Propylene Glycol excerpted from NTP-CERHR expert panel on the reproductive and developmental toxicity of propylene glycol - The Center For The Evaluation Of Risks To Human Reproduction (CERHR) NTP-CERHR-PG-03 May 2003

The National Toxicology Program (NTP) and the National Institute of Environmental Health Sciences (NIEHS) established the NTP Center for the Evaluation of Risks to Human Reproduction (CERHR) in June 1998. The purpose of the Center is to provide timely, unbiased, scientifically sound evaluations of human and experimental evidence for adverse effects on reproduction, including development, caused by agents to which humans may be exposed.

Propylene glycol was selected for evaluation by the CERHR based on its high production and widespread public exposure due to its use as an antifreeze and de-icing agent, as well as its use in paints, coatings, foods, drugs, and cosmetics.

This evaluation results from the efforts of a nine-member panel of government and non-government scientists that culminated in a public expert panel meeting held February 11-13, 2003. This report has been reviewed by CERHR staff scientists and by members of the Ethylene Glycol / Propylene Glycol Expert Panel. Copies have been provided to the CERHR Core Committee, which is made up of representatives of NTP-participating agencies. This report is a product of the expert panel and is intended to (1) interpret the strength of scientific evidence that propylene glycol is a reproductive or developmental toxicant based on data from in vitro, animal, or human studies, (2) assess the extent of human exposures to include exposures of the general public, occupational groups, and other sub-populations, (3) provide objective and scientifically thorough assessments of the scientific evidence that adverse reproductive/developmental health effects may be associated with such exposures, and (4) identify knowledge gaps to help establish research and testing priorities to reduce uncertainties and increase confidence in future assessments of risk.

The Expert Panel Report on Propylene Glycol will be a central part of the subsequent NTP CERHR Monograph. The monograph will include the NTP CERHR Brief, the expert panel report, and all public comments on the expert panel report The NTP CERHR Monograph will be made publicly available and transmitted to appropriate health and regulatory agencies.

The NTP-CERHR is headquartered at NIEHS, Research Triangle Park, NC and is staffed and administered by scientists and support personnel at NIEHS and at Sciences International, Inc., Alexandria, Virginia.

Reports can be obtained from the website (http://cerhr.niehs.nih.gov) or from:

Michael D. Shelby, Ph.D., NIEHS EC-32 PO Box 12233, Research Triangle Park, NC 27709, 919-541-3455 shelby@niehs.nih.gov

This report is prepared according to the Guidelines for CERHR Panel Members established by NTP/NIEHS. The guidelines are available from the CERHR web site (http://cerhr.niehs.nih.gov/). The format for Expert Panel Reports includes synopses of studies reviewed and an evaluation of the Strengths/Weaknesses and Utility (Adequacy) of the study for a CERHR evaluation. Statements and conclusions made under Strengths/Weaknesses and Utility evaluations are those of the Expert Panel and are prepared according to the NTP/NIEHS guidelines. In addition, the Panel often makes comments or notes limitations of the study in the synopses. Bold, square brackets are used to enclose such statements. As discussed in the guidelines, square brackets are used to enclose key items of information not provided in a publication, limitations noted in the study, conclusions that differ from authors, and conversions or analyses of data conducted by the Panel.

The Chemical Abstracts Service Registry Number (CAS RN) for propylene glycol is 57-55-6. Synonyms or trade names for propylene glycol include: 1,2-Propanediol; 1,2-Dihydroxypropane; Methylethylene glycol; Trimethyl glycol; 1,2-Propylene glycol; monopropylene glycol; propane- 1,2-diol; alpha-propylene glycol; dowfrost; PG 12; sirlene; solar winter ban; propanediol (1); 2- dihydroxypropanol; methylethyl glycol; methyl glycol; 2,3 propanediol; and alpha propylene glycol (2). The American Chemistry Council (ACC) (3) stated that the name Sirlene is no longer used.

Figure 1-1. Formula and Molecular Weight (mw) of Propylene Glycol. Physicochemical properties are listed in Table 1-1.

According to the ACC (3), impurities of propylene glycol include chlorides (1 ppm max), iron (1.0 ppm max), water (0.2 wt% max), and dipropylene glycol (<0.2%).

Manufacturers of propylene glycol are The Dow Chemical Company, Freeport, TX and Plaquemine, LA; Lyondell Chemical Company in Pasadena, TX; Huntsman Corporation in Port Neches, TX; and Arch Chemicals, Inc., in Brandenburg, KY (3).

Commercial propylene glycol is manufactured by direct hydrolysis of propylene oxide by water (5). Propylene oxide is made using the chlorohydrin process where the propylene oxide is recovered as a pure product before conversion to the glycol. In 1999, 1,083 million pounds of propylene glycol were produced in the U.S. with apparent consumption of 854 million pounds (5).

Of the 854 million pounds of propylene glycol consumed in the U.S., uses included (in million pounds and % wt) as a chemical intermediate in the manufacture of unsaturated polyester resins (228, 26.7%), cosmetics and personal care products, pharmaceuticals, and human food (170, 19.9%), liquid detergents (135, 15.8%), deicing fluids (85, 10%), antifreeze/engine coolant (55, 6.4%), paints and coatings (40, 4.7%), tobacco humectant (25, 2.9%), other fluids (32, 3.8%), and other applications (84, 9.8%) (5). Propylene glycol is also used in the production of plasticizers (e.g., polypropylene adipate), 2-methylpiperazine, 1,2-propylene diamine, hydroxylated polyester, polyester-type fluorescent resin matrix, and polyether polyols (2).

The following summary obtained from the Agency for Toxic Substances and Disease Registry (ATSDR) (4) and the Hazardous Substances Data Bank (HSDB) (2) provides information about propylene glycol uses and exposures:

Propylene glycol is used as a softener for cellulose films in the United Kingdom (2, 6). Propylene glycol is Food and Drug Administration (FDA) approved for use in food, tobacco, and pharmaceutical products as an inert ingredient (7). It is considered to be generally recognized as safe (GRAS) for direct addition to foods (7). GRAS substances, such as propylene glycol, are also permitted in packaging materials as long as the substances “are used in amounts not to exceed that required to accomplish their intended physical or technical effect” (7). Inert ingredients are required to be listed in over-the-counter drugs (8).

Propylene glycol is a humectant in pet food products, but not in cat foods. Because of the sensitivity of the cat erythrocyte to Heinz body formation (denatured proteins, primarily hemoglobin) by propylene glycol and the possibility of inducing anemia in cats, propylene glycol was removed from cat food products (semi-moist cat food) by the FDA in 1996 (9).lycol.

1.2.3 Occurrence

Propylene glycol is released into the environment from industrial disposal and from consumer products containing this chemical. Airports are required by the Environmental Protection Agency (EPA) (10) to monitor storm water runoff and to recycle deicing solutions. Propylene glycol is water-soluble and has the potential to leach into groundwater, but is rapidly degraded. The half-life of propylene glycol in water is estimated to be 1−4 days under aerobic and 3−5 days under anaerobic conditions (4). No information was found on this compound in any environmental medium. Propylene glycol was not listed as an organic wastewater contaminant in a recent report by Kolpin et al. (11).

1.2.4 Human Exposure

1.2.4.1 General Population Exposure

The general population can be exposed to propylene glycol through dermal contact with consumer products such as cosmetic products, antifreeze solutions, coolants, windshield deicers, or pharmaceutical creams. Oral exposure to propylene glycol can occur through its use in food and tobacco products and as a solvent for pharmaceutical products (2). In Japan, average daily intake of propylene glycol as a food additive has been reported to be 43.0 mg/person [43 mg/60kg = 0.71 mg/kg bw/day] (Louekari et al. (12) [from Market Basket Study, Japan 1982]).

Data for per capita daily intake of propylene glycol in food products have been estimated for the United States in a recent report by the United Nations Joint Food and Agriculture Organization/World Health Organization (FAO/WHO) Expert Committee on Food Additives (13). In reviewing the annual volume of production of 31 flavoring agents, propylene glycol per capita consumption was estimated at 2,400,000 µg/day [34.28 mg/kg bw/day for a 70 kg person]. (This value was based upon the 1995 update of data collected since 1972 by the Flavor and Extract Manufacturers’ Association.)

In a review by the Cosmetic Ingredient Review Expert Panel (14), data on the percent concentration and use of propylene glycol in cosmetics was summarized; these data are presented in Table 1-2. These data were based upon information provided to the FDA in 1984 on propylene glycol use in cosmetic formulations and consisted of a total of 5,676 cosmetic products in 74 categories, with 2,597 product formulations containing between 1 and 10% propylene glycol.

Table 1-2. Product Formulation Data for Propylene Glycol (Adapted from Cosmetic Ingredient Review (14)).

Propylene glycol is rapidly degraded in water and CERHR was unable to locate any information on propylene glycol in drinking water.

Propylene glycol may be released by some carpeting (2). In a technical study by Hodgson et al. (15), emissions of volatile organic compounds (VOC) from four different types of new carpets were measured. Exposure chamber air samples were collected onto multisorbent samplers packed with Tenax-TA, Ambersorb XE-340, and activated charcoal, in series. The chemicals were thermally desorbed from the sampler, concentrated, and injected into a capillary gas chromatograph with a mass spectrometer used as a detector. One carpet with a polyvinyl chloride backing emitted propylene glycol, vinyl acetate, formaldehyde, isooctane, and 2-ethyl-1-hexanol. Propylene glycol and vinyl acetate had the highest concentrations and emission rates for this carpet. The estimated emission rates ranged from 690 µg/m²/hr 24 hours after installation to 193 µg/m²/hr at 168 hours after installation. The other three carpet types did not emit propylene glycol.

The FDA estimated that the human daily dietary intake of propylene glycol to be a ‘few mg per kg [body weight] per day’ (16). [No details were given on how exposures were estimated.] In a 2002 report by the United Nations Joint FAO/WHO Expert Committee on Food Additives (13), per capita consumption of propylene glycol in the United States was estimated at 2,400,000 µg/day [34.28 mg/kg bw/day for a 70 kg person]. The average daily dietary intake of propylene glycol in Japan was estimated to be 43 mg/person [0.7 mg/kg bw/day based on a 60 kg person] (12). The WHO food additive series (17) lists the acceptable human daily intake of propylene glycol at <25 mg/kg bw/day.

Propylene glycol is used in some pharmaceuticals that are administered intravenously (Table 2-8). This represents a unique exposure route for certain subpopulations.

Occupational exposure to propylene glycol may occur through direct dermal contact while handling products containing this compound or through inhalation of airborne propylene glycol resulting from heating or spraying processes (2).

Neither the Occupational Safety and Health Administration (OSHA) nor the American Conference of Governmental Industrial Hygienists (ACGIH) has established exposure limits for propylene glycol vapors. No Threshold Limit Value (TLV) has been defined for propylene glycol, but an American Industrial Hygiene Association (AIHA) Workplace Environmental Exposure Level (WEEL) guide of 50 ppm (total exposure) and inhalation aerosol exposure of 10 mg/m³ has been determined (18).

A 1981-1983 National Occupational Exposure Survey (NOES) of U.S. workers led NIOSH to estimate that 1,748,454 people were potentially exposed to propylene glycol at the workplace (2). Ninety-eight percent of exposures are with trade name products containing propylene glycol, rather than in the production of propylene glycol itself (2).

Norbäck et al. (19) studied the exposure of Swedish painters to VOCs from indoor application of water-based paints. VOCs were sampled on different sorbents within the personal breathing zone of the painter and analyzed by gas chromatography (GC)/mass spectroscopy (MS). Propylene glycol was one of the VOC constituents measured. Exposure measurements for propylene glycol were taken over a 1-hour period of water-based paint application for 20 batches of paint from 5 different manufacturers. Propylene glycol was detected in 12 of the 20 samples. Personal exposure to propylene glycol during application of water-based paints yielded a geometric mean of 350 µg/m³ with a maximum value of 12,700 µg/m³.

Laitinen et al. (20) examined exposure to ethylene and propylene glycol in Finnish motor servicing workers. Ten male mechanics from five different garages participated in the study. The only protective equipment used by some workers was leather gloves. Ten age-matched male office workers served as controls. Differences between groups were evaluated by Student’s t-test. Air concentrations of ethylene glycol and propylene glycol were measured during the entire shift. Neither ethylene glycol nor propylene glycol vapors were detected in the breathing zones of workers; detection limits for each compound were given as 1.9 cm³/m³ and 3.2 cm³/m³, respectively. Urine samples were collected after the work shift and analyzed for ethylene glycol, oxalic acid, and propylene glycol [method of urine collection, storage, and extraction and quality control not reported]. There were no differences found between controls and propylene glycol-exposed mechanics.

Deicing fluids are low viscosity glycols used to remove ice or snow that would increase drag on the aircraft. The antifreeze components in a deicing solution vary with the manufacturer, usage, and environmental conditions. Commercial Type I fluid is applied hot as a mixture of fluid and hot water to deice the exterior of aircraft. Type IV fluids are usually applied after the aircraft is deiced to keep ice from reforming. Approximately 90% of Type I fluids and 50% of Type IV fluids are propylene-glycol based (3, 5). Performance criteria for deicing fluids are governed by specifications of the Aerospace Division of the Society of Automotive Engineers (SAE) (21). Both inhalation and dermal exposures to workers using deicing solutions can occur.

The levels of propylene glycol in aircraft deicing workers (n=7, age 31-52 years, sex not given) using either undiluted or water-diluted propylene glycol heated to 60°C was measured in urine

samples collected pre- and post-shift (22). Workers were wearing coats, rubber gloves, and masks. The detection limit for the method used to measure propylene glycol in urine was 20 µg/L. Urine samples were also collected from a comparison group of non-exposed persons (n=16, sex and age not given). For the exposed workers, the median pre-shift urine level was 1.49 mg/L (range 0.72-13.44 mg/L) and 1.67 mg/g creatinine (range 0.41-10.58 mg/g creatinine) and the median post-shift urine level was 2.07 mg/L (range 0.77-9.04 mg/L) and 2.46 mg/g creatinine (range 1.22-10.27 mg/g creatinine). Propylene glycol concentrations in the post-shift worker urine samples were only slightly higher than those of the unexposed comparison group.

In a study simulating concentrations of propylene glycol mist used in aviation emergency training, Wieslander et al. (23) concluded that short (1 minute), high exposure (geometric mean concentration of 309 mg/m³, range 176-851 mg/m³) to propylene glycol mist may cause acute ocular and upper airway irritation. The duration of these effects was not measured, as measurements were taken within 15 minutes of exposure.

A Health Hazard Evaluation (HHE) on occupational exposure to propylene glycol during aircraft deicing operations was conducted by NIOSH (24). Evaluation of deicing procedures was conducted at the Denver International Airport (DIA) in March 1996. At DIA, United Airlines uses a 50% solution of propylene glycol in water, heated to 180° F for deicing aircraft. Trucks with dual 800-gallon tanks, spray hoses, and booms are used. The amount of fluid used for deicing each plane ranges from 50 to 200 gallons. Personal breathing-zone air samples were collected from six ground sprayers, one basket man, and one truck driver. Air samples were collected on XAD-7 OVS tubes at a flow rate of 0.5 L/min for 6 hours and analyzed by GC/MS for propylene glycol according to NIOSH Method 5523. Seven workers (Table 1-3) had a range of exposures from 10 to 21 mg/m³ with a mean of 15 mg/m³, based on a 6-hour collection.

*Air sample was visibly contaminated with liquid propylene glycol. This was caused by a worker being accidentally sprayed with the deicing fluid during sampling.

The author concluded that "there was no hazard from overexposure to deicing fluid. . . .Airborne exposure to propylene glycol was low and propylene glycol has low toxicity."

Propylene glycol does not bioaccumulate in organisms and rapidly biodegrades in the soil and in water (25). However, this process is oxygen-demanding and can deplete dissolved oxygen levels in water (26). The Clean Water Act requires airports to implement plans for deicer management to control storm water contamination. Therefore, airports must monitor propylene glycol storm water runoff and scavenge and recycle deicing solutions (10).

Limited human exposure data for propylene glycol were available for Expert Panel review. The utility of the occupational exposure data available is limited by either the small sample size or a high proportion of non-detected values. Estimates of propylene glycol workplace exposures are based on a 1981-1983 NOES of U.S. workers and may not reflect current occupational exposure. These data are insufficient to evaluate occupational exposure to propylene glycol.

An estimate of U.S. consumer exposure was available from a 2002 report by the United Nations Joint FAO/WHO Expert Committee on Food Additives (13). In reviewing the annual production volume of 31 flavoring agents, per capita consumption of propylene glycol was estimated at 2,400,000 µg/day [34.28 mg/kg bw/day for a 70 kg person]. This value exceeded the estimated per capita consumption in Japan (1982) by approximately 50-fold [43 mg/60 kg = 0.71 mg/kg bw/day]. These estimates of human exposure are for food products and do not include exposure from pharmaceutical products or exposure through inhalation. Propylene glycol is found in many pharmaceuticals that are administered intravenously. There are limited data on the effects and exposure levels of chronic (intravenous) administration of propylene glycol in infants and children and no information was found on chronic exposure in pregnant women.

In 1999, 1,083 million pounds of propylene glycol were produced in the U.S. with apparent consumption of 854 million pounds (5). Of the apparent amount consumed, uses included, in million pounds and percentages, unsaturated polyester resins (228, 26.7%); cosmetics and personal care products, pharmaceuticals, and human food (170, 19.9%); liquid detergents (135, 15.8%); deicing fluids (85, 10%); antifreeze/engine coolant (55, 6.4%); paints and coatings (40, 4.7%); tobacco humectant (25, 2.9%); other fluids (32, 3.8%); and other applications (84, 9.8%) (5). Propylene glycol is approved by the FDA for use in food, tobacco, and pharmaceutical products and has GRAS status for direct addition to foods.

The general population is exposed to propylene glycol by oral intake, dermal contact, and inhalation. The average daily intake of propylene glycol from food products in the United States has been estimated at 2,400 mg/day [34 mg/kg bw/day for a 70 kg person] (13). In Japan, the estimated average daily intake of propylene glycol as a food additive was reported to be 43 mg per person [43 mg/60 kg=0.71 mg/kg bw/day] (Louekari et al. (12) [from Market Basket Study, Japan 1982]). The Joint FAO/WHO Expert Committee on Food Additives (13) concluded that "the safety of these substances [propylene glycol and propylene glycol stearate] would . . . not be expected to be of concern." Since propylene glycol has GRAS status and may not be listed as a specific ingredient in some foods, dietary intake based upon product labeling would result in an underestimation of intake. Propylene glycol is an inert ingredient in some pharmaceutical preparations and is also found in many pharmaceuticals that are administered intravenously, which represents a unique exposure route for certain subpopulations.

Occupational exposure to propylene glycol may occur through dermal contact or through inhalation of airborne propylene glycol from heating or spraying processes. No TLV has been defined for propylene glycol, but an AIHA WEEL guide of 50 ppm (total exposure) and an inhalation aerosol exposure of 10 mg/m³ have been determined. NIOSH estimated that 1,748,454 people (1981-1983 NOES survey as cited in NIOSH report, 1983 (2)) are potentially exposed to propylene glycol in the workplace, primarily through contact with trade name products containing propylene glycol.

Several small occupational exposure studies measuring propylene glycol were located. In a study by Laitinen et al. (20), motor-servicing worker exposure to propylene glycol and ethylene glycol was measured. Propylene glycol was below the detection level in air and levels in the urine of exposed workers did not differ from urinary levels in unexposed controls. As dermal exposure to workers was not measured, it was not possible to determine whether urinary levels of propylene glycol found in the workers were due to low exposure or to low dermal absorption.

Norbäck et al. (19) measured airborne propylene glycol exposure of Swedish painters during indoor application of water-based paints. Propylene glycol was detected in 12 of 20 samples with a geometric mean of 350 µg/m³ and a maximum value of 12,700 µg/m³.

The levels of propylene glycol in aircraft deicing workers (n=7, age 31-52 years, sex not given) using either undiluted or water-diluted propylene glycol heated to 60^°C was measured in urine samples collected pre- and post-shift (22). Urine samples were also collected from a comparison group of non-exposed persons (n=16, sex and age not given). For the exposed workers, the median pre-shift urine level was 1.49 mg/L (range 0.72-13.44 mg/L) and 1.67 mg/g creatinine (range 0.41-10.58 mg/g creatinine). For the exposed workers, the median post-shift urine level was 2.07 mg/L (range 0.77-9.04 mg/L) and 2.46 mg/g creatinine (range 1.22-10.27 mg/g creatinine). For the unexposed comparison group, the median urine level was 1.35 mg/L (range 0.29-10.7 mg/L) and 1.18 mg/g creatinine (range 0.46-18.77 mg/g creatinine).

In a Health Hazard Evaluation (HHE) conducted by NIOSH on workers (n=8) using propylene glycol during aircraft deicing operations (24), personal breathing-zone air samples over a 6-hour period were collected. Seven workers had exposures ranging from 10 to 21 mg/m³ with a mean of 15 mg/m³ (1 worker sample excluded due to a suspect high value).

The toxicokinetics and metabolism data for propylene glycol were initially examined by consulting authoritative reviews (4, 27) and an independent review (28). The toxicokinetics sections in those reviews were somewhat brief, and a decision was made by CERHR to review relevant original studies in humans and studies in animals pertinent to reproductive and developmental toxicity.

Studies of the pharmacokinetics of propylene glycol in humans have been conducted primarily in conjunction with on-going patient therapy where propylene glycol was administered as a vehicle for medications.

Yu et al. (29) examined the pharmacokinetic profile of propylene glycol during multiple oral-dosing regimens. The 22 subjects were outpatients who participated in a phenytoin bioavailability study where propylene glycol was used as a solvent. In one study, 16 adults received a 20.7 g/dose 3 times daily for a minimum of 3 days. In another study, 6 individuals received a 41.4 g/dose twice daily for a period of 3 days. These oral doses were given in conjunction with 100 mg phenytoin in 7.25 mL of alcohol USP, 6 µL of Peach Flavor, 5 mL of glycerin USP, and 8 mL of 70% (w/w) fructose. Propylene glycol was rapidly absorbed from the gastrointestinal tract with maximum plasma concentrations obtained within 1 hour of dosing. The average serum half-life of propylene glycol for the study with 16 and 6 individuals was determined by the authors to be 3.8 and 4.1 hours, respectively. The average total body clearance was determined by the authors to be approximately 0.1 L/kg/hr, although there was significant variability in clearance rate among individuals. The apparent volume of distribution was determined by the authors to be approximately 0.5 L/kg, which approximates the volume of distribution of total body water (29).

Strength/Weaknesses: This study by Yu et al. (29) provides data on the oral absorption of propylene glycol as well as on serum half-life, and apparent volume of distribution and total body clearance after repeated oral doses of either 20.7 g 3 times daily or 41.4 g 2 times daily, for a minimum of 3 days. The results are in agreement with expectations for a highly water-soluble, small molecule: rapid absorption, distribution into total body water, relatively short half-life, and rapid total body clearance. One study limitation is the study subjects’ concomitant exposure to ethanol; propylene glycol and ethanol are substrates that compete for alcohol dehydrogenase in the initial step of metabolism. While the doses of propylene glycol were high, the data do indicate ready bioavailability of the chemical. The half-life estimates are generally consistent with the results of Speth et al. (30) to be discussed later.

Utility (Adequacy) for CERHR Evaluation Process: Data in the Yu et al. (29) study are generally adequate to estimate kinetic parameters, but inadequate for quantitative determination of bioavailability.

In a study using human volunteers, Kollöffel et al. (31) studied rectal absorption and other kinetic parameters in children and adults. Propylene glycol and water (1:1) were used as solvents in the formulation of a rectal solution of paracetamol. Absorption of propylene glycol through the rectum was rapid with peak concentrations obtained at 1±0.6 hour (average ±SD) in children (5-12 years old) and 1.5±0.3 hours in adults. Peak plasma concentrations were measured at 171 mg/L [2.2 mM] in 4 children dosed with 0.173 g/kg bw propylene glycol and 119 mg/L [1.6 mM] in 10 adults dosed with 8.64 g propylene glycol [123 mg/kg bw assuming a 70 kg bw]. The serum half-life was determined to be 2.8±0.7 hours in adults and 2.6±0.3 hours in children. The apparent volume of distribution was 0.79±0.30 L/kg in adults and 0.77±0.17 L/kg in children (31).

Strength/Weaknesses: Kolloffel et al. (31) determined Cmax ^and_Tmaxand then used a linear curve-fitting program to recalculate Cmax ^and_Tmax,values as well as half-life, apparent volume of distribution, and clearance after different doses of propylene glycol were administered per rectum to adults and children. The small number of children (n=4) and the age range (5-12 years) does not permit a judgment as to whether bioavailability may differ as a function of age within childhood or between children and adults. The values reported are in the expected range providing confirmatory evidence for the reliability of kinetic parameters determined by Speth et al. (30). Plasma levels in children (age 5-12 years) were only slightly higher than in adults. The half-life was virtually the same in children as in adults, which is in agreement with alcohol dehydrogenase activity reaching adult levels by the age of 5 years (32). The extent of oral absorption cannot be judged from these data but a visual inspection of plasma concentrations after intravenous (IV) infusion (30) and rectal administration (31) indicate very high bioavailability. Thus, oral bioavailability will also be very high. Although it appears that children absorb propylene glycol significantly faster and attain higher peak plasma concentration than adults, the differences are modest and of doubtful toxicological significance.

Utility (Adequacy) for CERHR Evaluation Process: The study by Kolloffel et al. (31) is useful to indirectly assess bioavailability.

There is limited information on the absorption of propylene glycol through intact human skin. In a study of human skin biopsy specimens from adults 19-50 years of age, MacKee (33) found no penetration of radioactive tracer materials after up to 1 hour permeation time using propylene glycol alone as a vehicle [visual evidence of tracer uptake into biopsied skin, but no analytical confirmation provided]. Enhancers, such as surfactants, increased absorption.

Three studies are described briefly below that involved patients with significant medical complications. In 45 patients (0.5-87 years old) with second- and third-degree burns on 21-95% of their body, propylene glycol was absorbed through skin following dermal treatment with sulfadiazine in a propylene glycol vehicle; serum levels of propylene glycol in those patients ranged from 0 to 0.98 g/dL [0 to 129 mM] (4, 34). In an 8-month-old infant with second- and third-degree burns and complicating toxic epidermal necrolysis over 78% of his body, dermal treatment with silver sulfadiazine in propylene glycol resulted in a peak propylene glycol blood level of 1.059 g/dL [139 mM] (35). A blood propylene glycol level of 0.070 g/dL [9.2 mM] in an infant was attributed to Mycostatin cream usage for diaper rash (36).

Strength/Weaknesses of Dermal Studies: The MacKee study (33) showed what is expected of a highly water-soluble substance: that dermal absorption of propylene glycol through the intact skin is very limited. Weaknesses of this study are the insensitive, non-quantitative method for assessing chemical uptake and the extensive manipulation of the skin following the permeation period (excision which apparently produced bleeding), which may have lead to losses of both skin and permeated chemical from handling the tissue. The three clinical studies (34-36) present evidence of propylene glycol bioavailability in circumstances that preclude confident extrapolation to a healthy general population. They do indicate that once the stratum corneum is impaired (removed such as in burns or irritated), dermal absorption may become a significant source of exposure.

Utility (Adequacy) for CERHR Evaluation Process: The MacKee (33) study has minimal utility for drawing conclusions regarding propylene glycol penetration across healthy human skin. However, when combined with the rat dermal penetration in vitro study (37) also showing no uptake, and given the difficulty water soluble molecules generally have penetrating the stratum corneum, the Panel concluded that the dermal absorption rate across intact skin is likely to be slow. Therefore, it can also be expected that any dermal exposure to propylene glycol will result in systemic levels far below saturation of metabolic clearance.

Bau et al. (38) [as reported in HSDB (2)] reported that less than 5% of a technetium-labeled aerosol containing 10% propylene glycol [propylene glycol not directly measured] in deionized water was taken up by humans after inhalation for 1 hour in a mist tent. The authors measured the aerosol mass median diameter to be 4.8-5.4 microns, a size small enough to have enabled penetration to the deep lung. Ninety percent of the dose was found in the nasopharynx and it rapidly entered the stomach with very little entering the lungs. Propylene glycol was not measured. The low vapor pressure (0.07 mmHg, approx equal to ~90 ppm ~270 mg/m³) of propylene glycol in combination with the short half-life before saturation of metabolism does not allow the build up of toxicologically relevant doses.

Strength/Weaknesses: Since propylene glycol was not directly measured by Bau et al. (38), absorption through the nasal mucosa cannot be determined. However, the low dose rate from inhalation exposure and the small surface area would not lead to significant absorption of propylene glycol.

Utility (Adequacy) for CERHR Evaluation Process: Since inhalation of chemicals is kinetically related to IV infusion, it is of interest to know if propylene glycol is efficiently absorbed from the lungs. As a small, water soluble molecule, it is reasonable to predict that propylene glycol would be absorbed by the lungs. However, with a low vapor pressure (0.07 mm Hg), inhalation of toxicologically relevant doses of propylene glycol is not possible unless heated to higher temperatures. Therefore, the remaining question is whether propylene glycol in a carrier medium could lead to significant exposure by inhalation. Bau et al. (38) provides a quantitative answer. Of an average of 263 mL of nebulized aerosol, 8.1 mL containing 10% propylene glycol was retained per hour, corresponding to about 0.8 g of compound, which in turn amounts to 0.09 g/kg per 8 hours. Therefore, it can be concluded that under normal conditions of exposure, propylene glycol via inhalation is of limited toxicological relevance.

Animal studies demonstrate that propylene glycol is rapidly absorbed following oral exposure. ATSDR (4) reports the findings of a study by Christopher et al. (39) in which plasma levels of propylene glycol were measured at 19.1 and 8.4 mM in 2 cats fed a diet with 12% propylene glycol [1.60 g/kg bw/day] for 5 weeks. Morshed et al. (40) found that propylene glycol blood concentration (41.04 mM) reached its maximum level 1 hour after 4 New Zealand White (NZW) rabbits were administered 38.66 mmol/kg bw [2.942 g/kg bw] as a 28.4% aqueous solution by gavage. Morshed et al. (41) orally administered an aqueous solution of propylene glycol at 4.83-77.28 mmol/kg bw [0.368-5.881 g/kg bw] to 6 male Wistar rats/group and found that absorption occurred by a first order process; time to peak absorption was related to dose and ranged from roughly 10 minutes at the low dose to 2 hours at the high dose. An older study by Lehman and Newman (42) demonstrated peak blood levels of propylene glycol approximately 2-3 hours after oral dosing in dogs.

Strength/Weaknesses of the Cited Studies: The Christopher et al. (39) study provides very limited data (one time point only) on plasma concentration of propylene glycol after repeated administration of one of two dose rates administered in the diet. It is impossible to derive any kinetic information from such a study other than the qualitative statement that propylene glycol is absorbed to some extent by the cat from the diet.

In contrast, Morshed et al. (41) provided a more complete set of data indicating dose-dependent Tmax for propylene glycol in the dose range of 0.4-5.9 g/kg. The authors did not calculate absorption half-lives or determine the extent of absorption. They concluded that gastrointestinal absorption occurred by a first order process because of the linear rise of plasma concentration at each of the five doses. [This is an improper conclusion. Data are plotted on an arithmetic scale from which calculation of kinetic rate constants is not possible. There is no indication of curve stripping to calculate kabs. The fact that elimination appears linear on an arithmetic scale indicates a zero order process. If absorption were first order, the absorption rate should increase with increasing concentration in the gastrointestinal tract. The fact that absorption rate did not increase in this manner suggests some limitation with higher bolus doses – e.g., possible delayed gastric emptying. In any case, more complete information is needed to assess bioavailability from the oral route (e.g., Vd, AUC, total body clearance rate, or a comparison IV study in rats).] The other Morshed et al. (40, 43) papers and the Lehman and Newman (42) paper also do not provide data suitable for quantitative evaluation. There are reliable quantitative data for the gastrointestinal absorption of diethylene glycol in the rat (44) with absorption half-lives ranging from 5 to 40 min (average 16 min) amounting to 80-100% of the dose. Since diethylene glycol has a higher molecular weight but comparable hydrophilicity, it is likely that very rapid gastrointestinal absorption occurs also for propylene glycol. This is also the case for ethylene glycol as indicated by rapid urinary excretion (45).

Utility (Adequacy) for CERHR Evaluation Process: Available animal data are not well suited for quantitative estimation of gastrointestinal absorption of propylene glycol. Nevertheless, all data including structure-activity relationships point toward very rapid and complete absorption. This is plausible for a highly water-soluble small molecule which will cross membranes with bulk flow of water across aqueous pores.

Information on in vivo dermal absorption of propylene glycol in animals was not located. ATSDR notes that "In vitro studies of the penetration of propylene glycol through the rat abdominal stratum corneum have been conducted" (4). Fresh abdominal skin from male Wistar rats was used in experiments in which propylene glycol, or a mixture of propylene glycol and oleic acid, were evaluated for absorption properties (46). When propylene glycol was applied for up to 2 hours, no compound was detected in the dermis. However, when 0.15 M oleic acid was added to the propylene glycol, it was detected in the dermis after 30 minutes of exposure, but not after 5 or 15 minutes (46). ATSDR (4) reported that hairless mouse skin overestimates absorption of propylene glycol by human skin while shed snake skin underestimates absorption. Therefore, the authors concluded that human skin should be used for absorption studies if possible.

Speth et al. (30) reported on the pharmacokinetics of IV administration of propylene glycol involving six cancer patients who were sufficiently healthy to care for themselves and had normal liver and kidney function. They reported that clearance decreased as dose increased over a dose-range of 3-15 mg/m². There was a first order elimination with an average terminal half-life of 2.3±0.7 hours. Considerable interpatient variation was noted. The apparent volume of distribution ranged from ¨ 0.55-0.94 L/kg. In other studies with oral or rectal exposure, apparent volumes of distribution ranged from ¨ 0.52-0.79 L/kg (29, 31).

Strength/Weaknesses: This study (30) provides sound pharmacokinetic data from a limited number of individuals who were exposed intravenously to propylene glycol. However, the Speth et al. conclusion that clearance of propylene glycol in humans occurs by a first order process is questionable, as is the calculation of an average half-life of 2.3±0.7 hours.

Utility (Adequacy) for CERHR Evaluation Process: This human study with IV exposure and those with oral and rectal exposure indicate that propylene glycol is uniformly distributed in total body water without a significant distribution to specific tissues. It can be predicted with certainty that propylene glycol will distribute into the water compartment of the placenta and fetus.

In what is considered to be the main pathway of propylene glycol metabolism in mammals (4, 39), propylene glycol is oxidized by alcohol dehydrogenase to lactaldehyde, then to lactate by aldehyde dehydrogenase. The lactate is further metabolized to pyruvate, carbon dioxide, and water. Lactate also contributes to glucose formation through gluconeogenic pathways (39). Lactate, via phosphoenol pyruvate, can be detoxified into glucose and stored as glycogen, as has been demonstrated by Wittman et al. (47) for propylene glycol in rats. Excess production of lactic acid resulting from very large exposures to propylene glycol can produce a metabolic anion gap [anion gap = (Na⁺) – (Cl^- + total CO₂)] and metabolic acidosis (4). Serum levels of >180 mg/L [2.37mM] can result in toxicity (48).

In most mammals, part of the absorbed propylene glycol is eliminated unchanged by the kidney, while another portion is excreted by the kidneys as a glucuronic acid conjugate (2, 28). The amount of propylene glycol eliminated by the kidneys has been estimated for humans at 45% (48), for dogs at 55-88% (49), and for rabbits at 24-14.2% (50). Cats do not have the ability to produce the glucuronidated metabolite (28). Alternate stereo-specific reaction pathways have

In adult humans, the mean serum half-life of propylene glycol is approximately 2-4 hours (30). Kelner and Bailey (51) studied the pharmacokinetics of propylene glycol in humans in conjunction with the IV administration of medications. Propylene glycol concentrations were measured in sera and cerebral spinal fluids (CSFs) from five patients receiving medication containing propylene glycol as a vehicle; lactate and pyruvate concentrations were also measured. The authors stated that all patients had normal hepatic and renal function based upon laboratory tests. The authors found a significant (p<0.01) correlation of lactate concentrations in the serum and CSF to the corresponding propylene glycol concentrations in these fluids. The authors concluded that although the increase in serum lactate could be due to the patients’ clinical conditions, it was unlikely in light of the correlation between propylene glycol and lactate concentrations. For two patients, the authors had propylene glycol/creatinine clearance ratios and were able to calculate the serum half-life of propylene glycol. The authors estimated this to be 4.7 and 5.6 hours, respectively, for these patients. [The dose was not stated, but because of the severe lactic acidosis, the results suggest that it must have been higher than the 2x41.4g dose/day for 3 days administered by Yu and Sawchuck (50), which did not cause lactic acidosis.]

While it is clear that total body clearance of propylene glycol occurs by metabolism and by renal excretion of the parent compound, there are no data in humans from which to assess the percentage fate of propylene glycol by these mechanisms. In the rabbit, Yu and Sawchuck (50) observed that metabolic clearance accounts for 85.8-97.6% of total clearance at lower doses. Morshed et al. (41) provided evidence that the rate-determining step in the metabolic clearance of propylene glycol in the rat is the NAD-dependent alcohol dehydrogenase. Using the dehydrogenase inhibitor pyrazole, there was a dose-dependent inhibition of the dehydrogenase leading to a dose-dependent increase in urinary excretion of propylene glycol. They found that the maximum metabolizing capacity in the rat was 8.33 mmole of propylene glycol/kg bw/hour, which they stated would extrapolate to 1.06 kg bw/day for a 70 kg human.

The Expert Panel believes that Speth et al. (30) supports the conclusion that humans clear propylene glycol similarly to rats and rabbits. However, saturation of metabolic clearance seems to occur at lower doses in humans than in rats and rabbits. Speth et al. (Table 2 of the study) (30) indicates that saturation of metabolic clearance seems to occur at a dose of about 7 g/day in some patients but not in others. Metabolic clearance does not seem to be affected at ¨ 5 g/day (although no lower dose was used to prove it conclusively) and is uniformly decreased above 12.6 g/day. Speth et al. (30) provide evidence of metabolic saturation in propylene glycol metabolism at doses of approximately 7 g/day as seen by lengthening half-life and nonlinear increases in AUC and Cmax. When this dose is converted to mmol/kg based upon the body weights reported for the three subjects receiving this dose, the value is 1.6 mmole/kg, which is considerably lower than the K_m reported by Morshed et al. in rats. Therefore, the half-life of propylene glycol before saturation of metabolic clearance when it would occur by a first order process is 1.6±0.2 (±SD) hours. This increased to above 3 hours after metabolic saturation of doses above 12 g/day, when metabolic clearance occurs by a zero order process. This is confirmed by Yu et al. (29) who found a "terminal elimination" half-life of ¨ 4 hours in patients administered even higher doses (3 x 20.7 and 2 x 41.4 g/day) of propylene glycol. Unlike the half-life of a compound cleared by a first order process, which is constant, the half-life of a chemical cleared by a zero order process is dose-dependent as is amply documented for propylene glycol.

Synthesis of propylene glycol results in a 1:1 ratio of D and L stereoisomer forms. There is some, although incomplete, information in the literature about stereospecificity of the enzymes in the propylene glycol metabolic pathways (Figure 2-1). In the main metabolic pathway, D and L forms of lactaldehyde and lactate are formed (4, 39). In the horse and rabbit, ADH will oxidize the L form of propylene glycol and lactaldehyde more efficiently than the D form (52). L-lactic acidosis has been observed in both humans and animals following exposure to propylene glycol (39, 40).

The conversion of lactaldehyde to methylglyoxal by ADH and then to D-lactate by glyoxalase and reduced glutathione is thought to be an alternate route of metabolism (Figure 2-1). D-lactate is cleared more slowly than L-lactate and is considered a poor substrate for gluconeogenesis.

Methylglyoxal synthetase can convert the substrate, dihydroxyacetone phosphate, to methylglyoxal. However, in conditions where ketone levels are high, such as diabetes or starvation, methylglyoxal synthetase activity is increased, producing more methylglyoxal and D-lactate. Excessive production of D-lactate may result in its accumulation, especially in the brain, which has a low level of catabolizing enzymes (39). Therefore, in cases of ketosis, excess levels of D-lactate may be exacerbated by propylene glycol.

In a third possible metabolic pathway, propylene glycol can be phosphorylated, converted to acetol phosphate, lactaldehyde phosphate, lactyl phosphate, and lactic acid (see Figure 2-2) (49). Metabolism of D and L forms of propylene glycol in this pathway is species-specific. The rabbit converts the L-form of phosphorylated propylene glycol to lactic acid, whereas the rat and mouse can convert both forms (52, 53).

A limited number of studies were summarized in detail since they demonstrate evidence of in vitro stereospecificity of ADH (52), L-lactatemia in rabbits (40), and increased D-lactate formation in cats (39).

Stereospecificity of ADH was studied by Huff (52). In vitro rabbit liver ADH K_s values were obtained for ethanol, L-propylene glycol, and D-propylene glycol substrates and were 0.63, 3.6, and 33.3 µmoles/mL, respectively. K_s values obtained for acetaldehyde, L-lactaldehyde, and D-lactaldehyde were 3.6, 1.4, and 3.7 µmole/mL, respectively. A similar trend in values was observed with horse liver ADH. Therefore, ADH from horse and rabbit liver exhibited stereospecific preference for L-propylene glycol and L-lactaldehyde.

Strength/Weaknesses: Stereospecificity of metabolism should be considered because technical grade propylene glycol contains the stereoisomers in a 1:1 ratio. Huff (52) determined the K_m values for oxidation of the D- and L-forms by alcohol dehydrogenase and found that L-propylene glycol is 5-9 times more readily metabolized to L-lactaldehyde by rabbit and horse alcohol dehydrogenase than is the D-form. Therefore, it is plausible that D-propylene glycol will be cleared more slowly since this is the rate-determining step in the metabolic clearance of these compounds. Moreover, accumulation of D-lactate has been documented in cats (39) and humans (54), which was partially attributed to D-lactate being a poor substrate for gluconeogenesis, a detoxification pathway for L-lactate. In addition, D, L-lactaldehydes are oxidized to methyl glyoxal with loss of the chirality center, which glyoxylase with GSH as co-substrate converts stereospecifically to D-lactate.

Another pathway occurs by phosphorylation of propylene glycol followed by oxidation steps without loss of the chirality center. Here, species differences were found; rabbits converted the L-form more readily to lactic acid, but rats and mice did it equally well with both forms (52, 53). Due to incomplete time-point sampling and a lack of quantitative numbers regarding fluxes through the different pathways, it is not possible to put together a complete picture of stereospecific metabolism of D, L-propylene glycol.

It is of no toxicological consequence whether L- or D-lactatemia develops because both can contribute to the development of lactic acidosis. The longer half-life of D-lactate can be easily

factored in via the Michaelis-Menten equation into a physiologically based pharmacokinetic (PBPK) model. The weakness of this approach is that D-lactate was shown to be efficiently utilized in man (54), but its tubular reabsorption was shown to be retarded, particularly at higher concentrations (>3 meq/L). Since chirality is lost during oxidation of D, L-lactate, the preferential use of L-lactate must be due to a lower K_m of lactate dehydrogenase for L- than for D-lactate. In any event, reduced tubular reabsorption enhances overall clearance of D-lactate, whereas reduced utilization for gluconeogenesis runs counter to this effect, apparently outweighing both its reduced tubular reabsorption and its utilization in the Krebs cycle that produces CO₂.

The overall conclusion from all data is that acute exposure to D, L-propylene glycol can cause L-lactic acidosis (if the dose is very high) due to the more rapid biotransformation (alcohol dehydrogenase being the rate-determining step) of L-propylene glycol to L-lactate, whereas subchronic/chronic exposure leads to D-lactic acidosis due to accumulation of D-lactate derived from the glyoxylase/GSH pathway and from being a poor substrate for gluconeogenesis.

Utility (Adequacy) for CERHR Evaluation Process: The database is sufficient to understand and predict metabolic clearance of D, L-propylene glycol in man.

The role of propylene glycol metabolism in lactatemia in the rabbit was investigated by Morshed et al. (40). Propylene glycol was administered to NZW rabbits by gavage in a single dose of 38.66 mmol/kg [2.942 g/kg] (1 mL 28.4% (v/v)) aqueous solution per 100 g bw. Whole blood was withdrawn from the marginal ear vein after a 24 hour fast and at 0.25, 1, and 3 hours after administration of propylene glycol. Blood pH and the levels of propylene glycol and D- and L-lactate and pyruvate were determined. The level of propylene glycol was estimated colorimetrically and the levels of lactate and pyruvate were estimated enzymatically. Data were evaluated by analysis of variance for repeated measures and were expressed as mean ± SD; a value of P < 0.05 was statistically significant. As noted in Table 2-1, blood propylene glycol concentrations were at a maximum 1 hour post-dosing. Treatment with propylene glycol significantly (P<0.01) increased the concentration of L-lactate, which reached a plateau at 0.25 hours following exposure. D-lactate levels were significantly increased and reached maximum concentration at 3 hours after administration of oral propylene glycol. Although significant, the authors considered the increase in D-lactate to be negligible and noted that L-lactate levels were similar to total lactate levels. Levels of pyruvate remained unaffected before and after administration of propylene glycol. Blood pH was not significantly altered when compared to control values. The authors note that these findings are different than the results from oral administration of propylene glycol to the rat (55).

Strength/Weaknesses: The Morshed et al. (40) paper provides some useful information about the early phase of metabolism of propylene glycol in rabbits; its usefulness for propylene glycol kinetics is limited because of poor sampling intervals. Blood levels of propylene glycol dropped from a maximum of 41.0 mM at 1 hour after dosing to 36.6 mM at 3 hours after dosing. A very rough estimate under the assumption of the first order one compartment model would indicate a half-life of about 12 hours in the rabbit. It must be emphasized that neither assumption may be correct, because the high dose and the very slow flux of L-lactate indicates that the system operated according to a zero order process. [In any event, neither Morshed et al. (40, 55) paper is properly interpreted.] The study in rats (55) did not determine blood levels of propylene glycol although it used many doses and a sufficient number of time points. Lactate levels are plotted on an arithmetic scale, which allows half-life estimates by a visual inspection but no exact calculation. The statement "The elimination time ranged from 1.40 to 5.82 hour which followed apparent first order kinetics" is contradictory. The half-life of first order processes is a constant and independent of dose. Except for the two lower doses (0.4 and 0.8 mg/kg), which were below saturation of metabolic clearance, the higher doses (1.6, 3.2, and 6.0 ml/kg) were above saturation of metabolic clearance and therefore the metabolite (lactate) reflected the kinetics of the parent compound (saturation of alcohol dehydrogenase being the rate-determining step) with dose-dependent increase in its half-life.

The time course evaluated for propylene glycol-induced lactatemia in rabbits was too short to allow for any conclusions regarding D- or L-lactate half-life in the study of Morshed et al. (40). That study also contains contradictory data in that blood L-lactate concentrations peaked at the earliest time point (0.25 hours) and declined thereafter (see Table 2-1 above). However, the propylene glycol concentration peaked at 1 hour and fell only slightly by 3 hours. This irregular decline of primary metabolite in the face of increasing parent compound concentrations is not readily interpretable. One might conclude from this paper that L-lactate is orders of magnitude more important as a metabolite of propylene glycol than is D-lactate. However, it should be made

clear that this may only be true for the rabbit, as Morshed et al. point out that rat ADH is more efficient in metabolizing D-propylene glycol than is rabbit ADH, which leads to slightly greater overall lactate levels from propylene glycol metabolism in rats than in rabbits. The lack of information of D- vs. L-lactate formation in humans makes it unclear whether humans are more like the rat or rabbit.

Utility (Adequacy) for CERHR Evaluation Process: The usefulness of the Morshed et al. (40, 55) data is limited for reproductive and developmental considerations. It is clear from these papers that high doses of propylene glycol will result in sustained hyperlactemia, probably without lactic acidosis, because of the efficient removal of lactate via gluconeogenesis.

In a study examining clinical chemistry abnormalities, 5 or 6 cats of each sex were fed a diet containing 12% propylene glycol (low dose, 1.60 g/kg bw/day) for 5 weeks (a dose equivalent to that found in commercial soft-moist cat foods), or a high dose diet containing 41% propylene glycol (8.00 g/kg bw/day) for 22 days (39). Propylene glycol (99.7% purity) was a racemic mixture of D- and L-isomers. Predosing observations were made such that each group of cats served as its own control. Clinical chemistry analyses were conducted on serum samples. L- (+) lactate was determined enzymatically using L-lactate dehydrogenase and D-(-) lactate was determined on days 0, 10, and 24 of the low-dose diet and days 0, 6, 10, and 24 of the high-dose diet. Data were analyzed by ANOVA and significance was at the p<0.05 level. Plasma levels of propylene glycol were measured in two of the low-dose cats. Propylene glycol levels on day 24 of dosing were 19.1 and 8.4 mM and propylene glycol was not detected in the control plasma. The authors reported a linear correlation between increases in anion gap [anion gap = (Na⁺) -(Cl^- + total CO₂)] and D-lactate in cats fed the low dose. Serum levels of D-lactate increased with days of propylene glycol ingestion and levels of L-lactate decreased in low-dose cats (Table 2-2). The authors noted previous observations where propylene glycol was found to produce L-lactic acidosis in humans and animals including cats shortly after exposure. Because their study first measured lactic acid exposure at 1 week following exposure, it is unknown if acute increases in L-lactate concentration occurred in the cats.

			0 days ingestion  10 days ingestion 	24 days ingestion
D-lactate (1.6 g/kg) 	0.08±0.03 mmol/L  1.90±0.80 mmol/L 	1.96±0.75 mmol/L
L-lactate (1.6 g/kg) 	1.02±0.18 mmol/L 			0.60 (approx)*
D-lactate (8.0 g/kg) 			  4.21±1.95 mmol/L 	7.12±0.14 mmol/L

* Value taken from graph; 0.32 ± 0.10 mmol/L lactate at 35 days ingestion.
** Christopher (39).

Strength/Weaknesses: The Christopher et al. (39) paper is important because it links the anion gap with D-lactate levels in plasma in cats after repeated doses of propylene glycol. Plasma levels of propylene glycol were determined in two low dose (1.6 g/kg bw/day) cats, which in itself is not suitable for any kind of kinetic modeling. Nevertheless these data (19.1 and 8.4 mmol/L) are in agreement with the Morshed et al. (41) results, which showed that administration of a single dose (1.6 g/kg) of propylene glycol resulted in peak plasma concentration in the same concentration range (about 8 mmol/L). Thus, it appears that the half-life of propylene glycol is short in cats as well since there seems to be no accumulation of it after repeated administration.

Utility (Adequacy) for CERHR Evaluation Process: Christopher et al. (39) is a useful study linking human data (54) with animal data regarding D-lactatemia.

It appears that high, acute doses of propylene glycol can lead to lactic acidosis. Unless the dose is very high, L-lactate is efficiently converted (detoxified) to glucose. However, D-lactate is not readily converted in the gluconeogenic pathway and therefore tends to accumulate after subacute/chronic dosing leading to D-lactic acidosis. Logically, lactate dehydrogenase must have a much higher affinity for L-lactate than for D-lactate because chirality is lost at the level of pyruvate and D- and L-lactate derived intermediates become indistinguishable upstream of pyruvate.

It may be more likely that at high propylene glycol doses and plasma lactate loads, lactate clearance via utilization in intermediary metabolism is saturated. Limited evidence for this is suggested in the D, L-lactate dosing study of Oh et al. (54). Ten male volunteers received one of two different infusion rates (n=5 per group) of D, L-lactate in which a doubling in the D-lactate blood level yielded only a 1.5-fold increase in D-lactate utilization rate but a 3.5-fold increase in D-lactate urinary excretion. The levels of D-lactate in this study were in the same range as those reported for total lactate at the high doses in rats (55). The rate of L-lactate excretion and utilization were not reported in the human study (54).

2.1.3.3 Developmental and Species Specific Variations in Metabolism and Enzyme Activities

Activities of enzymes such as ADH and ALDH can affect how fast propylene glycol is cleared from the body, thus affecting potential toxicity. A number of studies examined both the activities of these enzymes in human placenta and the age-related activity of the enzymes. Although most studies focused on ethanol metabolism, they are still relevant to propylene glycol metabolism, since ADH and ALDH activities are investigated. Therefore, CERHR conducted a brief review of the data.

Studies in humans and rodents suggest that the placenta has extremely limited capacity to metabolize propylene glycol. Pares et al. (56) isolated Class III ADH from full term human placenta and found it had low activity for ethanol and a K_m value for octanol that was 100-times higher than the Class I ADH enzyme found in human liver. Zorzano and Herrera (57) found that ALDH from full-term human placentas had a lower activity and Vmax, and a higher K_m value than ALDH isoenzymes from liver.

In rats, placenta was found to have no ADH activity and ALDH activity in placenta was found to be 4-7% of liver activity (58).

Sjoblom et al. (58) found that in Wistar rats ADH activity in liver was low before birth, being 5 and 16 % of adult activity on gd 15 and 20, respectively. There was a rapid increase at birth: 53% of adult levels on postnatal (pnd) 1 with a continued gradual increase with age to 82% of adult activity on pnd 47. Similar developmental patterns were noted for ALDH in rat liver.

Pikkarainen and Raiha (32) measured in vitro ADH activity in the livers of human fetuses, children, and adults (n=1–3/age group) using ethanol as a substrate. The ADH activity in 2-month-old fetal livers was about 3–4% that of adults. In 4–5-month-old fetuses, ADH activity was roughly 10% that of adults, and in infancy, activity was about 20% that of adults. ADH activity increased in children with age, and at 5 years of age, activity reached a level within ranges noted for adults. Great variation was noted in adult ADH activity.

Somewhat different results were reported subsequently by Smith et al. (59) who examined human liver ADH activity using ethanol as a substrate and also examined the ontogeny of individual ADH class I isoforms. They reported total ADH activity in 9-22-week-old fetal liver that was 30% of adult values, and in premature infants and children less than 1 year old, activity was 50% of adult values. Individual enzyme activity was determined using starch gel electrophoresis with an in situ assay. A total of 222 liver samples were assayed, 56 from fetuses (9-22 weeks gestation), 37 from premature infants and infants less than 1 year of age, and 129 from adults greater than 20 years of age. In fetal liver samples with a mean gestational age of 11 weeks, only the ADH1A enzyme was detectable. By 17 weeks, both ADH1A and ADH1B were measurable, although ADH1A predominated. By 19 weeks, products from all three loci were observed, with ADH1A greater than ADH1B, and ADH1B greater than ADH1C. At 30 weeks, ADH1A and ADH1B levels were equivalent, but still greater than ADH1C, but by 36 weeks, ADH1B expression dominated. In the adult, hepatic ADH1A expression was nondetectable, whereas expression from the ADH1B and ADH1C loci were equivalent. Interestingly, this progressive change in expression was tissue-specific. In lung, there were no observed differences between the fetal and adult samples and only ADH1C was detectable. ADH expression in the intestine and kidney was low and did not change appreciably with age.

Thus, it would appear that human liver ADH is expressed early in development and may well contribute to propylene glycol metabolic disposition. However, given the paucity of knowledge regarding isoform specificity towards propylene glycol, it is uncertain how these data on ethanol metabolism might be extrapolated. Assuming the enzyme most active in ethanol metabolism, ADH1B, is also most active in propylene glycol metabolism, the significant fetal metabolism is not predicted to occur until later in gestational development (20-36 weeks).

Strength/Weaknesses: There are consistent data in both animals and humans showing that alcohol dehydrogenase is much lower prenatally. In humans, adult levels were reached by the age of 5 years and in rats on day 47 after parturition.

Utility (Adequacy) for CERHR Evaluation Process: D, L-lactate, metabolites of D, L-

propylene glycols, are postulated to be associated with toxicity in mammalian species. Therefore a lack of in situ conversion in the fetus would seem to decrease the toxicity of propylene glycol. Since lactate distributes into total body water, the fetus will share the mother’s metabolic load and associated acidosis, if present. The lower metabolism capability in newborns and infants, however, may partially protect them from metabolic acidosis after ingestion of propylene glycol.

Zorzano and Herrera (60, 61) found different ADH isoenzymes in liver homogenates from humans (class I ADH) and rats (ADH-3), which differed greatly in kinetic properties. Using ethanol as a substrate at a pH of 10.5, activity, K_m, and _Vmaxin humans was measured at 6.24 Units/g tissue, 2.10 mM, and 7.70 Units/g tissue, respectively, while activity, K_m, and Vmax in rats was measured at 2.72 Units/g tissue, 1.02 mM, and 2.96 Units/g tissue, respectively. Two different low K_m ALDH isoenzymes were found in humans and rats but they had similar activities using acetaldehyde as the substrate at pH 8.8 (humans: Km=9 µM and _Vmax=0.85Units/g tissue; rats: Km=10 µM and V_max=0.87 Units/g tissue).

Reviews by Agarwal (62), Bosron and Li (63), Pietruszko (64), and Burnell et al. (65) discussed genetic polymorphisms for ADH and ALDH in humans. Class I ADH, the primary ADH in human liver, is a dimer composed of randomly associated polypeptide units encoded by three loci (ADH1A, ADH1B, and ADH1C). Polymorphisms resulting in altered phenotypes are observed at the ADH1B and ADH1C loci.

There are two primary ALDH isoenzymes in human liver, ALDH2 (also referred to as E₂,ALDHI, or ALDH2 ) and ALDH1 (also referred to as an E1, ALDHII, or ALDH1) (62-64). About 50% of Japanese and Chinese carry a phenotypically null variant of the ALDH2 enzyme.

In mammals, part of the propylene glycol dose is eliminated unchanged by the kidney and part is metabolized by the liver to lactic acid and further metabolized to pyruvic acid; in mammals, with the exception of cats, the remainder is conjugated with glucuronic acid (2) and eliminated in the urine. The amount of propylene glycol eliminated by the kidneys has been estimated for humans at 45% (48), for dogs at 55–88% (49), and for rabbits at 2.4–14.2% (50). Morshed et al. (41) provided evidence in the rat that increasing doses of propylene glycol increased elimination by the kidneys. Dosages of 19, 38, and 77 mmole/kg bw resulted in 2.3, 7, and 17% renal excretion of propylene glycol. Maximum urinary excretion of propylene glycol was determined using pyrazole (1.0 mmole/kg bw), a competitive inhibitor of propylene glycol. High urinary clearance was observed with 75% excretion of the ingested dose within 24 hours.

In human adults receiving 20.7 or 41.4 g propylene glycol 2–3 times daily for a minimum of 3 days, the total body clearance was dependent on serum concentration and was approximately 0.1 L/kg bw/hour; elimination half-life in those same subjects was about 4 hours (29). [The influence of ethyl alcohol administration must be considered when interpreting results since it will compete with propylene glycol for the dehydrogenase enzymes.] In a study where adults and children were rectally exposed once to ¨ 123–173 mg/kg bw propylene glycol [blood levels 1.6– 2.2 mM], the clearance rate was 0.2 L/hour/kg and half-life was 2.6–2.8 hours (31). In 6 adults

receiving propylene glycol intravenously, blood levels of propylene glycol were measured at 48– 425 µg/mL [0.63–5.6 mM] and an average half-life of 2.3 hours was estimated (30).

A small number of studies suggest that elimination of propylene glycol in infants is slower than in adults. In an 8-month-old infant exposed to propylene glycol through medication applied to burns, the propylene glycol blood level was 1.059 g/dL [139 mM] and the elimination half-life was measured at 16.9 hours (35). Ten infants exposed to 10 mL [10.36 g] propylene glycol in a parenteral vitamin solution daily for 5 days had propylene glycol blood levels of ¨ 65-950 mg/dL [8.5–125mM] and elimination half-lives of 10.8–30.5 hours, with a mean of 19.3 hours (36).

Excretion of propylene glycol has been studied in patients with second and third degree burns over more than 20% of their total body surface (34). According to ATSDR (4), "Sulfadiazine preparations containing propylene glycol were applied dermally over a period of 3–7 days after admission to the hospital. Serum and urinary levels of propylene glycol were measured. Propylene glycol was detected in the serum of 24 of 45 patients and in the urine of 40 of 45 patients. Average urinary levels were 1.3 mg/mL with a range of 0–17.9 mg/mL for patients who lived, and 2.9 mg/mL with a range of 0–23 mg/mL for patients who died. Propylene glycol levels correlated with total burn surface area and total third degree burn surface area."

Strength/Weaknesses: Elimination kinetics of propylene glycol are well understood. Speth et al. provides the major kinetic parameters needed for calculations. The saturation of metabolic clearance occurs in humans at about 7 g, which is somewhat lower than in animals. Kollöffel et al. (31) provide data in 10 adults which indicate that at a dose of 8.64 g, elimination of propylene glycol was zero order because it was nearly linear on an arithmetic scale. At a dose of 5.1 g/day the half-life of propylene glycol was 1.6±0.20 hours, at doses of 7.2 to 7.7 g/day it was 1.9±0.15 hours, and at doses of 12.6 to 21.0 g/day it was 3.2±0.12 hours (30). The data of Kollöffel et al. provide 2.6±0.2 hours as half-life in adults at a dose of 8.64 g/day. At a dose of 3 x 20.7 to 2 x 41.4 g/day, Yu et al. (29) estimated an elimination half-life of about 4 hours. Thus, the half-life of propylene glycol increased from 1.6 to 4 hours as the dose increased from 5.1 to 2 x 41.4 g/day. The half-life of chemicals eliminated by first order processes is independent of dose. Therefore, it is certain that in humans propylene glycol is eliminated by zero order kinetics at or above a dose of 5.1 g/day. Clearance data and AUCs verify this conclusion.

The infant studies suggest prolonged half-lives of propylene glycol (35, 36) in the range of 10.8-30.5 hours in infants receiving a dose of about 3 g propylene glycol. While such data are consistent with very low alcohol dehydrogenase activity perinatally (32), they cannot be considered definitive due to confounding effect(s) associated with the disease treatment and the drugs associated with such therapy. In addition, the Kulick et al. (34) paper is not suitable for determination of elimination kinetics because only one time point was measured. Prolonged half-life of propylene glycol in infants is also supported by a recent report showing that it accumulated to very high levels (up to 2,000 µg/mL) in serum of these children (66).

Utility (Adequacy) for CERHR Evaluation Process: There are sufficient data available on the elimination kinetics of propylene glycol in humans to model elimination in adults; data in infants and in the fetus are less certain.

"Dose-dependent elimination of propylene glycol is seen in rats, with saturation of the pathways at doses above 5.88 g/kg. An apparent maximum elimination rate of 8.3 mmol/kg/hour (0.63 g/kg/hour) was observed" (4).

Yu and Sawchuk (50) studied the metabolism and elimination of propylene glycol after acute or chronic IV administration to NZW male rabbits. Rabbits were exposed acutely by IV injection to either 0.5,1.0, or 2.0 g/kg bw (three rabbits per dose group). There was evidence of a saturation of propylene glycol metabolism at the 2.0 g/kg bw acute dose, as evidenced by the decreased metabolic clearance. The half-life and the terminal elimination phase rate constant was not significantly affected over this dose range. An additional few rabbits were exposed by continuous IV infusion to propylene glycol delivered at various rates (2.8-6.3 mg/min/kg bw) over the course of 51-52 hours. Both Vmax and K_m were lower in the case of prolonged exposure, but the Vmax/Km ratio was approximately 3-fold greater than under acute dosing. Plots of metabolic clearance from single rabbits dosed acutely versus continuously indicate higher metabolic clearance rates from continuous exposure. [This raises the possibility of the induction of a second, low K_mform of ADH during the 51–52 hours of infusion.] The authors concluded that metabolism of propylene glycol was the dominant disposition pathway with a concentration-dependent metabolic clearance; renal excretion of propylene glycol was only 2.4–14.2% of the total dose after acute administration, most likely due to kidney reabsorption. Authors also concluded that for both acute and chronic administration of propylene glycol, the clearance of propylene glycol is lower at higher plasma concentrations and the rate of elimination of propylene glycol was dependent upon urine flow.

Ruddick (49) cited an earlier study by Lehman and Newman (42) where dogs were force fed 8 mL/kg and 12 mL/kg of a 50% aqueous solution of propylene glycol. Blood concentrations were 1.3 g/dL [171 mM] 2 hours after dosing and 0.9 g/dL [118 mM] 4 hours after dosing. Recovery of 12–45% of the unchanged administered dose in the urine led the authors to conclude that the compound was eliminated by the kidney and a large portion of unexcreted chemical was metabolized.

Strength/Weaknesses: Animal data are consistent with human data regarding the elimination kinetics (practically the same elimination half-life before saturation of metabolic clearance) of propylene glycol, although minor species differences may be present. Saturation of metabolic clearance occurs at somewhat higher doses in animals; therefore, the half-life of elimination

Utility (Adequacy) for CERHR Evaluation Process: It is useful to have mechanistic insight into the process of elimination of propylene glycol as represented by the Yu and Sawchuk (50) paper on the urinary flow dependence of elimination as well as on the dose-dependence of metabolic clearance.

The Ruddick (49) and Lehman and Newman (42) papers are not suitable for quantitative kinetic evaluation.

The majority of information in this section is summarized from the reviews by ATSDR (4) and LaKind et al. (28) and from the SIDS Initial Assessment Report for 11^th SIAM (27) and the EPA Health and Environmental Effects Document on Propylene Glycol (67). No toxicity studies have

been located on propylene glycol subsequent to the 2001 SIDS Initial Assessment Report. A very limited number of toxicity studies included an examination of the reproductive organs and those studies are discussed in detail.

Propylene glycol has very low systemic toxicity in experimental animals and very high doses are used in most acute studies to determine a toxic level. It is primarily metabolized to lactic acid and pyruvic acid, both of which are normal constituents of the citric acid cycle. CNS, hematologic, hyperosmotic, and cardiovascular effects have been noted in humans and animals and high serum concentrations of propylene glycol may result in lactic acidosis and hyperosmotic changes in the blood (4, 27, 49). Symptoms of acute propylene glycol intoxication in animals include CNS depression and narcosis. Individuals with compromised hepatic or renal function would be less apt to clear propylene glycol, and hence would be more susceptible to toxicity due to high blood levels (2, 4, 68). No system or organ has been established as a target for the acute oral lethal effects of propylene glycol (69).

Lactate can be detoxified into glucose and stored as glycogen as has been demonstrated by Wittman et al. (47) in propylene glycol-exposed rats. Doses of 0.5-2.0 g/kg of propylene glycol were administered to female rats and liver glycogen content and blood glucose were determined 90 minutes after dosing. Liver glycogen content nearly doubled and fasting blood glucose increased from 88 to about 140 mg%. Lactic acidosis was not reported. [Lactic acidosis is not expected at these relatively low doses of propylene glycol. However, lactic acidosis can develop if these two detoxification pathways cannot remove excess lactic acid sufficiently.]

ATSDR (4) stated that "The mechanism of action of propylene glycol is not well understood". [In fact, much is known about the mechanism of action.] Lactatemia has been well documented in animals and there are supporting human data. Cats administered 12 (1.6 g/kg bw/day) or 41% (8.0 g/kg bw/day) propylene glycol in the diet (dry weight) for 22 days, showed a time-dependent increase in plasma lactate and in anion gap (39). Morshed et al. (40, 43) produced more data on the dose-dependence of blood lactate and/or pyruvate in rats and rabbits given propylene glycol orally. Finally, a human case report (48) demonstrated that repeated infusions of lorazepam dissolved in propylene glycol can lead to lactic acidosis with increased osmolar gap (21mOsm/L). Furthermore, increased blood glucose (296 mg/dL) and elevated pyruvate level (1.01 mg/dL) indicate that the same metabolic pathways of detoxification occur in humans as in animals. Glasgow et al. (36) reported a good correlation between osmolality gap and serum propylene glycol concentrations in ten infants. The half-life was reported as 19.3 hours (range 10.8-30.5 hours), which is about 10 times longer than in adults. Alcohol dehydrogenase activity is up to 10 times lower in infants (32) than in adults providing an explanation for the prolonged half-life in the latter and at the same time further evidence that this enzyme is the rate-determining enzyme in the clearance of propylene glycol. Other endpoints of toxicity are anesthesia, probably by the same mechanism as other alcohols, and hemolysis, which may be due to the osmolality gap.

Strength/Weaknesses: There is an adequate database to assess the toxicity of propylene glycol (4, 27, 28, 67). Very high doses of propylene glycol cause CNS, hematologic/hyperosmotic, and perhaps cardiovascular effects, as well as lactic acidosis. Animals lethally intoxicated undergo CNS depression, narcosis, and eventual respiratory arrest.

Utility (Adequacy) for CERHR Evaluation Process: There are sufficient reliable reviews to obtain any information needed for informed toxicological judgment.

A lethal oral dose of propylene glycol has not been reported for humans (28), but it is estimated that the human lethal oral dose is >15 g/kg or >32 fl oz for a 150 lb person (2). In adults, serum levels of >180 mg/L [2.37mM] have resulted in toxicity (48). In one case, an 11-year-old child receiving oral doses of 2-4 mL per day for 13 months as a component of a vitamin D preparation (estimated dose 4–8 g/kg bw/day) resulted in seizures and CNS depression (28). In acutely ill infants, death has occurred after repeated exposure to propylene glycol in medication; CNS depression and seizures have been reported after multiple oral doses (36, 70) [see Section 2.5 Potentially Sensitive Subpopulations]. According to HSDB (2), the acceptable daily intake of propylene glycol as a food additive is 25 mg/kg body weight.

Contact dermatitis has been reported from propylene glycol exposure in a wide variety of topical preparations (28) and ingestion of propylene glycol in sensitized individuals has produced flares of dermatitis (28). Skin irritation resulting from topical exposure is manifest as erythematous reactions restricted to sites of exposure. The irritation potential is enhanced after prolonged dermal exposure, under dermal occlusion, and in combination with triethanolamine-stearate, a cosmetic emulsifier (71, 72). The nature of the skin reaction of propylene glycol-sensitive patients has been a matter of controversy (73, 74). In a study by Hannuksela and Forstrom (73), primary irritant reactions to the skin and type IV delayed hypersensitivity reactions were observed following oral ingestion or topical application of propylene glycol. However, in most cases, the skin reaction was due to a primary irritation, not to an allergic reaction (72).

Propylene glycol is a component of theatrical fog and is used for special effects. The Actors’ Equity Association and the League of American Theaters and Producers sponsored a study which included an examination of the health effects of theatrical fog in response to actors’ concerns about exposure (75). The health endpoints selected for investigation were irritant effects to the respiratory tract and eyes. This study was conducted over 2 years with 439 actors from 16 musicals, and consisted of a baseline questionnaire, daily checklists, and medical evaluation. There was no clinically significant adverse impact on pulmonary function or in rates of asthma associated with exposure to propylene glycol. However, "peak exposures to elevated localized air concentrations following release of glycol smoke are associated with increased reporting of respiratory, throat, and nasal symptoms, and findings of vocal cord inflammation." The study authors recommended that exposures to propylene glycol by actors not exceed peak or ceiling concentrations of 40 mg/m³.

NIOSH conducted a study in 1990 on the use of theatrical fog in Broadway theaters (76). Personal breathing zone and general area air sampling and a questionnaire on irritant effects (130 questionnaires from productions with theatrical smoke, 90 questionnaires from productions without theatrical smoke) were collected from personnel from four productions using theatrical smoke and five productions without theatrical smoke. Air samples collected yielded propylene glycol concentrations <2.1 mg/m³. However, there was a significant (p<0.05) increase in the reporting of respiratory irritant symptoms such as runny nose, stuffy nose, and sneezing by personnel from productions using theatrical smoke.

In a study by Cohen and Crandall (77) [reviewed by LaKind et al. (28)], propylene glycol was recommended as a vehicle for administration of bronchodilator drugs. No adverse clinical effects were observed after subjects were exposed to an inhalant mist of isoproterenol-HCl containing 40% propylene glycol for 15 minutes at a temperature of 115–124° F.

Wieslander, Norbäck, and Lindgren (23) examined experimental exposure of volunteers to propylene glycol mist simulating concentrations routinely used in aviation emergency training. Twenty-seven non-asthmatic volunteers (22 males, 5 females) were exposed in an aircraft simulator to propylene glycol mist over a 1-minute period (average concentration 360 mg/m³; range 176–851 mg/m³). Average age was 44±11 years. None of the subjects had previous occupational exposure to propylene glycol. Medical examinations were performed both within 15 minutes before and after the exposure. Exams included an estimate of tear film stability breakup time, nasal patency by acoustic rhinometry, lung function by dynamic spirometry, and a self-rated symptom questionnaire. After 1 minute of exposure there was a statistically significant difference when compared to pre-exposure levels in tear film stability (decreased; P=0.02) and ocular and throat irritation ratings (both increased; P<0.001) [P values determined by Student’s t test for paired comparisons]. The forced expiratory volume in 1 second over the forced vital capacity was slightly reduced and the self-rating of severity of dyspnea increased. There were no apparent changes in nasal patency, vital capacity, forced vital capacity, nasal symptoms, dermal symptoms, smell of solvents, or any other systemic symptoms. The authors concluded that short exposure to propylene glycol mist from artificial smoke generators may cause acute ocular and upper airway irritation.

Hemolysis, CNS depression, hyperosmolality, and lactic acidosis have been reported after IV administration of propylene glycol (68). Rapid IV infusion of concentrated propylene glycol-containing drugs has been associated with respiratory depression, arrhythmias, hypotension, and seizures. Propylene glycol is used as a vehicle for IV administration of drugs such as lorazepam, etomidate, phenytoin, diazepam, digoxin, hydralazine, esmolol, chlordiazepoxide, multivitamins, nitroglycerin, pentobarbital sodium, phenobarbital sodium, and trimethoprim-sulfamethoxazole. Therefore, patients, especially children and infants, receiving IV drugs can be at risk for propylene glycol toxicity (28) [see Section 2.5 Potentially Sensitive Subpopulations].

Information on the dose of propylene glycol necessary to induce toxicity is limited. Some reports describing the dose of propylene glycol given and the serum concentration measured in cases of toxicity in humans are contained in Table 2-8 in Section 2.5, Potentially Sensitive Subpopulations.

General toxicity studies in animals are discussed in the sections below and summarized in Table 2-4.

LD₅₀ oral toxicity values are listed in Table 2-3. A wide range of LD₅₀ values has been reported for the rat. In a study by Morshed et al. (43), 6 male Wistar rats were dosed by gavage with saline or 2.942 g/kg bw/day propylene glycol in water for 10, 20, or 30 days. No deaths occurred over any of the time intervals. However, a 41% reduction in body weight was noted at 10 days and an increase in body weight was noted at 20 and 30 days in treated animals as compared to respective saline controls.

Strength/Weaknesses: This study by Morshed et al. (43) does not have strengths, only weaknesses. Controls gained 16.9 g during the first 10 days (1.69 g/day on average), 23.3 g after 20 days (1.17 g/day on average), and 40.15 g after 30 days (1.34 g/day on average). Well-maintained rats do not display such weight gain variability.

In a study by Weatherby and Haag (78) [reviewed by OECD (27)] in rats, only minimal kidney changes were observed and the LD₅₀ value was determined to be 33.5 g/kg.

Strength/Weaknesses: This is an older study (78) which characterized acute toxicity of propylene glycol in rats and rabbits by various routes of administration. As expected, propylene glycol was most toxic when administered IV. Toxicity decreased IV>IM>subcutaneous>oral. There was no apparent species difference. Information provided on the chronic administration of propylene glycol is sparse but the hemolysis experiment with human blood in vitro demonstrates conclusively the hemolytic potential above 0.111 M.

Utility (Adequacy) for CERHR Evaluation Process: This study by Weatherby and Haag (78) is useful for the characterization of acute toxicity, but is less useful for chronic toxicity.

Acute oral toxicity in rabbits was studied by administering a 20% aqueous solution of propylene glycol by stomach tube over a 1-hour period (15.75-21.00 g/kg) (79) [reviewed in LaKind et al. (28); OECD (27)]. Animals exhibited an increased respiratory rate, loss of equilibrium, depression, analgesia, coma, and died by 36 hours post dosing. The minimum fatal dose was determined to be 18.9 g/kg (3 of 9 deaths), with 100% mortality at a dose of 21 g/kg (4 of 4 deaths).

Strength/Weaknesses: The Braun and Cartland (79) paper predates the Weatherby and Haag (78) publication and represents a less extensive but nevertheless reliable documentation of the acute toxicity of propylene glycol administered IM and subcutaneously to rats and orally to rabbits. Results of the two studies are very similar. Data on chronic toxicity are scant.

Utility (Adequacy) for CERHR Evaluation Process: This report is useful for the characterization of acute, but not chronic, toxicity.

Chronic toxicity studies reflect that propylene glycol has a very low order of toxicity. In the following toxicity studies by Morris et al. (80) and Gaunt et al. (81), reproductive tissues were examined.

Albino rats (inbred strain, male and female, 20 rats/group) were administered 0, 2.45, and 4.9% of propylene glycol in the diet (0, 1.23, and 2.45 g/kg bw/day, respectively) for 2 years. Other glycol chemicals were also part of this chronic study. Body weights and food consumption were determined at weekly intervals. No changes were noted when compared to control animals for growth rate, food and water consumption, and animal survival. There were no differences between control and propylene glycol groups in gross and microscopic lesions in the lung, heart, liver, spleen, kidney, adrenal glands, and testes [individual data or summary tables not reported]. The authors noted that there were no bladder stones or signs of chronic kidney damage and no change in the gross morphology of the testes when compared to control animals. "Slight liver damage" [authors’ words] was observed in the propylene glycol exposed group (80) [reviewed in LaKind et al. (28); OECD (27)]. [No statistical analyses were performed and the histopathology of the liver is not described.]

Strength/Weaknesses: The Morris et al. (80) paper predates standardized chronic toxicity test protocols and some may view it as poorly controlled. However, the experiment is well-described including the limitations. Therefore, it appears reasonable to accept that daily doses of 4.9% propylene glycol in the diet (¨ 3 g/kg) caused centrilobular atrophy, bile duct proliferation, and fatty degeneration in the liver even though it is not stated in the paper at which dose slight liver damage was observed. The highest doses (1.7 to 2.1g/kg) used by Gaunt et al. (81) were close to the lower dose in this study and no liver effect was reported by Gaunt et al. Therefore, the lower dose probably did not cause liver damage. Failure to conduct statistical analyses weakens this study.

Utility (Adequacy) for CERHR Evaluation Process: The Morris et al. (80) study can only serve as a modest indicator that 3 g/kg propylene glycol chronically might cause slight liver injury.

In 2-year and 15-week toxicity studies in rats given propylene glycol in the diet (81), body weight, renal concentration tests, organ weights, histology, and incidence of neoplasms were described. Necropsy at the end of the study included gross and microscopic examination of the male and female reproductive tracts. Charles River CD rats from a Specific Pathogen Free (SPF) breeding colony were used in this study. At the start of the study, the weight range of the males was 120–150 g and of the females was 120–140 g. [Statistical methods were not described and standard errors for treatment groups were not presented.] The studies were run concurrently.

For the short-term study, groups of 15 male and 15 female rats were fed diets containing 0, or 50,000 ppm propylene glycol [Shell Co. Ltd., >99% purity] for 15 weeks. Body weights and food

consumption were not recorded. During the last week of treatment, renal concentration tests were estimated over a 6-hour water deprivation period. At necropsy, blood was collected for hematology and blood concentrations of urea, glutamic-oxalacetic, and glutamic-pyruvic transaminases were determined. At necropsy, brain, heart, liver, spleen, kidneys, adrenals, gonads, and pituitary were weighed. In the short-term study, the authors reported no differences between the control rats and those fed the 50,000 ppm diet for the parameters measured, including the urine and serum analyses, blood chemistry, and organ weights [data not reported].

In the long-term study, groups of 30 male and 30 female rats were fed diets containing either 0, 6,250, 12,500, 25,000, or 50,000 ppm propylene glycol for 2 years. Animals and food consumption were monitored daily and body weights recorded at 2 week intervals. Blood was collected from the tail vein of 8 male and 8 female rats in the 0, 25,000, and 50,000 ppm dose groups at 13, 21, 52, and 80 weeks of the study; and in the 0, 6,250, and 12,500 ppm groups at week 54 of the study. A urinary concentration test was done on selected rats from the 0, 25,000, and 50,000 ppm dose groups. Measurements were made of both specific gravity and urine volume over a 6-hour water deprivation period, during a 2-hour period after a 25 mL/kg water load, and then during a 4-hour period beginning 16 hours after the water load. At necropsy, brain, heart, liver, spleen, kidneys, adrenals, gonads, stomach, small intestine, and cecum were weighed. Samples of these organs, the following organs, and any tissue that appeared abnormal were preserved in 10% buffered formalin: salivary gland, trachea, aorta, thymus, lymph nodes, pituitary, urinary bladder, colon, rectum, pancreas, uterus, and muscle.

In the 2-year study, the mean daily intakes of propylene glycol were approximately 0, 0.2, 0.4, 0.9, and 1.7 g/kg in males and 0, 0.3, 0.5, 1.0, and 2.1 g/kg in females for the 0, 6,250, 12,500, 25,000, or 50,000 ppm propylene glycol dose groups, respectively. [The authors did not provide daily food consumption or bi-monthly animal weight data.] No abnormalities were observed among groups in deaths, behavior, or food consumption. The authors reported no significant differences between the control and treated groups with respect to blood chemistry or renal concentration tests. Organ weights (including gonads) and organ weights relative to terminal body weight were similar between control and treated groups. Incidences of histological findings and the incidence of neoplasms in various tissues were presented, but the tabulated data did not include reproductive organs. Abnormalities cited were similar for the control and treated groups. The authors noted that the changes observed were consistent with those of aging rats and concluded that a "no-untoward-effect level" found in this study was 2.1 g/kg for male rats and 1.7 g/kg for female rats [highest dose used].

Strength/Weaknesses: Gaunt et al. (81) is a well-conducted carcinogenicity bioassay which clearly demonstrates that an average daily dose of 1.7 g/kg in male rats and an average daily dose of 2.1g/kg in female rats had no adverse effect (NOAEL) on body weight gain, mortality, hematology, urinary cell excretion, renal function, serum chemistry, or absolute and relative organ weights. The histopathological changes were consistent with those expected in aging rats. No malignancy could be attributed to treatment. Although reference is made in the text to "no statistically significant differences," it is not stated what statistical methods were used. However, the reputation of the British Industrial Biological Research Association (BIBRA) and of the authors of this paper lend credibility to the statement. It is unfortunate that a higher dose was not used, because as conducted, the Panel did not learn anything about the chronic toxicity in rats, only about propylene glycol’s safety. Up to 78 weeks there is no discernible effect on body weight but thereafter, there might have been a slight body weight effect. Unfortunately, no standard error is given and mortality was high in all groups, which was at least partially due to a high rate of pulmonary infection.

Utility (Adequacy) for CERHR Evaluation Process: This study by Gaunt et al. (81) establishes a highly credible NOAEL for propylene glycol in terms of chronic toxicity in both male and female rats. This information could be very useful when evaluating reproductive/developmental toxicity (i.e., a maternal NOAEL).

Propylene glycol administered in the drinking water of rats at doses >13.2 g/kg bw/day for 140 days resulted in CNS depression and minor liver injury (reviewed by Mortensen (72) and LaKind et al. (28)). In a 2-year drinking water study in rats (dosed up to1.834 g/kg bw/day), no renal pathology and very slight liver damage was found (28).

The Seidenfeld and Hanzlik (82) paper predates all other publications thus far evaluated. It includes detailed observation of the animals. A mix of acute and subchronic studies was conducted in rats and rabbits. Acute studies provided the dose ranges for the later, more detailed experiments of Braun and Cartland (79) and Weatherby and Haag (78). [The Panel notes that even though the style of the Seidenfeld and Hanzlik publication may appear outdated, the data seem reliable. In fact, the dose x time product for slight vacuolization of the liver is 1,862g x day in this study and 2,160g x day in the Morris et al. (80) report. Thus, it can be concluded that slight hepatic injury could be expected in rats at a daily intake of 2 g/kg bw of propylene glycol. The study by Seidenfeld and Hanzlik is useful because now the Morris et al. (80) report can be viewed as confirmatory evidence for the slight liver damage as a high dose effect.]

Utility (Adequacy) for CERHR Evaluation Process: This study is useful because now the Morris et al. (80) report can be viewed as confirmatory evidence for the slight liver damage as a high dose effect.

Propylene glycol was fed to dogs as a carbohydrate source in the diet at a concentration of 8% (2 g/kg bw/day) and 20% (5 g/kg bw/day) for 2 years; a control group was fed an equal caloric amount of dextrose and a second control group did not receive the dextrose. No adverse effects were observed in the low-dose group. In the high-dose group, there was evidence of RBC destruction (packed cell volume and hemoglobin values were lower and reticulocytes were higher than control values). There were no differences in kidney weights compared to the control group and no other indications of toxicity (67, 83).

Strength/Weaknesses: Weil et al. (83) studied the toxicity of propylene glycol in beagle dogs fed in the diet at 2 and 5 g/kg bw/day for 2 years. A roughly isocaloric diet to the propylene glycol containing dextrose was fed to a positive control group. After appropriate statistical evaluation, the authors concluded that 5 g/kg bw/day of propylene glycol in the diet resulted in enhanced erythrocyte destruction with signs of increased erythropoiesis. Use of a positive control group was useful to identify this effect as caused by propylene glycol. The NOAEL for chronic toxicity in dogs (2 g/kg bw/day) was essentially identical to the rat NOAEL.

Utility (Adequacy) for CERHR Evaluation Process: This paper is very useful because it has a dose that was actually toxic, which allows judgement of the ratio between LOAEL and NOAEL.

No effects were found on the kidneys in studies by VanWinkle and Newman (84) in dogs. Female dogs were administered 5% propylene glycol in drinking water two times a day for up to 9 months; male dogs were allowed to drink 600 mL of 10% propylene glycol daily. Kidney function was measured by phenosulfonphthalein excretion and liver function by rose bengal in the blood and galactose and uric acid in the urine. No pathological changes were found in these organs (28).

Strength/Weaknesses: In these experiments (84), liver and kidney function of dogs provided drinking water containing 5% propylene glycol (5 .1 cm³=5.3 g/kg body weight) were determined and found not to be effected. However, dogs given water with 10% propylene glycol died and those provided with 10% propylene glycol containing water in the morning and clean water in the evening showed impaired renal function as indicated by increased blood urea. Authors stated that control values ranged from 14 to 24 mg% and after drinking the glycol for 6 months the range was 12-33 mg%. Statistical analysis was not performed and if it had been, it certainly would have shown no difference. There are no hematology measurements.

Utility (Adequacy) for CERHR Evaluation Process: The studies of Van Winkle and Newman (84) may be considered inadequate by today’s standards, but they still provide useful data as confirmatory evidence for the NOAEL of 2 g/kg bw/day established by Weil et al. (83) in dogs.

Propylene glycol was tested on the clipped skin of NZW rabbits according to three protocols (the cosmetic protocol, the Association Francaise de Normalization protocol, and the OECD protocol); in all three tests, propylene glycol was classified as a nonirritant (28).

Strength/Weaknesses: Irritation potential of propylene glycol, although minimal, has been established in man.

Utility (Adequacy) for CERHR Evaluation Process: None. 2.2.2.3 Inhalation Exposure

The ATSDR review (4) states that studies available on inhalation exposure of animals to propylene glycol are inconclusive. An acute inhalation study with 10% propylene glycol [mg/L not stated] for 20 or 120 minutes in rabbits resulted in degenerated goblet cells in the trachea (88). However, a subchronic exposure study in rats (85) did not support these findings. Rats exposed to 321 ppm over 90 days had thickened respiratory epithelium and enlarged goblet cells (85). Monkeys (n=29) and rats [number not specified] were continuously exposed to propylene glycol vapor at doses of 32–113 ppm for 13 months. At 113 ppm, hemoglobin levels were slightly increased; there were no adverse effects noted on body weight or on the renal, respiratory, gastrointestinal, hepatic, or endocrine systems (4).

Strength/Weaknesses: Konradova et al. (88) demonstrated that a 10% propylene glycol mist inhaled by rabbits resulted in enhanced mucolytic activity (+69%) of respiratory goblet cells. This is not surprising from a surface tension lowering agent. In fact, the effect of pure propylene glycol was less pronounced than that of clinically used mucolytics (Broncholysin, Histabron). Other conclusions regarding ciliated cells are difficult to assess because of the smallness of the effect. Moreover, a much more thorough study of inhalation of a propylene glycol aerosol did not confirm these findings (85).

The Suber et al. (85) paper appears to be a well-conducted subchronic, nose-only inhalation study by a contract laboratory. Nominal doses were 0.0, 0.16, 1.0, and 2.2 mg/L of propylene glycol with an air flow rate of 1.0-1.5 L/min to each animal. Absorption was not determined, but system toxicity could not be expected even if 100% of the highest dose had been absorbed. As is clear in Bau et al. (38), only a fraction of inhaled propylene glycol will be absorbed into the systemic circulation through the lungs. Nasal hemorrhage is compatible with the known irritation potential of propylene glycol. Goblet cell score was significantly increased in the nasal turbinates, which is plausible for a surface-active agent facilitating the discharge of mucous from the swollen goblet cells.

Utility (Adequacy) for CERHR Evaluation Process: This is a useful study (85) that confirms the view arrived at for kinetic reasons that exposure by inhalation to propylene glycol does not pose a significant toxicological problem.

Robertson et al. (86) examined the chronic toxicity of propylene glycol by inhalation in Rhesus monkeys and rats. This is a very interesting study because both rats and monkeys were exposed continuously to saturated/supersaturated air of propylene glycol (55–113 ppm) for up to 1 year. At the highest dose, hemoglobin levels seemed to have increased. However, since no standard

error is given and no statistical analysis was performed, it is uncertain whether this is a real effect. Otherwise, no adverse effects were found in spite of extensive gross and histopathologic examination. In fact, both rats and monkeys inhaling propylene glycol gained more weight than the controls. The health status of monkeys was poor, which was not uncommon in 1947.

Assuming Rhesus monkeys inhale about 2 m³ of air per day, the data indicate that primates may safely inhale about 1 g of propylene glycol per day. Although this paper uses unusual reporting methods by today’s conventions, it certainly appears reliable and interpretable.

Utility (Adequacy) for CERHR Evaluation Process: Continuous exposure to propylene glycol vapor (without vehicle) in a primate species provides important evidence.

Results from animal studies indicate that intermediate and chronic exposure to propylene glycol may lead to hemolysis of RBCs. After a 90-day inhalation exposure to 321 ppm of propylene glycol, female rats had decreased white blood cell count, while exposure to 707 ppm of propylene glycol decreased hemoglobin concentrations. No dose-related changes in RBCs were observed in male rats (85). After exposure of rats to 5% propylene glycol in the diet for 2 years, there were no hematological effects noted (81). However, Saini et al. (90) [reviewed by OECD (27)] found that a single oral dose of either 0.73 or 2.94 g/kg bw given to female Wistar rats, produced a reversible, statistically significant decrease in hemoglobin, packed cell volume, and RBC counts for 2 days. Electron microscopy revealed a rough RBC surface. However, in an early study by Robertson et al. (86), Rhesus monkeys continuously exposed to concentrations of propylene glycol in air up to 112 ppm for 13 months had a slightly greater increase [statistical analyses not reported] in RBCs and hemoglobin content than the control animals.

Cats exposed to oral administration of propylene glycol developed Heinz bodies in RBCs and experienced decreased RBC survival (89, 91). Heinz bodies are composed of denatured proteins, primarily hemoglobin. Cats exposed orally to 1.2, 1.6, 2.4, and 3.6 g/kg bw/day of propylene glycol for 2, 5, or 17 weeks developed increased numbers of RBCs with Heinz bodies. The cat is very sensitive to propylene glycol toxicity, with a 0.44 mg/kg bw/day dose reported to result in Heinz body formation in erythrocytes (reviewed by OECD (27)). This sensitivity occurs at concentrations that were present in soft moist cat foods and lead the FDA to remove propylene glycol from cat foods in 1996 (9).

In a study by Weil et al. (83) dogs were fed propylene glycol at 2 and 5 g/kg bw/day through the diet. Significant hematological changes were noted in the high dose group after two years; hemoglobin, hematocrit, and total erythrocyte counts were lower, whereas, poikilocytes and reticulocytes were increased.

Strength/Weaknesses: There are few and inconsistent changes in hematologic parameters in the Suber et al. (85) study. No inferences can be made for erythropoiesis.

Strength/Weaknesses: Saini et al. (90) reported hematologic effects of propylene glycol in rats administered single doses of 0.7 or 3 g/kg bw by gavage. There is sufficient experimental detail given to deem the results reliable. However, Gaunt et al. (81) did not find any hematologic effect after feeding about 2 g/kg bw/day for 2 years. It is very likely that the acute changes seen by Saini et al. (90) have been overcome by 2 years due to adaptation.

Utility (Adequacy) for CERHR Evaluation Process: This is a useful report (90) that confirms that the hematopoietic system is also a target of propylene glycol in rats, albeit at higher chronic doses than in cats, dogs, and probably monkeys.

Strength/Weaknesses: The Robertson et al. (86) study has a very large uncertainty attached to it, as discussed earlier, and provides marginal evidence of a hematologic effect in non-human primates.

Utility (Adequacy) for CERHR Evaluation Process: The hemolytic capability of propylene glycol has been demonstrated in vitro in human erythrocytes (78). However, the primate data presented by Robertson et al. (86) do not provide evidence of a hematological effect of propylene glycol on primates.

Strength/Weaknesses: Christopher et al. (91) reported D-lactic acidosis and Heinz body formation in cats administered daily 1.6 or 8 g/kg propylene glycol for up to 35 days. Authors conclusively demonstrated a dose-dependent reduction of erythrocyte survival. Bauer et al. (89) confirms in essence the findings of Christopher et al. (91) and refines the dose response on Heinz body formation and eryth