Blood gas control
This application is a continuation-in-part of our copending patent application Ser. No. 170,600 filed July 21, 1980, now U.S. Pat. No. 4,299,728. 
    
    
     This invention relates to blood gas control liquids for quality control and/or calibration of blood gas analyzer equipment. 
     Blood gas analyzers are utilized to measure parameters of blood such as pH, partial pressure of carbon dioxide (expressed as pCO 2 ), and partial pressure of oxygen (expressed as pO 2 ). Such blood gas analyzers require frequent calibration and quality control checks to insure the analyzer is operating properly and accurately. In connection with such quality control and calibration procedures, it is convenient to use a prepared blood gas control liquid of constant, known composition to monitor the accuracy of such analyzers. For practical purposes, the blood gas control liquid should have long term physical and chemical stability, constant pH, pCO 2 , and pO 2  levels, flow characteristics comparable to blood, and should be readily cleaned from and not otherwise complicate or impair analyzer accuracy. Commercially available, prepared aqueous blood gas control liquids adequately mimic blood for pH and pCO 2 , but do not have adequate oxygen buffering capacity as they are unable to dissolve an adequate amount of oxygen. Such controls are therefore prone to inaccuracy in the presence of relatively small amounts of outside oxygen contamination, and also may falsely indicate certain types of instrument malfunctions. 
     Other blood gas control products have been proposed which are based on components of human blood (e.g., Louderback U.S. Pat. No. 3,973,913) or components proposed for use as blood substitutes such as fluorocarbon and silicone compound emulsions (e.g., Sorenson et al. U.S. Pat. No. 4,163,734). One blood gas control product proposed by Sorenson et al. contains a perfluorotributylamine compound, an emulsifying agent to provide a stable suspension of the perfluorinated compound, a phosphate buffer system, and a bicarbonate ion-carbon dioxide buffer system. Sterilization, which is necessary for stability, is accomplished by means of radioactive irradiation. Such a control liquid has several undesirable limitations. A perfluorotributylamine-water emulsion with an oxygen solubility coefficient of ten requires a high fluorocarbon:water ratio (in excess of 20% fluorocarbon) which results in an undesirable viscosity that is much higher than that of blood. The emulsion leaves bubbles in the measuring chambers of blood gas analyzer equipment, creating cleaning difficulties and causing control-to-sample carry over. The specific surfactant described in Sorenson et al., like all polyol surfactants, must be used in an amount that further undesirably increases viscosity. Also the phosphate buffer system is conducive to growth of aerobic bacteria, which impair pO 2  stability, and efforts to sterilize the product by means of radioactive irradiation are themselves detrimental to oxygen stability. 
     We have discovered a superior blood gas control liquid that includes a stable oil in water emulsion that has an oxygen solubility coefficient of at least 10 ml O 2  per 100 ml emulsion and a viscosity in the range of blood (a viscosity of less than 10 centipoises). The emulsion includes water, a water insoluble non-protein organic compound, and a nonionic surfactant that is partially soluble in both the water and the oil phases. The control liquid also includes a pH buffering agent and a microbial growth inhibiting agent that is substantially nonreactive with the buffering agent. Control liquids in accordance with the invention have emulsions of long term stability (at least six months without breaking), superior oxygen buffering capacity, good pH buffering and viscosity of less then ten centipoises. 
     Preferred control liquids are formulated to provide pH, pCO 2 , and pO 2  values at three different levels (acidosis, normal, and alkalosis) and include coloring agents that simulate, both visually and analytically, corresponding hemoglobin value ranges, the acidosis level liquid having the appearance of normal venous blood, the normal level liquid having the appearance of normal arterial blood, and the alkalosis level liquid have the appearance of a low hemoglobin, high pO 2  blood, and each of these control liquids providing distinctive hemoglobin values when measured with hemoglobin analyzers. The density (at 20° C.) of a preferred blood gas control liquid is less than 1.12. Microbial growth in the liquids is discouraged without the use of irradiation or expensive sterile fill procedures; the liquids do not foam or form bubbles to a significant degree; and they are easily cleaned out of blood gas measuring equipment. 
     Blood gas control liquids in accordance with the invention are preferrably supplied enclosed in gas-tight, sealed ampuls and contain known concentrations of dissolved carbon dioxide and oxygen. A preferred liquid includes a water-based emulsion containing a water-insoluble perfluorinated compound material, a fluorocarbon-based surfactant capable of emulsifying the perfluorinated compound material, a preservative in a concentration sufficient to inhibit microbial growth without interfering with the quality control functions of the liquid, and a pH buffering agent substantially non-reactive with said preservative. 
     According to the invention, the perfluorinated compound material must have a high enough oxygen solubility coefficient, and be present in sufficient quantity, to yield a blood gas control liquid having an effective oxygen solubility coefficient of at least 10 ml O 2  /100 ml of emulsion. Further, the perfluorinated compound material must have a high enough oxygen solubility coefficient to allow it to be used in an amount which constitutes less than about 40% of the liquid by volume, because larger volumetric amounts would raise viscosity to an unacceptable level. To provide a margin of safety, the effective oxygen coefficient of the perfluorinated compound material used in the invention should be, at one atmosphere pressure, 25° C., and a volumetric proportion of 15%, at least 10 mL O 2  /100 mL emulsion. 
     A preferred perfluorinated compound material of the invention is manufactured by 3M Company under the designation &#34;FC-77&#34; and is a mixture of perfluoroalkanes and perfluorocyclic ethers. We have found that FC-77 may be advantageously used in combination with perfluorotributylamine, sold by 3M Company under the designation &#34;FC-43&#34; and/or with a dimethyl siloxane polymer that has a density of about 0.8 grams per milliliter, sold by Dow-Corning under the designation &#34;200 Fluid-1 centistoke&#34; (hereinafter &#34;200-1&#34;). Oil in water emulsions of these materials have the oxygen solubility coefficients shown in Table I below. Preferred fluorocarbon-based surfactants of the invention include fluoroalkylpoly(ethyleneoxy)ethanol, sold by DuPont Corporation under the designation Zonyl FSN; fluoracylpolyoxyethylene, sold under the designation Lodyne S-107 by Ciba-Geigy; fluorinated acylpolyoxyethylene ethanol, sold under the designation FC-170-C by 3M Company; and Monflor 51, a polymer of about 20-25 units of polyethyleneoxide with between one and four tetrafluoroethylene groups at each end, sold by Imperial Chemical Industries Ltd. 
     
                       TABLE I                                                     
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VOLUME SOLUBILITY IN ml O.sub.2 ml EMULSION                               
(at one atmosphere and 25° C.)                                     
              % Oil in Water                                              
              10%      15%      20%                                       
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200-1            4.12%      4.78     5.44                                 
FC 43           6.12        7.78     9.44                                 
FC 77           8.12       10.78    13.44                                 
3 FC 77 -1 FC 43                                                          
                7.62       10.03    12.44                                 
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     The buffering agent used in the invention is selected so that it is substantially non-reactive with the preservative being used. Because the preferred preservatives are the aldehydes, and particularly formaldehyde and gluteraldehyde, the preferred buffering agents are those containing a tertiary amine; such buffering agents do not react with aldehydes, and also are relatively non-conducive to microbial growth. The preferred tertiary amine buffering agent for use with an aldehydic preservative is HEPES (N-2-Hydroxyethyl-piperazine-N&#39;-2-ethane sulfonic acid), its pK being centered in the appropriate physiological pH range. Another effective tertiary amine buffering agent for use with aldehydic preservatives is triethanolamine. 
     The blood gas control liquids are preferrably formulated to simulate three physiological levels of pH, pCO 2 , and pO 2  values; and have coloring agents that simulate hemoglobin values and visual appearances of a range of blood conditions. A preferred blood gas control liquid has a density (at 20° C.) of about 1.08, facilitating its use in &#34;open junction&#34; types of blood gas analyzers where mixing of the blood gas control liquid and the potassium chloride electrolyte would interfere with and produce inaccurate measurement. 
    
    
     A preferred embodiment of the invention has the following formulation: 
     
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Compound             Concentration                                        
______________________________________                                    
FC-77                10.95% (v/v)                                         
FC-43                3.65% (v/v)                                          
200-1                19.8% (v/v)                                          
Fluoroalkyl                                                               
poly (ethyleneoxy)                                                        
ethanol (Zonyl FSN)  2.5% (v/v)                                           
HEPES Buffer salts   32.8 mM                                              
NaOH                 30.50-35.23 mM                                       
NaCl                 26.32-48.99 mM                                       
Formaldehyde         43.5 mM                                              
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     The first step in the preparation of this liquid was to prepare the antibacterial/surfactant/water solution by mixing together 1250 grams of 40% Zonyl FSN and 120 mL of formaldehyde and bringing the solution up to 16.0 liters with distilled water. The 200-1 silicone oil is then poured into the hopper of a Gaulin Homogenizer (Model 15M) and the aqueous phase is added at the rate of 725 mL/minute. The mixture is homogenized at a pressure of 2500 psig and repeatedly passed through an ice bath and recirculated to the hopper until all of the oil had been dripped into it. The pressure was then increased to 8000 psig and two discrete passes through the homogenizer are carried out to form a stable emulsion. 
     Next, an oil mixture was prepared by combining 2.20 L FC-77 with 0.73 L FC-43. The silicone emulsion was placed in the hopper of a Gaulin Homogenizer (Model 15M) and the fluorocarbon oil mixture was pumped into the hopper at the rate of 180 mL/min. while the liquid in the hopper was homogenized at a pressure of 2500 psig. The liquid was repeatedly passed through an ice bath and recirculated to the hopper until all of the oil had been dripped into it. At this point, the pressure was increased to 8000 psig and nine discrete passes through the homogenizer were carried out. A stable emulsion was formed by the ninth pass. 
     The emulsion was allowed to stand for 24 hours, and was then filtered through Whatman #3 filter paper using vacuum filtration with a Buchner funnel. 
     In a separate operation, buffers were prepared to be added to the emulsion. Three different buffers were made using HEPES, NaOH, and NaCl in different concentrations. After the buffers were added to the emulsions, each of the three emulsions was equilibrated with a different gas mixture, producing three different control liquids to be used under appropriate conditions. The three levels differed in pH, pCO 2 , and PO 2 , the levels having been chosen to reflect pH and gas partial pressure of the blood of normal patients, patients suffering from alkemia, and those suffering from acidemia. The three levels were: 
     
         ______________________________________                                    
Acidosis       Normal      Alkalosis                                      
______________________________________                                    
pH      7.15-7.25  7.34-7.41   7.55-7.65                                  
pCO.sub.2                                                                 
        65-75 mm Hg                                                       
                   40-47 mm Hg  18-24 mm Hg                               
pO.sub.2                                                                  
        60-70 mm Hg                                                       
                   95-110 mm Hg                                           
                               140-160 mm Hg                              
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     The buffer formulations corresponding to the three levels were: 
     
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Acidosis          Normal     Alkalosis                                    
______________________________________                                    
NaOH   30.50 mM/L emulsion                                                
                      33.91 mM/L 35.23 mM/L                               
HEPES  32.80 mM       32.80 mM/L 32.80 mM/L                               
NaCl   26.32 mM       48.99 mM/L 48.94 mM/L                               
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     The buffer formulations were added to the emulsion and each level was then given a distinctive color using the following dyes: 
     
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Acidosis       Normal       Alkalosis                                     
______________________________________                                    
1.667 g/L      1.25 g/L     0.781 g/L                                     
Red #33        Red #33      Red #33                                       
3.333 g/L      1.25 g/L     1.406 g/L                                     
Yellow #5      Yellow #5    Yellow #5                                     
0.01667 g/L                                                               
Blue #1                                                                   
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     The emulsions, containing dyes and dissolved buffer salts and having viscosities of about 8 centipoises (measured with a Brookfield Viscometer at 60 rpm-spindle #1), O 2  solubility coefficients of about 12.5, and densities of about 1.08 (at 20° C.), were then placed in a container which was thermally controlled to 25° C.±0.5° C. The appropriate equilibration gas mixture was then bubbled through each emulsion at the rate of 3 L/min. until the pH, pO 2 , and pCO 2  reached equilibrium values, as determined by blood gas measuring equipment. The equilibration gas mixtures had the following compositions: 
     
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       Acidosis  Normal      Alkalosis                                    
______________________________________                                    
CO.sub.2 6.7%        4.15        2.0                                      
O.sub.2  9.0%        14.0        21.5                                     
N.sub.2  84.3%       81.85       76.5                                     
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     After equilibration, glass ampuls which had been purged with the same gas equilibration mixture used for the emulsion were filled with 2 mL of the appropriate emulsion, up to below a hand-breakable line, and the ampuls heat-sealed. 
     The formulated control liquids, in their sealed ampuls, were used successfully to check the calibration of blood gas analyzers. An ampul containing the appropriate emulsion is opened and the control liquid is placed in the blood gas analyzer, which measures and registers the pH, pCO 2  and pO 2  of the emulsion. A reading outside of the pH and gas partial pressure ranges known to exist in the liquid indicates the need for re-calibration or repair of the analyzer. 
     The appearances and hemoglobin values of these formulated control liquids as set out in the following table: 
     
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       ABL 1 &amp; 2                                                          
               IL 282    Appearance                                       
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Acidosis 17-22 g/dL                                                       
                   11-14 g/dL                                             
                             Normal venous blood                          
Normal   14-18      8-12     Normal arterial                              
                             blood                                        
Alkalosis                                                                 
          8-12      6-9      High pO.sub.2, low                           
                             hemoglobin blood                             
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     Another embodiment of the invention has the following formulation: 
     
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Compound             Concentration                                        
______________________________________                                    
FC-77                15% (v/v)                                            
FC-43                5% (v/v)                                             
Fluoroalkyl                                                               
poly (ethyleneoxy)                                                        
ethanol (Zonyl FSN)  2% (v/v)                                             
HEPES Buffer salts   40 mM                                                
NaOH                 38.7-42.57 mM                                        
NaCl                 85.47-89.29 mM                                       
Formaldehyde         53 mM                                                
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     The first step in the preparation of this liquid was to prepare the antibacterial/surfactant/water solution by mixing together 500 grams of 40% Zonyl FSN and 60 mL of formaldehyde and bringing the solution up to 8.0 liters with distilled water. Next, an oil mixture was prepared by combining 1.5 L FC-77 with 0.5 L FC-43. 
     The aqueous solution was placed in the hopper of a Gaulin Homogenizer (Model 15M) and the oil mixture placed in a separatory funnel mounted above the hopper. Oil was dripped into the hopper at the rate of 100 mL/min. while the liquid in the hopper was homogenized at a pressure of 2500 psi. The liquid was repeatedly passed through an ice bath and recirculated to the hopper until all of the oil had been dripped into it. At this point, the pressure was increased to 8000 psi and nine discrete passes through the homogenizer were carried out. A stable emulsion was formed by the ninth pass. 
     The emulsion was allowed to stand for 24 hours, and was then filtered through Whatman #3 filter paper using vacuum filtration with a Buchner funnel. 
     In a separate operation, buffer solutions were prepared to be added to the emulsion. Three different buffer solutions were made using HEPES, NaOH, and NaCl in different concentrations. After the buffer solutions were added to the emulsions, each of the three emulsions was equilibrated with a different gas mixture, producing three different control liquids to be used under appropriate conditions. The three levels differed in pH, pCO 2 , and PO 2 , the levels having been chosen to reflect pH and gas partial pressure of the blood of normal patients, patients suffering from alkemia, and those suffering from acidemia. The three levels were: 
     
         ______________________________________                                    
Acidosis       Normal      Alkalosis                                      
______________________________________                                    
pH      7.15-7.25  7.34-7.41   7.55-7.65                                  
pCO.sub.2                                                                 
        65-75 mm Hg                                                       
                   40-47 mm Hg  18-24 mm Hg                               
pO.sub.2                                                                  
        60-70 mm Hg                                                       
                   95-110 mm Hg                                           
                               140-160 mm Hg                              
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     The buffer formulations corresponding to the three levels were: 
     
         ______________________________________                                    
Acidosis          Normal     Alkalosis                                    
______________________________________                                    
NaOH   38.7 mM/L emulsion                                                 
                      40.62 mM/L 42.57 mM/L                               
HEPES  40.00 mM       40.00 mM/L 40.00 mM/L                               
NaCl   89.29 mM       87.38 mM/L 85.43 mM/L                               
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     The buffer formulations were added to the emulsion and each level was then given a distinctive color using the following dyes: 
     
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Acidosis        Normal      Alkalosis                                     
______________________________________                                    
100 mg/l Yellow #5                                                        
                100 mg/l    66.67 mg/l                                    
                Amarath     Amarath,                                      
                            133.33 mg/l                                   
                            Yellow #5                                     
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     The emulsions, containing dyes and dissolved buffer salts and having viscosities of about 4.5 centipoises, O 2  solubility coefficients of about 12.4, and a density of about 1.15 (at 20° C.), were then placed in a container which was thermally controlled to 25° C.±0.5° C. The appropriate equilibration gas mixture was then bubbled through each emulsion at the rate of 2 L/min. until the pH, pO 2 , and pCO 2  reached equilibrium values, as determined by blood gas measuring equipment. The equilibration gas mixtures had the following compositions: 
     
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       Acidosis  Normal      Alkalosis                                    
______________________________________                                    
CO.sub.2 7.0%        4.17        1.98                                     
O.sub.2  9.5%        14.0        21.5                                     
N.sub.2  83.5%       81.83       76.52                                    
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     After equilibration, glass ampuls which had been purged with the same gas equilibration mixture used for the emulsion were filled with 1 mL of the appropriate emulsion, up to below a hand-breakable line, and the ampuls heat-sealed. 
     Another embodiment of the invention employs a mixture of perfluoroalkanes and perfluorocyclic ethers (FC-77) and a polymer of about 20-25 units of polyethyleneoxide with between one and four tetrafluoroethylene groups at each end (Monflor 51). Two liters of FC-77 were emulsified, as described above for the FC-77/FC-43 emulsion, with an antibacterial surfactant/water solution consisting of 150 g Monflor 51 (about 1.5% by volume of emulsion) and 60 ml formaldehyde brought up to 8.00 liters with distilled water. After all the FC-77 had been added, the emulsion was subjected to seven discrete passes through the Gaulin Homogenizer. 
     Because Monflor 51 often contains HF as an impurity, the emulsions were neutralized with NaOH. After neutralization, buffers and dyes were added as described above for FC-77/FC-43, they were equilibrated with the appropriate gas mixtures, and the emulsions placed in ampuls. The liquids had viscosities of about 5 centipoises, O 2  solubility coefficients of about 13.5, and densities of about 1.15. In still another embodiment, liquids with stable emulsions of about 15% FC-77, 15% 200-1 silicone oil and Monflor 51 surfactant formulated and equilibrated as above described have densities of about 1.09, O 2  solubility coefficients of about 12.5, and viscosities of less than 10 centipoises. 
     The embodiments described above do not include added bicarbonate, but these emulsions nevertheless, owing to the gas equilibration step, effectively contain bicarbonate. Alternatively, it is of course possible to add bicarbonate, in appropriate amounts, along with HEPES. 
     The emulsions, in their sealed ampuls, were used successfully to check the calibration of blood gas analyzers. An ampul containing the appropriate emulsion is opened and the control liquid is placed in the blood gas analyzer, which measures and registers the pH, pCO 2  and pO 2  of the emulsion. A reading outside of the pH and gas partial pressure ranges known to exist in the liquid indicates the need for re-calibration or repair of the analyzer. 
     While particular embodiments of the invention have been described, various modifications will be apparent to those skilled in the art, and it is not intended that the invention be limited to the disclosed embodiments or to details thereof and departures may be made therefrom within the spirit and scope of the invention.