**3. Direct measurement of VO2 using respiratory mass spectrometry**

The concept of mass spectrometry (measuring fractional proportions of substances in a mixture, according to their molecular mass-charge ratios) was first introduced at the end of the 19th century. The recruitment of mass spectrometry into respiratory physiology in the 1940s, and its subsequent refinement over the decades, has established a 'state-of-the-art' method for measuring VO2 using highly accurate and rapid multiple gas analysis. The mixed expirate method (Davies and Denison 1979) enabled use of the mass spectrometer alone to measure metabolic gas exchange and ventilation volume. This technique has been used widely to measure VO2 in a broad spectrum of clinical and experimental conditions. We have adapted the method to continuously measure VO2 with a variety of pediatric ventilators and anesthesia ventilators before, during, and after CPB, using the AMIS 2000 Medical Respiratory Mass Spectrometer System (Innovision A/C Odense, Denmark). Our combination of techniques and equipment makes respiratory mass spectrometry a unique and powerful tool in multiple settings: in the cardiac catheterization laboratory (Schulze-Neick, Li et al. 2002 ; Shekerdemian, Bush et al. 1997; Shekerdemian, Bush et al. 1997), in the operating room (Li, Stokoe et al. 2004), and in the ICU (Li, Hoschtitzky et al. 2004; Li, Schulze-Neick et al. 2000; Li, Zhang et al. 2006; Li, Zhang et al. 2006; Li, Zhang et al. 2007; Li, Zhang et al. 2007; Li, Zhang et al. 2008; Schulze-Neick, Li et al. 2001).

### **3.1 Principles of mass spectrometry**

Mass spectrometers analyze substances in the gas phase by performing a sequence of five operations: *1)* accept and vaporize a minute controlled quantity of sample; *2)* reduce the sample vapor to a very low pressure; *3)* ionize a representative part of the vapor; *4)* separate the ionized particles produced, according to their mass-to-charge ratio; and *5)* read the abundance of particles at specific values of the mass-to-charge ratio.

#### **3.2 Hardware design**

298 Congenital Heart Disease – Selected Aspects

particular, 'fat mass' is relatively higher in younger children (Fomon, Haschke et al. 1982; Moukarzel, Salas et al. 2003) and VO2 in fat mass is about 20 times lower than in muscular

Fig. 4. The trends in relation to age of (A) estimated VO2, (B) measured VO2, and (C) their

We conclude that estimation of VO2 is unacceptably inaccurate for clinical decision-making and research. Direct measurement of VO2 is appropriate for young children with congenital heart defects undergoing cardiac catheterization and in the ICU after CPB. Direct and continuous measurement of VO2 is essential, particularly in those younger than 3 years and

VO2 can also be calculated by the reverse Fick method from the cardiac output, directly measured by the thermodilution technique, for example. This method has obvious limitations for clinical application in patients with congenital heart defects. First, the calculation is clearly intermittent. Second, the presence of intracardiac shunting and tricuspid regurgitation can significantly affect the accuracy of the calculation. Most important, certain complex circulations preclude the use of the thermodilution technique (e.g., after the Norwood procedure, bidrectional caval pulmonary shunt, or the Fontan operation), because of anatomical (e.g., parallel systemic and pulmonary circulations) and

difference, in 75 patients undergoing cardiac catheterization.

in the early postoperative period after CPB.

**2.2 Inadequacy of the reverse Fick method** 

methodological (e.g., inadequate mixing) limitations.

mass (Moukarzel, Salas et al. 2003).

#### Features of a mass spectrometer are outlined in Figure 5.

A Teflon capillary tubing (A) with an internal diameter 0.3 mm and a length of 3 to 6 m provides the gas transport from the sampling site (B) to the vacuum system. There are three sample inlets, one for the on-line continuous monitoring of O2 and CO2 fractional concentrations, the other two for the inspiratory and expiratory gas sampling. The gas sample at atmospheric pressure is drawn at a sampling rate of 10 to 20 mL/min down the tubing (A), passing through the three static electro-magnetic valves (C), which are selected in turn as appropriate to the sequence of gas analysis needed for the metabolic calculations. The sample is drawn by the inlet rotary vacuum pump (D) into the sample chamber (E), and all but a small fraction of it is pumped away continuously at the low-pressure end of the inlet tubing. The turbo molecular pump (F), supported by a second backing rotary pump (G), provides a very high vacuum environment of around 5 10-7 mBar in the dispersion chamber (H), housing ionization chamber (I), mass filter (J), and ion detector (K). Thus, the high vacuum provided by the turbo molecular pump draws the remaining gas sample through the molecular leak (L) into the ionization chamber, where the gas molecules are ionized. The ions are focused into a tight beam, enter the aperture of the quadrupole mass filter, and are separated by the quadrupole mass filter (J). The use of appropriate voltages in this field allows only ions of a definite mass interval to pass and reach the ion detector (K),

Accurate Measurement of Systemic

STPD = Standard temperature and pressure dry.

of the mixing chamber

**with anesthesia ventilators** 

Oxygen Consumption in Ventilated Children with Congenital Heart Disease 301

FMO2, FMCO2, FMTr: = measured fractional concentrations of O2, CO2 and tracer gas at the outlet

**3.4 Setup of respiratory mass spectrometer in the cardiac catheterization laboratory** 

laboratory to anesthesia ventilators with a partial rebreathing system (Figure 6).

Fig. 6. The setup of the AMIS2000 respiratory mass spectrometer sampling inlets, mixing box, and the circuit of the anesthesia ventilator in the cardiac catheterization laboratory.

Inlet 3 samples the 'effluent' mixed expirate from the distal end of the mixing box.

Most children undergoing cardiac catheterization are paralyzed and mechanically ventilated by an anesthesia ventilator with a partial rebreathing system, whereby the exhaled gas is recirculated via a CO2 absorber and fresh gas mixture is continuously supplied. The mixing box is inserted in the expiratory limb of the ventilator circuit to collect the expired gas. Inlet 1 is placed at the mouth piece close to the endotracheal tube, for continuous on-line monitoring of breath-to-breath oxygen and carbon dioxide fractional concentrations. This checks that the steady state had not been perturbed. Another two inlets are used for the measurement of VO2. Inlet 2, which is placed in the inspiratory limb of the ventilator circuit, samples inspiratory gas;

Accurate measurement of minute ventilation relies on the complete collection of expired gas and therefore a leak-free circuit. Ideally, patients are intubated with cuffed endotracheal tubes. Precise VO2 measurement requires a steady-state period, thus patients are sedated and paralyzed to obviate the confounding effects of movement, agitation, and pain on VO2. We have adapted the AMIS2000 respiratory mass spectrometer in the cardiac catheterization

suppressing the inevitable 'noise' which would otherwise be created by the contaminants that cross the mass filter as a result of scatter. The ions are then collected and pre-amplified in a current-voltage pre-amplifier prior to signal processing. The resulting signal is therefore made as pure as possible before being amplified by the secondary electron multiplier (SEM). Amplification enables a higher sensitivity and faster operation. The resulting output of the detection unit is proportional to the ion current, which in turn is proportional to the partial pressure of the gas species. (Pressures indicated are at normal operating conditions).

Fig. 5. Design of AMIS 2000 respiratory mass spectrometer.

#### **3.3 The measurement of VO2**

The mass spectrometer measures metabolic gas exchange by the 'inert gas dilution method'. 'A known mass flow of a marker gas is injected into expired gas upstream of a mixing box, and the resulting gas composition downstream used to deduce the mass flows of all its components' (Davies and Denison 1979). VO2 is then calculated every 30 seconds as:

$$\text{VO}\_2\text{ (STPD)} = \text{V}\_{\text{Tr}}\text{(STPD)} \bullet \left[\text{F}\_{\text{IC2}} \bullet \left(\text{1-F}\_{\text{MCO2}-\text{F}\_{\text{MTr}}}\right)\text{-F}\_{\text{MO2}} \bullet \left(\text{1-F}\_{\text{F}\text{CO2}-\text{F}\_{\text{ITr}}}\right)\right] / \text{D} \tag{3}$$

D = FMTr (1- FIO2- FICO2) - FItr (1- FMO2- FMCO2) VTr = added flow of indicator gas (tracer, Argon)

FIO2, FICO2, FItr = constant fractional concentrations of O2, CO2 and tracer gas in inspired air

FMO2, FMCO2, FMTr: = measured fractional concentrations of O2, CO2 and tracer gas at the outlet of the mixing chamber

STPD = Standard temperature and pressure dry.

300 Congenital Heart Disease – Selected Aspects

suppressing the inevitable 'noise' which would otherwise be created by the contaminants that cross the mass filter as a result of scatter. The ions are then collected and pre-amplified in a current-voltage pre-amplifier prior to signal processing. The resulting signal is therefore made as pure as possible before being amplified by the secondary electron multiplier (SEM). Amplification enables a higher sensitivity and faster operation. The resulting output of the detection unit is proportional to the ion current, which in turn is proportional to the partial

pressure of the gas species. (Pressures indicated are at normal operating conditions).

Molecular leak

Fig. 5. Design of AMIS 2000 respiratory mass spectrometer.

Inlet rotary pump 0.5 mBar

Ionisation chamber 10-4 mBar

H

J

I

Sampling site 1000 mBar


A

<sup>E</sup>

Inlet Valve

D = FMTr (1- FIO2- FICO2) - FItr (1- FMO2- FMCO2) VTr = added flow of indicator gas (tracer, Argon)

The mass spectrometer measures metabolic gas exchange by the 'inert gas dilution method'. 'A known mass flow of a marker gas is injected into expired gas upstream of a mixing box, and the resulting gas composition downstream used to deduce the mass flows of all its components' (Davies and Denison 1979). VO2 is then calculated every 30

Backing rotary pump

Turbo molecular pump

K

<sup>+</sup> <sup>L</sup>

5 10 -7 mBar

G

F

Dispersion chamber

Ion Detector (SEM)

10 -2 mBar

Signal out

Ion deflector


VO2 (STPD) = VTr (STPD) [FIO2 (1-FMCO2-FMTr)-FMO2 (1-FICO2-FITr)] / D (3)

FIO2, FICO2, FItr = constant fractional concentrations of O2, CO2 and tracer gas in inspired air

**3.3 The measurement of VO2**

D

Sample chamber 10 mBar

C

B

seconds as:

#### **3.4 Setup of respiratory mass spectrometer in the cardiac catheterization laboratory with anesthesia ventilators**

Accurate measurement of minute ventilation relies on the complete collection of expired gas and therefore a leak-free circuit. Ideally, patients are intubated with cuffed endotracheal tubes. Precise VO2 measurement requires a steady-state period, thus patients are sedated and paralyzed to obviate the confounding effects of movement, agitation, and pain on VO2. We have adapted the AMIS2000 respiratory mass spectrometer in the cardiac catheterization laboratory to anesthesia ventilators with a partial rebreathing system (Figure 6).

Fig. 6. The setup of the AMIS2000 respiratory mass spectrometer sampling inlets, mixing box, and the circuit of the anesthesia ventilator in the cardiac catheterization laboratory.

Most children undergoing cardiac catheterization are paralyzed and mechanically ventilated by an anesthesia ventilator with a partial rebreathing system, whereby the exhaled gas is recirculated via a CO2 absorber and fresh gas mixture is continuously supplied. The mixing box is inserted in the expiratory limb of the ventilator circuit to collect the expired gas. Inlet 1 is placed at the mouth piece close to the endotracheal tube, for continuous on-line monitoring of breath-to-breath oxygen and carbon dioxide fractional concentrations. This checks that the steady state had not been perturbed. Another two inlets are used for the measurement of VO2. Inlet 2, which is placed in the inspiratory limb of the ventilator circuit, samples inspiratory gas; Inlet 3 samples the 'effluent' mixed expirate from the distal end of the mixing box.

Accurate Measurement of Systemic

**4.1 The Fick principle** 

substance.' (Fick 1870).

2004; Li, Schulze-Neick et al. 2000; Li, Zhang et al. 2007).

Oxygen Consumption in Ventilated Children with Congenital Heart Disease 303

surgery in children with congenital heart defects (Li, Hoschtitzky et al. 2004; Li, Hoskote et al. 2005; Li, Schulze-Neick et al. 2000; Li, Zhang et al. 2006; Li, Zhang et al. 2006; Li, Zhang et al. 2007; Li, Zhang et al. 2007; Li, Zhang et al. 2008; Zhang 2008; Zhang, Holtby et al. 2008).

The importance of oxygen transport is increasingly appreciated in the care of critically ill patients, particularly after CPB (Gilbert, Haupt et al. 1986; Haupt, Gilbert et al. 1985; Hoffman, Ghanayem et al. 2000; Li, Hoschtitzky et al. 2004; Li, Hoskote et al. 2005; Li, Schulze-Neick et al. 2000; Li, Zhang et al. 2006; Li, Zhang et al. 2006; Li, Zhang et al. 2007; Li, Zhang et al. 2007; Li, Zhang et al. 2008; Oudemans-van Straaten, Jansen et al. 1996; Powers, Mannal et al. 1973; Tweddell, Hoffman et al. 1999; Zhang 2008; Zhang, Holtby et al. 2008). The fundamental requirement of ICU management is to match systemic oxygen delivery (DO2) to VO2, to sustain cellular metabolism and end-organ function. Many patients requiring ICU support have reduced DO2, usually as a result of decreased myocardial function. However, in many patients the reduced cardiac output and DO2 are further compounded by secondary abnormalities of VO2 that amplify the deficiency of DO2 and contribute to the overall imbalance of oxygen transport. This combination is seen in many situations such as sepsis and trauma (Gilbert, Haupt et al. 1986; Haupt, Gilbert et al. 1985; Powers, Mannal et al. 1973), but is a consistent feature of cardiac surgery (Hoffman, Ghanayem et al. 2000; Li, Hoschtitzky et al.

Most children experience a phase of reduced cardiac output during the first few hours after CPB (Hoffman, Ghanayem et al. 2000; Li, Zhang et al. 2007; Wernovsky, Wypij et al. 1995). At the same time, VO2 is increased and highly dynamic in relation to central body temperature (Li, Hoschtitzky et al. 2004; Li, Schulze-Neick et al. 2000), the systemic inflammatory response (Li, Hoschtitzky et al. 2004; Oudemans-van Straaten, Jansen et al. 1996), pharmacological manipulations (Hayes, Timmins et al. 1994; Li, Zhang et al. 2006), and ventilatory manipulations (Li, Hoskote et al. 2005; Li, Zhang et al. 2008). VO2 is an important constituent in the balance of oxygen transport, but has been largely ignored in ICU management. Traditional management of these patients has focused on augmenting myocardial performance through the use of inotropes, for example, to enhance cardiac output and DO2. However, inotropic agents may not effectively enhance cardiac output in the presence of myocardial injury, and may paradoxically stimulate systemic and myocardial oxygen consumption, offsetting any gains in DO2 (Fowler, Alderman et al. 1984; Hayes, Timmins et al. 1994; Li, Zhang et al. 2006). An alternative (and in some ways more rational) approach to improving the balance of oxygen transport may be to decrease VO2. For example, the use of skeletal muscle paralysis and assisted ventilation is a standard therapy directed at reducing metabolic demands. These simple maneuvers may reduce VO2 by up to 20 to 30% (Nisbet, Dobbinson et al. 1973; Westenskow, Jordan et al. 1978). Similarly, profound reductions in VO2 may be achieved simply by controlling central body temperature. We have shown in infants after cardiac surgery, for example, that central pyrexia increases VO2 by approximately 11% per degree Celsius (Li, Schulze-Neick et al. 2000). Therefore, in the current conceptualization of oxygen transport, emphasis should shift

beyond cardiac output and oxygen delivery to the balance between DO2 and VO2.

**4. Calculation of oxygen transport parameters using directly measured VO2** 

The Fick principle states that 'The total uptake or release of any substance by an organ is the product of blood flow to the organ and the arteriovenous concentration difference of the

#### **3.5 Setup of respiratory mass spectrometer in the ICU with pediatric ventilators**

We have also adapted the AMIS2000 respiratory mass spectrometer in the ICU to various pediatric ventilators with continuous flow (Figure 7). Pediatric ventilators in the ICU use continuous flow to supply fresh gas throughout the breathing cycle. This avoids increasing the amount of work required to trigger spontaneous breathing. In a setup different from the anesthesia ventilator, the mixing box is connected to the exit port of the pediatric ventilator to collect the expired gas, and is also connected to an 'expiratory' inlet (Inlet 3). The expiratory inlet allows sampling of the 'effluent' mixed expirate from the distal end of the mixing box. Inlets 1 and 2 are placed at the mouth piece and in the inspiratory limb respectively, in the same way as in the anesthesia ventilator.

Fig. 7. The setup of the AMIS2000 respiratory mass spectrometer sampling inlets, mixing box, and the circuit of the pediatric ventilator in the ICU.

#### **3.6 Clinical applications of respiratory mass spectrometry in the ICU**

Respiratory mass spectrometers were first produced for commercial applications in the late 1980s. The reported precision of the mass spectrometer was as low as 5% in spontaneous breathing at rest or during exercise. The precision increases in paralyzed and ventilated patients. Precision was further improved by avoiding the use of flow sensors. Versatility of the respiratory mass spectrometer is greatly increased by the use of long sampling probes up to 30 meters(Davies and Denison 1979). Long probes allow the study of subjects who otherwise might be inaccessible due to the size of the equipment, such as patients undergoing cardiac catheterization, intensive care, or surgery, where space at the bedside is severely limited. Long probes also enable simultaneous events to be examined sequentially, and permit a single mass spectrometer to be shared between several patients or laboratories. Respiratory mass spectrometers have become valuable clinical research tools. Our adaptation of the respiratory mass spectrometer to pediatric ventilators has allowed us to extensively study systemic hemodynamics and oxygen transport, and the factors affecting them, before and after cardiac

**Respiratory mass spectrometer**

M i x i n g b o x

Exp inlet (Inlet 3)

Fig. 7. The setup of the AMIS2000 respiratory mass spectrometer sampling inlets, mixing

Respiratory mass spectrometers were first produced for commercial applications in the late 1980s. The reported precision of the mass spectrometer was as low as 5% in spontaneous breathing at rest or during exercise. The precision increases in paralyzed and ventilated patients. Precision was further improved by avoiding the use of flow sensors. Versatility of the respiratory mass spectrometer is greatly increased by the use of long sampling probes up to 30 meters(Davies and Denison 1979). Long probes allow the study of subjects who otherwise might be inaccessible due to the size of the equipment, such as patients undergoing cardiac catheterization, intensive care, or surgery, where space at the bedside is severely limited. Long probes also enable simultaneous events to be examined sequentially, and permit a single mass spectrometer to be shared between several patients or laboratories. Respiratory mass spectrometers have become valuable clinical research tools. Our adaptation of the respiratory mass spectrometer to pediatric ventilators has allowed us to extensively study systemic hemodynamics and oxygen transport, and the factors affecting them, before and after cardiac

**3.5 Setup of respiratory mass spectrometer in the ICU with pediatric ventilators**  We have also adapted the AMIS2000 respiratory mass spectrometer in the ICU to various pediatric ventilators with continuous flow (Figure 7). Pediatric ventilators in the ICU use continuous flow to supply fresh gas throughout the breathing cycle. This avoids increasing the amount of work required to trigger spontaneous breathing. In a setup different from the anesthesia ventilator, the mixing box is connected to the exit port of the pediatric ventilator to collect the expired gas, and is also connected to an 'expiratory' inlet (Inlet 3). The expiratory inlet allows sampling of the 'effluent' mixed expirate from the distal end of the mixing box. Inlets 1 and 2 are placed at the mouth piece and in the inspiratory limb

respectively, in the same way as in the anesthesia ventilator.

**Pediatric ventilator**

box, and the circuit of the pediatric ventilator in the ICU.

end tidal CO2 (Inlet 1)

> Insp inlet (Inlet 2)

**3.6 Clinical applications of respiratory mass spectrometry in the ICU** 

surgery in children with congenital heart defects (Li, Hoschtitzky et al. 2004; Li, Hoskote et al. 2005; Li, Schulze-Neick et al. 2000; Li, Zhang et al. 2006; Li, Zhang et al. 2006; Li, Zhang et al. 2007; Li, Zhang et al. 2007; Li, Zhang et al. 2008; Zhang 2008; Zhang, Holtby et al. 2008).

The importance of oxygen transport is increasingly appreciated in the care of critically ill patients, particularly after CPB (Gilbert, Haupt et al. 1986; Haupt, Gilbert et al. 1985; Hoffman, Ghanayem et al. 2000; Li, Hoschtitzky et al. 2004; Li, Hoskote et al. 2005; Li, Schulze-Neick et al. 2000; Li, Zhang et al. 2006; Li, Zhang et al. 2006; Li, Zhang et al. 2007; Li, Zhang et al. 2007; Li, Zhang et al. 2008; Oudemans-van Straaten, Jansen et al. 1996; Powers, Mannal et al. 1973; Tweddell, Hoffman et al. 1999; Zhang 2008; Zhang, Holtby et al. 2008). The fundamental requirement of ICU management is to match systemic oxygen delivery (DO2) to VO2, to sustain cellular metabolism and end-organ function. Many patients requiring ICU support have reduced DO2, usually as a result of decreased myocardial function. However, in many patients the reduced cardiac output and DO2 are further compounded by secondary abnormalities of VO2 that amplify the deficiency of DO2 and contribute to the overall imbalance of oxygen transport. This combination is seen in many situations such as sepsis and trauma (Gilbert, Haupt et al. 1986; Haupt, Gilbert et al. 1985; Powers, Mannal et al. 1973), but is a consistent feature of cardiac surgery (Hoffman, Ghanayem et al. 2000; Li, Hoschtitzky et al. 2004; Li, Schulze-Neick et al. 2000; Li, Zhang et al. 2007).

Most children experience a phase of reduced cardiac output during the first few hours after CPB (Hoffman, Ghanayem et al. 2000; Li, Zhang et al. 2007; Wernovsky, Wypij et al. 1995). At the same time, VO2 is increased and highly dynamic in relation to central body temperature (Li, Hoschtitzky et al. 2004; Li, Schulze-Neick et al. 2000), the systemic inflammatory response (Li, Hoschtitzky et al. 2004; Oudemans-van Straaten, Jansen et al. 1996), pharmacological manipulations (Hayes, Timmins et al. 1994; Li, Zhang et al. 2006), and ventilatory manipulations (Li, Hoskote et al. 2005; Li, Zhang et al. 2008). VO2 is an important constituent in the balance of oxygen transport, but has been largely ignored in ICU management. Traditional management of these patients has focused on augmenting myocardial performance through the use of inotropes, for example, to enhance cardiac output and DO2. However, inotropic agents may not effectively enhance cardiac output in the presence of myocardial injury, and may paradoxically stimulate systemic and myocardial oxygen consumption, offsetting any gains in DO2 (Fowler, Alderman et al. 1984; Hayes, Timmins et al. 1994; Li, Zhang et al. 2006). An alternative (and in some ways more rational) approach to improving the balance of oxygen transport may be to decrease VO2. For example, the use of skeletal muscle paralysis and assisted ventilation is a standard therapy directed at reducing metabolic demands. These simple maneuvers may reduce VO2 by up to 20 to 30% (Nisbet, Dobbinson et al. 1973; Westenskow, Jordan et al. 1978). Similarly, profound reductions in VO2 may be achieved simply by controlling central body temperature. We have shown in infants after cardiac surgery, for example, that central pyrexia increases VO2 by approximately 11% per degree Celsius (Li, Schulze-Neick et al. 2000). Therefore, in the current conceptualization of oxygen transport, emphasis should shift beyond cardiac output and oxygen delivery to the balance between DO2 and VO2.
