Jeffrey S. Knutsen

Cellulase Retention and Sugar Removal by Membrane Ultrafiltration During Lignocellulosic Biomass Hydrolysis

Technologies suitable for the separation and reuse of cellulase enzymes during the enzymatic saccharification of pretreated corn stover are investigated to examine the economic and technical viability of processes that promote cellulase reuse while removing inhibitory reaction products such as glucose and cellobiose. The simplest and most suitable separation is a filter with relatively large pores on the order of 20–25 mm that retains residual corn stover solids while passing reaction products such as glucose and cellobiose to form a sugar stream for a variety of end uses. Such a simple separation is effective because cellulase remains bound to the residual solids. Ultrafiltration using 50-kDa polyethersulfone membranes to recover cellulase enzymes in solution was shown not to enhance further the saccharification rate or overall conversion. Instead, it appears that the necessary cellulase enzymes, including β-glucosidase, are tightly bound to the substrate; when fresh corn stover is contacted with highly washed residual solids, without the addition of fresh enzymes, glucose is generated at a high rate. When filtration was applied multiple times, the concentration of inhibitory reaction products such as glucose and cellobiose was reduced from 70 to 10 g/L. However, an enhanced saccharification performance was not observed, most likely because the concentration of the inhibitory products remained too high. Further reduction in the product concentration was not investigated, because it would make the reaction unnecessarily complex and result in a product stream that is much too dilute to be useful. Finally, an economic analysis shows that reuse of cellulase can reduce glucose production costs, especially when the enzyme price is high. The most economic performance is shown to occur when the cellulase enzyme is reused and a small amount of fresh enzyme is added after each separation step to replace lost or deactivated enzyme.

Cellulase recovery via membrane filtration

A combined sedimentation and membrane filtration process was investigated for recycling cellulase enzymes in the biomass-to-ethanol process. In the first stage, lignocellulose particles longer than approx 50 μm were removed by means of sedimentation in an inclined settler. Microfiltration was then utilized to remove the remaining suspended solids. Finally, the soluble cellulase enzymes were recovered by ultrafiltration. The perm eate fluxes obtained in microfiltration and ultrafiltration were approx 400 and 80 L/(m2·h), respectively. A preliminary economic analysis shows that the cost benefit of enzyme recycling may be as much as 18 cents/gal of ethanol produced, provided that 75% of the enzyme is recycled in active form.

Combined Sedimentation and Filtration Process for Cellulase Recovery During Hydrolysis of Lignocellulosic Biomass

A combined sedimentation and ultrafiltration process was investigated for recovering cellulase enzymes during the hydrolysis of lignocellulosic biomass. Lignocellulosic particles larger than approx 50 microm in length were first removed via sedimentation using an inclined settler. Ultrafiltration was then used to retain the remaining lignocellulosic particles and the cellulose enzymes, while transmitting fermentable sugars and other small molecules. The permeate flux from the ultrafiltration step for a feed consisting of 0.22 w/v% cellulase is 64+/-5 L/m2-h, while that for a feed consisting of the settler overflow from a mixture 0.22 w/v% cellulase and 10 wt% lignocellulose fed to the settler is 130+/-20 L/m2-h. The higher permeate flux in the latter case is presumably due to binding of a portion of the cellulase enzymes to the lignocellulosic particles during hydrolysis and filtration, preventing the enzymes from fouling the membrane. A filter paper activity assay shows little loss in enzymatic activity throughout the combined sedimentation/ultrafiltration separation process.

Worker Exposure to Methanol Vapors During Cleaning of Semiconductor Wafers in a Manufacturing Setting

An exposure simulation was conducted to characterize methanol exposure of workers who cleaned wafers in quality control departments within the semiconductor industry. Short-term (15 min) and long-term (2–4 hr) personal and area samples (at distances of 1 m and 3–6 m from the source) were collected during the 2-day simulation. On the first day, 45 mL of methanol were used per hour by a single worker washing wafers in a 102 m3 room with a ventilation rate of about 10 air changes per hour (ACH). Virtually all methanol volatilized. To assess exposures under conditions associated with higher productivity, on the second day, two workers cleaned wafers simultaneously, together using methanol at over twice the rate of the first day (95 mL/hr). On this day, the ventilation rate was halved (5 ACH). Personal concentrations on the first day averaged 60 ppm (SD = 46 ppm) and ranged from 10–140 ppm. On the second day, personal concentrations for both workers averaged 118 ppm (SD = 50 ppm; range: 64–270 ppm). Area concentrations measured on the first day at 1 m from the source and throughout the balance of the room averaged 29 ppm (SD = 19 ppm; range: 4–83 ppm) and 18 ppm (SD = 12 ppm; range: 3–42 ppm), respectively. As expected, area concentrations measured on the second day were higher than the first and averaged 73 ppm (SD = 25 ppm; range: 27–140 ppm) at 1 meter and 48 ppm (SD = 13 ppm; range: 21–67 ppm) throughout the balance of the room. The results of this simulation suggest that the use of methanol to clean semiconductor wafers without the use of local exhaust ventilation and with relatively low room ventilation rates is unlikely to result in worker exposures exceeding the current ACGIH® threshold limit value of 200 ppm. This study also confirmed prior studies suggesting that when a relatively volatile chemical is located within arms length (near field), breathing zone concentrations will be about two- to threefold greater than the room concentration when the air exchange rate is 5–10 ACH.

Airborne Concentrations of Benzene Associated with the Historical Use of Some Formulations of Liquid Wrench

The current study characterizes potential inhalation exposures to benzene associated with the historical use of some formulations of Liquid Wrench under specific test conditions. This product is a multiuse penetrant and lubricant commonly used in a variety of consumer and industrial settings. The study entailed the remanufacturing of several product formulations to have similar physical and chemical properties to most nonaerosol Liquid Wrench formulations between 1960 and 1978. The airborne concentrations of benzene and other constituents during the simulated application of these products were measured under a range of conditions. Nearly 200 breathing zone and area bystander air samples were collected during 11 different product use scenarios. Depending on the tests performed, average airborne concentrations of benzene ranged from approximately 0.2–9.9 mg/m3 (0.08–3.8 ppm) for the 15-min personal samples; 0.1–8 mg/m3 (0.04–3 ppm) for the 1-hr personal samples; and 0.1–5.1 mg/m3 (0.04–2 ppm) for the 1-hr area samples. The 1-hr personal samples encompassed two 15-min product applications and two 15-min periods of standing within 5 to 10 feet of the work area. The measured airborne concentrations of benzene varied significantly based on the benzene content of the formulation tested (1%, 3%, 14%, or 30% v/v benzene) and the indoor air exchange rate but did not vary much with the base formulation of the product or the two quantities of Liquid Wrench used. The airborne concentrations of five other volatile chemicals (ethylbenzene, toluene, total xylenes, cyclohexane, and hexane) were also measured, and the results were consistent with the volatility and concentrations of these chemicals in the product tested. A linear regression analysis of air concentration compared with the chemical mole fraction in the solution and air exchange rate provided a relatively good fit to the data. The results of this study should be useful for evaluating potential inhalation exposures to benzene and other volatile chemicals that occurred during the past use of some formulations of Liquid Wrench and perhaps for some similar products containing these chemicals.

Airborne Concentrations of Asbestos Onboard Maritime Shipping Vessels (1978-1992)

The exposure of shipyard workers to asbestos has been frequently investigated during the installation, repair or removal of asbestos insulation. The same level of attention, however, has not been directed to asbestos exposure of maritime seamen or sailors. In this paper, we assemble and analyze historical industrial hygiene (IH) data quantifying airborne asbestos concentrations onboard maritime shipping vessels between 1978 and 1992. Air monitoring and bulk sampling data were compiled from 52 IH surveys conducted on 84 different vessels, including oil tankers and cargo vessels, that were docked and/or at sea, but these were not collected during times when there was interaction with asbestos-containing materials (ACMs). One thousand and eighteen area air samples, 20 personal air samples and 24 air samples of unknown origin were analyzed by phase contrast microscopy (PCM); 19 area samples and six samples of unknown origin were analyzed by transmission electron microscopy (TEM) and 13 area air samples were analyzed by scanning electron microscopy (SEM). In addition, 482 bulk samples were collected from suspected ACMs, including insulation, ceiling panels, floor tiles, valve packing and gaskets. Fifty-three percent of all PCM and 4% of all TEM samples were above their respective detection limits. The average airborne concentration for the PCM area samples (n = 1018) was 0.008 fibers per cubic centimeter (f cc(-1)) (95th percentile of 0.040 f cc(-1)). Air concentrations in the living and recreational areas of the vessels (e.g. crew quarters, common rooms) averaged 0.004 f cc(-1) (95th percentile of 0.014 f cc(-1)), while air concentrations in the engine rooms and machine shops averaged 0.010 f cc(-1) (95th percentile of 0.068 f cc(-1)). Airborne asbestos concentrations were also classified by vessel type (cargo, tanker or Great Lakes), transport status (docked or underway on active voyage) and confirmed presence of ACM. Approximately 1.3 and 0% of the 1018 area samples analyzed by PCM exceeded 0.1 and 1 f cc(-1), respectively. This data set indicates that historic airborne asbestos concentrations on these maritime shipping vessels, when insulation-handling activities were not actively being performed, were consistently below contemporaneous US occupational standards from 1978 until 1992, and nearly always below the current permissible exposure limit of 0.1 f cc(-1).

A calibrated human PBPK model for benzene inhalation with urinary bladder and bone marrow compartments.

A physiologically-based pharmacokinetic (PBPK) model of benzene inhalation based on a recent mouse model was adapted to include bone marrow (target organ) and urinary bladder compartments. Empirical data on human liver microsomal protein levels and linked CYP2E1 activities were incorporated into the model, and metabolite-specific conversion rate parameters were estimated by fitting to human biomonitoring data and adjusting for background levels of urinary metabolites. Human studies of benzene levels in blood and breath, and phenol levels in urine were used to validate the rate of human conversion of benzene to benzene oxide, and urinary benzene metabolites from Chinese benzene worker populations provided model validation for rates of human conversion of benzene to muconic acid (MA) and phenylmercapturic acid (PMA), phenol (PH), catechol (CA), hydroquinone (HQ), and benzenetriol (BT). The calibrated human model reveals that while liver microsomal protein and CYP2E1 activities are lower on average in humans compared to mice, the mouse also shows far lower rates of benzene conversion to MA and PMA, and far higher conversion of benzene to BO/PH, and of BO/PH to CA, HQ, and BT. The model also differed substantially from existing human PBPK models with respect to several metabolic rate parameters of importance to interpreting benzene metabolism and health risks in human populations associated with bone marrow doses. The model provides a new methodological paradigm focused on integrating linked human liver metabolism data and calibration using biomonitoring data, thus allowing for model uncertainty analysis and more rigorous validation.

Airborne benzene exposures from cleaning metal surfaces with small volumes of petroleum solvents.

Airborne benzene concentrations were measured in a room with controlled air exchange during surface cleaning with two petroleum-based solvents (a paint thinner and an engine degreaser). The solvents were spiked with benzene to obtain target concentrations of 0.001, 0.01, and 0.1% by volume in the liquid. Personal samples on the worker and area samples up to 1.8m away were collected over 12 events (n=84 samples) designed to examine variation in exposure with solvent type, cleaning method (rag wipe or spatula scrape), surface area cleaned, air exchange rate, solvent volume applied, and distance from the cleaned surface. Average task breathing zone concentrations of benzene represented by 18-32 min time-weighted averages were 0.01 ppm, 0.05 ppm, and 0.27 ppm, when the solvents contained approximately 0.003, 0.008, and 0.07% benzene. Solvent benzene concentration, volume applied, and distance from the handling activities had the greatest effect on airborne concentrations. The studied solvent products containing 0.07% benzene (spiked) did not exceed the current OSHA permissible exposure limit of 1 ppm (averaged over 8h) or the ACGIH Threshold Limit Value of 0.5 ppm, in any of the tested short-term exposure scenarios. These data suggest that, under these solvent use scenarios, petroleum-based solvent products produced in the United States after 1978 likely did not produce airborne benzene concentrations above those measured if the concentration was less than 0.1% benzene.

Historical Analysis of Airborne Beryllium Concentrations at a Copper Beryllium Machining Facility (1964–2000)

Copper beryllium alloys are the most commonly used form of beryllium; however, there have been few studies assessing occupational exposure in facilities that worked exclusively with this alloy versus those where pure metal or beryllium oxide may also have been present. In this paper, we evaluated the airborne beryllium concentrations at a machining plant using historical industrial hygiene samples collected between 1964 and 2000. With the exception of a few projects conducted in the 1960s, it is believed that >95% of the operations used copper beryllium alloy exclusively. Long-term (>120 min) and short-term (<120 min) personal and area samples were collected during a variety of activities including machining of copper beryllium-containing parts, as well as finishing operations (e.g., deburring and polishing) and decontamination of machinery. A total of 580 beryllium air samples were analyzed (311 personal and 269 area samples). The average concentration based on area samples (1964-2000) was 0.021 microg m(-3) (SD 0.17 microg m(-3); range 0.00012-2.5 microg m(-3)); 68.8% were below the analytical limit of detection (LOD). The average airborne beryllium concentration, based on all personal samples available from 1964 through the end of 2000 (n = 311), was 0.026 microg m(-3) (SD 0.059 microg m(-3); range 0.019-0.8 microg m(-3)); 97.4% were below the LOD. Personal samples collected from machinists (n = 78) had an average airborne concentration of 0.021 microg m(-3) (SD 0.014 microg m(-3); range 0.019-0.14 microg m(-3)); 97.4% were below the LOD. Airborne concentrations were consistently below the Occupational Safety and Health Administration permissible exposure limit for beryllium (2 microg m(-3)). Overall, the data indicate that for machining operations involving copper beryllium, the airborne concentrations for >95% of the samples were below the contemporaneous occupational exposure limits or the 1999 Department of Energy action level of 0.2 microg m(-3) and, in most cases, were below the LOD.

Determinants of carbon monoxide exposure inside a motor home during on-board generator use

Carbon monoxide (CO) is a well-known asphyxiant. As part of an incident investigation involving two fatalities, a study was conducted to determine key factors that influence CO concentrations inside motor homes/ recreational vehicles. Test parameters examined included the condition of the on-board generator exhaust pipe (attached/detached), generator load (<1–20 amps), position of ventilation hatches (open/closed), parking location (adjacent/perpendicular to a masonry wall), and weather conditions (breezy/ calm). A tracer gas test was also performed of the motor home undercarriage because of concerns for possible damage (no visible damage was observed). Results showed that all five parameters affected the CO concentrations detected within the motor home, but the generator exhaust tailpipe was found to have the greatest impact. Further, a specific combination of conditions, along with documented invisible undercarriage leaks, was necessary for CO concentrations to become high enough to produce acutely toxic and fatal conditions inside the motor home.