% of the nominal dose (20 mg SCG).

Earlier investigations in infants and toddlers (2–36 months) also showed a lung deposition of <1% after SCG inhalation.3 16 Even if in intubated newborns the nebulised dose was delivered direct into the inspiratory line of the ventilator circuits, the lung deposition (using an ultrasonic nebuliser) was as low as about 1%.17 For comparison, we found in patients with cystic fibrosis (9–21 years) after SCG inhalation using the LC Plus nebuliser (mouthpiece) a lung deposition of about 10% of the nominal dose.14 15 In proportion to body weight, however, the mean lung dose of SCG in these patients (0.051 mg/kg) was of an order of magnitude identical with that noted in the young infants in this study—that is, 0.079 mg/kg (LC Star) 0.048 mg/kg (LS 290) and 0.041 mg/kg (Projet).

In this study, only 10 complete datasets were available for estimating lung deposition. Yet, the post hoc calculated power was >98% for the differences between the LC Star and the other two nebulisers.

To summarise: Using the marker substance SCG the relative pulmonary deposition can be evaluated even in preterm infants. Under the inhalation conditions chosen, the lung deposition after inhalation with the LC Star jet nebuliser, which produces the greatest proportion of droplets <2 μm, was higher than with LS 290 and Projet ultrasonic nebulisers. All in all, on average, not more than 1% of the nominal dose is deposited into the lungs in these very young infants. Further investigations will be required, while considering all age-related characteristics, to find the optimum parameters for a nebuliser device that will enhance effectiveness and efficiency of aerosol treatment in young infants.