sample="quota" bates="SF0823867" isource="ctr" decade="1970" class="ui" date="19760206" ANNUAL PROGRESS SUMMARY and TECHNICAL PROPOSAL to The Council for Tobacco Research-USA, Inc. on CHARACTERIZATION OF ANIMAL INHALATION EXPOSURE DEVICES and MICE DOSIMETRY STUDIES ON THE WALTON HORIZONTAL SMOKING MACHINE J. R. Stokely and J. E. Caton Analytical Chemistry Division Oak Ridge National Laboratory Oak Ridge, Tennessee 37830 February 6, 1976 This Document Subject to Confidentiality Agreement This Document Subject to Confidentiality Agreement ANNUAL PROGRESS SUMMARY CHARACTERIZATION OF ANIMAL INHALATION EXPOSURE DEVICES I. INTRODUCTION This report summarizes progress on the project to characterize animal exposure devices for the period January, 1975 to January 1976. The primary objectives of the project are to establish methodology for evaluating inhalation exposure devices, to apply this methodology to devices of interest to CTR, and to formulate recommendations for elimination of discovered shortcomings in the devices. II. CURRENT STATUS In this report period, emphasis and priority have been placed on expediting development of the Process and Instruments Smoke Exposure Machine (SEM). A prototype model (SEM 1) has received a thorough evaluation of operational characteristics to discover possible shortcoming. Recommendations for improvement in the SEM were made and are incorporated into a new model of the smoking machine (SEM II) which has been built and is presently undergoing tests by the manufacturer. Evaluation and study of the new model (SEM II) will be performed by us in the near future. Prototype models of mice containment/exposure units designed by Process and Instruments Corp. (P&I) have been evaluated. In cooperation with P&I, we have made design changes in the containment/exposure units to decrease animal stress, prevent animal injury, and provide for more relevant inhalation exposures. Chemical studies were made with the prototype unit to establish the number of mice which could be simultaneously exposed with minimal animal effects on smoke composition and carbon dioxide build-up. A final model of the mice containment/exposure unit is presently being constructed for testing by us and installation at Microbiological Associates. Study and evaluation of the Walton Horizontal smoking machine were extensive during previous contract periods. Additional characterization/evaluation of this smoking machine have been made during this contract period using newly developed methodology. An air humidification accessory was designed and tested This Document Subject to Confidentiality Agreement to provide air 60% relative humidity to the animals. Puff pressure profiles were measured and compared with small sampling of human puff profiles. New analytical methods for analysis of specific smoke components in smoke inhalation exposure atmospheres are presently being used to investigate smoke composition in the Walton exposure chamber and the possible effect of July animals on smoke composition. It is anticipated that most proposed studies on the Walton smoking machine will be completed by the end of this contract period (July, 1976). Oral presentations of studies on the Walton have been made, and papers for open literature publication are being prepared. Work on the Lorillard LACS II exposure system has been minimal during this contract period due to failure of electronic components in the system. Major problems remain with the system in the design of the smoke distribution valve and construction of electronic components of the system. Cost estimates were obtained for upgrading the electronic components of the system for possible future use by CTR. At our recommendation and with approval of CTR, efforts on the LACS II were redirected to expedite development of the SEM exposure system. III. MAJOR PROJECT ACTIVITIES FOR THE PERIOD JANUARY, 1975 - JANUARY, 1976 A. Characterization and Design Improvements in the P&I SEM Exposure System 1. It has been found that significant differences in puff volumes are possible for cigarettes smoked on the SEM. Experiments with unlit cigarettes show that the puff volume depends on the airflow resistance of the cigarette, which can be highly variable from cigarette-to-cigarette and puff-to-puff. (The SEM is a constant pressure smoking machine whereas most other smoking machines are constant volume puffing which assures puff volume consistency.) Direct measurement of puff volumes of individual lit cigarettes on the SEM is not yet possible, but calculations show that puff volume variabilities as high as ±10ml may be obtained. Methodology for direct measurement of the puff volumes on individual, lit cigarettes is under development to further investigate this potentially serious shortcoming of the new model (SEM II) exposures system. This Document Subject to Confidentiality Agreement 2. The relative humidity of air in the dome area of the SEM system was found to decrease to below 10% during exposures because dry, pressurized air was used for the dome air supply. Because of the possible effects of low humidity on the burning rate of cigarettes, the chemical composition of the smoke, and deposition Humidity of smoke particulates in the animals, several methods for humidifying the dome air supply were investigated. A humidification system which involved forcing the air supply through a water saturator proved satisfactory and is included in the final model (SEM II) of the system. 2. A light scattering device has been designed and built for continuous monitoring of tobacco smoke particulates in inhalation exposure systems. The device is useful for studies aimed at establishing the uniformity of smoke in exposure systems, the age of the smoke when it reaches the exposure site, puff time, and other exposure conditions. The device has been used to study the spatial and time distribution of smoke in the Walton Exposure chamber (section B-2). Tests made with the device show that the response is linearly related to smoke particulate concentration and that This Document Subject to Confidentiality Agreement the response is independent of cigarette type and apparently depends only on the particulate matter concentration in the exposure chamber (Figure 8). There are numerous applications of this device to characterize the Walton and SEM exposure systems. Current plans call for utilization of the device as a smoke monitor in the SEM II exposure system at Microbiological Associates to prevent accidental deaths of animals due to exposure to toxic levels of smoke. 3. A new method for sampling smoke from inhalation exposure systems has been developed. Previous methods used by us were designed for smoke sampling from static (Walton) exposure system and are not applicable to sampling of continuos streams of smoke as generated by the SEM and LACS II smoking machines. In the new method, smoke is sampled by a motor-driven syringe which removes smoke at a selected, constant rate (0.1-40 ml/min). Initiation and termination of sampling is controlled by two timers which supply power to the syringe drive. The two timers permit sampling for a selected interval during any part of the exposure cycle. The timers can be by-passed to provide continuous sampling over the entire exposure cycle. Smoke is withdrawn from the exposure system through a micro Cambridge filter pad which retains smoke particulates, and the gas phase is collected in the glass syringe. Analysis are then made on either or both smoke fractions to determine constituents of interest. This sampling method is being used to sample smoke from the Walton and SEM exposure system to establish smoke concentration and composition (section B-4). IV. Presentation of Results A. Oral Presentations 1. "Chemical Indicators and Methods for Determination of Tobacco Smoke Concentration in Inhalation Exposure Systems", J. R. Stokely, J. H. Moneyhun, and M. R. Guerin, presented at the 29th Tobacco Chemists' Research Conference, College Park, Maryland, October 8-10, 1975. This Document Subject to Confidentiality Agreement 2. "Depletion of Constituents in Contained Smoke Aerosols for Inhalation Exposure Dosimetry", J. E. Caton, J. R. Stokely, and M. R. Guerin, presented at the 29th Tobacco Chemists' Research Conference, College Park, Maryland, October 8-10, 1975. 3. "Organic Gas Phase Composition of Contained Smoke Aerosols Used for Inhalation Testing", C. E. Higgins, J. R. STokely, and M. R. Guerin, presented at the 29th Tobacco Chemists' Research Conference, College Park, Maryland, October 8-10, 1975. 4. "Electron Microscope Measurement of Methyl Cyanoacrylate Fixed Tobacco Smoke Particles", R. W. Holmberg, J. R. Stokely, and M. R. Guerin, presented at the 29th Tobacco Chemists' Research Conference, College Park, Maryland, October 8-10, 1975. 5. "Chemical and Physical Properties of Smoke in Inhalation Exposure System", J. R. Stokely, presented at NCI Contractors' Conference, Atlantic City, N. J., July 17-18, 1975. B. Reports 1. "Initial Evaluation of a Prototype Animal Containment Unit for the P&I SEM", J. H. Moneyhun and J. R. Stokely, CTR/ORNL Topical Report #1 - 9-24-75. 2. "Pressure Measurements in a Prototype Animal Containment Unit for the P&I SEM", J. H. Moneyhun and J. R. Stokely, CTR/ORNL Topical Report #2 - 9-29-75. 3. "Observations and Suggestions on the Prototype Animal Containment Unit for the P&I SEM", J. H. Moneyhun and J. R. Stokely, CTR/ORNL Topical Report #3 - 11-3-75. 4. "Monitoring Smoke Concentration in the Walton Exposure Chamber Using a Light-Scattering Sensor", C. E. Higgins, T. M. Gayle, and J. R. Stokely, CTR/ORNL Topical Report #4 - 1-23-76. This Document Subject to Confidentiality Agreement TABLE I LOSSES OF GAS PHASE COMPONENTS FROM SMOKE PASSING THROUGH TUBING OF VARIOUS MATERIALS Tubing length: 10 ft, smoke generated from 1A1 cigarettes by P&I SEM Smoking machine, flow rate through tubing: 10.5 1/minute - 10% smoke and 1.05 1/minute - 100% smoke. Smoke sampled at entrance and exit of tubing by pulse sampling with gas sampling valve and transferred to gas chromatograph. High resolution chromatogram obtained under standard conditions. 10. Techniques will be developed to measure carbon dioxide produced by CO2 in SEM ? ? animals undergoing smoke exposure on the SEM II. This technique will be utilized in dosimetry studies (Part II, this proposal) to determine if carbon dioxide production by the animals can be empirically correlated with the dose of smoke particulates deposited in the respiratory tract. If this method proves valid, then it will be possible to continuously monitor dosimetry in mice and chronic inhalation exposures with the SEM II. 11. A collaborative effort will be initiated with Microbiological Associates in the selection and quality control of cigarettes used in chronic mouse exposure experiments. Cigarettes considered for use in the exposures will be analyzed for smoke constituents deemed to This Document Subject to Confidentiality Agreement be important and for which routine analysis methods are available. Quality control procedures will be devised and methodology will be made available to Microbiological Associates to provide quality control of cigarettes used in inhalation studies. III. REPORTING OF RESULTS Informal reporting of the status and experimental results derived from this project will be made to the CTR project officer at monthly intervals. CTR/ORIL Topical Reports will be prepared and submitted to CTR when deemed appropriate by the principle investigator or CTR project officer. A comprehensive report of work carried out under this contract will be prepared and submitted to CTR upon completion of experimental studies. Results of experimental studies, if deemed appropriate by in-house review, will be submitted for open literature publication or oral presentation. CTR will be informed before publication or presentation of experimental findings. Selected topics resulting from this working will be included in the Tobacco Smoke Research Program Annual Progress Report published by ORNL. IV. STAFFING AND BUDGET Man Year Allocation Name and Function 0.50 J. R. Stokely (Ph.D); Principle Investigator, experimental planning and design. 0.75 R. W. Holmberg (Ph.D); Development of particle size distribution methodology and application to exposure systems. 1.00 L. B. Yeatts (Ph.D); Routine smoke composition studies, sampling and analysis for smoke concentration measurements. 1.00 J. H. Moneyhum (B.S.); Operational studies on SEM 11, evaluation and design of animal containment unit, chemical analysis development and application 0.75 C. E. Higgins (B.S.); High resolution gas chromatographic analysis, applied studies with continuous smoke concentration monitors, routine chemical analysis. 0.25 T. M. Gayle (B.S.); Mechanical and electronic design and construction. 1.00 D. D. Pair (Tech); Routine chemical analysis, animal care and handling. This Document Subject to Confidentiality Agreement Cost Distribution 1. Salary and Overhead (229,000). To include 2.25 man-yrs Ph.D chemist, 2.0 man-yrs E. S. chemist, and 1.00 man-yr non-degreed analyst. 2. Materials and Supplies (16,000). To include normal in-house stores supplies and outside procurement of consumables. Major items include: (a) Gas chromatographic supplies (carrier and detector fuel gases, syringes, column packings, tubings, fittings, chart paper, septums, etc.). (b) Chemicals (standards, tracers, solvents, eluent, reagent, etc.). (c) Smoke generation and collection (filter pads, filter assemblies, cold traps, saran/tedlar bags, cigarette conditioning vessels, etc.). (d) Components and raw materials (valves, timers, switches, pumps, piping, instrument components, etc.). 3. Construction and Maintenance (20,000). To include in-house fabrication of items under development or not commercially available and maintenance of existing instrumentation. Major items include: (a) Monitoring systems (particulate sensor, chemical sensors). (b) Sampling systems (continuous and pulse). (c) Smoke generation systems (for comparisons of push/pull puffing, to generate reference data, to study TPM/gas phase distributions, etc.). (d) Cigarette loader (to prepare cigarettes doped with specific constituents). (e) Particle size (sampling and filtration systems) (f) Routine maintenance. 4. Travel (7,000). To include trips to Microbiological Associates, Process and Instruments, and participation in CTR-USA, Inc. contractors meeting, NCI Smoke Inhalation Bioassay workshop and Tobacco Chemists Research Conference. 5. Miscellaneous (5,000). To include in-house analytical services, technical information services to prepare materials for publication and presentation, computing services to assist data management. This Document Subject to Confidentiality Agreement 6. Unusual Costs (13,000). (a) Pressure transducer - for continuous measurement of puff volumes and dome pressure on SEM II (1,000). (b) Non-dispersive Infra-red Analyzer - for Carbon Dioxide Analysis on SEM animal containment unit (5,000). (c) Rapid response-time strip chart recorder - for use with item (a) and (b) (1,500). (d) Harvard Infusion - Pump - for sampling SEM and Walton exposure systems (1,500). (e) Sulfur and Nitrogen Specific Gas Chromatographic Detectors - for analysis of hydrogen cyanide and hydrogen sulfide (4,000) Cost Summary Dollars 1. Salary and Overhead $229,000 2. Materials and Supplies 16,000 3. Construction and Maintenance 20,000 4. Travel 7,000 5. Miscellaneous 5,000 6. Unusual Costs 13,000 $290,000