72 Fermentation and Biochemical Engineering Handbook The equipment needed in warehousing are fork lift trucks, floor- washing machines, etc. Special materials must be on hand to clean up spills quickly, according to federal regulations. Good housekeeping and pest control are essential 2.4 Media Preparation or Batching Area For good housekeeping, all equipment should be on or above the floor and no pits should be used. On the other hand, grated trenches make it easy to clean the floors and minimize the number of floor drains The number, shape and volume of batching tanks that different companies use show personal preference and are not very important. Usually two or three different sized tanks are used smaller batching tanks are for inoculum tanks and the larger tanks for feed and fermenter mediapreparation The type of agitation varies widely. Batching tanks, 10,000 gallons and smaller, could be specified as 304 stainless steel, dished or flat bottom and heads, HD ratio about 0. 7 to keep a working platform low, a slow speed (60 to 90 rpm)top-entering agitator with airfoil type impellers, horsepower approximately 1. 25 per 1000 gallons. The tanks need to be equipped with submerged (bottom)nozzles which are supplied with both steam and air. Hot and cold water are usually piped to the top. The hatch, with a removable grate of v/ 2 " S/S rod on 6"x6"centers, should be as large as a 100 lb bag of raw materials. a temperature recorder is the minimum instrumentation. The cyclone, with a rotary air lock valve to permit material additions from the bulk storage silos, is normally located above the tank(s). For tanks larger thar 10,000 gallons, the bottom head should be dished, the H/D ratio made l to 2, and airfoil type agitators used The size and number of batching tanks depend upon whether the plant uses continuous sterilizers or batch sterilization. The difference is that in the latter case, the tanks can be large(50 to 80%of the size of the fermenter), and usually all the materials are mixed together. For continuous sterilizers, there is usually a minimum of four smaller tanks so that proteins, carbohydrates and salts can be batched and sterilized separately. In this case, the tanks are consider ably smaller than the fermenter The media preparation area is also where hydrolysates of proteins, and starches, as well as special processing of steep liquor, molasses and other crude materials takes place. Very strict accuracy of weights, volumes, pH adjustments and processing instructions are the first step to reproducible fermentation results. A well-run batching area depends upon purchasing a uniform quality of raw materials, adequate equipment, detailed batching
72 Fermentation and Biochemical Engineering Hun dbook The equipment needed in warehousing are fork lift trucks, floorwashing machines, etc. Special materials must be on hand to clean up spills quickly, according to federal regulations. Good housekeeping and pest control are essential. 2.4 Media Preparation or Batching Area For good housekeeping, all equipment should be on or above the floor and no pits should be used. On the other hand, grated trenches make it easy to clean the floors, and minimize the number of floor drains. The number, shape and volume of batching tanks that different companies use show personal preference and are not very important. Usually two or three different sized tanks are used; smaller batching tanks are for inoculumtanks and the largertanks for feed and fermenter media preparation. The type of agitation varies widely. Batching tanks, 10,000 gallons and smaller, could be specified as 304 stainless steel, dished or flat bottom and heads, WD ratio about 0.7 to keep a working platform low, a slow speed (60 to 90 rpm) top-entering agitator with airfoil type impellers, horsepower approximately 1.25 per 1000 gallons. The tanks need to be equipped with submerged (bottom) nozzles which are supplied with both steam and air. Hot and cold water are usually piped to the top. The hatch, with a removable grate of %" S/S rod on 6" x 6" centers, should be as large as a 100 lb. bag of raw materials. A temperature recorder is the minimum instrumentation. The cyclone, with a rotary air lock valve to permit material additions from the bulk storage silos, is normally located above the tank(s). For tanks larger than 10,000 gallons, the bottom head should be dished, the WD ratio made 1 to 2, and airfoil type agitators used. The size and number of batching tanks depend upon whether the plant uses continuous sterilizers or batch sterilization. The difference is that in the latter case, the tanks can be large (50 to 80% of the size ofthe fermenter), and usually all the materials are mixed together. For continuous sterilizers, there is usually a minimum of four smaller tanks so that proteins, carbohydrates and salts can be batched and sterilized separately. In this case, the tanks are considerably smaller than the fermenter. The media preparation area is also where hydrolysates of proteins, and starches, as well as special processing of steep liquor, molasses and other crude materials takes place. Very strict accuracy of weights, volumes, pH adjustments and processing instructions are the first step to reproducible fermentation results. A well-run batching area depends upon purchasing a uniform quality of raw materials, adequate equipment, detailed batching
Fermentation Design 73 instructions and well trained, reliable personnel. Record keeping of batch quantities, lot numbers, pH, temperatures, etc are necessary for quality and good manufacturing practices 2.5 The Seed Fermenter Layout Some companies prefer to locate all the seed fermenters in one area so that a group of workmen become specialists in batch sterilizing, inoculating, and coddling the first(plant)inoculum stage to maturity. Other companies locate the seed fermenters adjacent to the fermenters. Small plants cannot afford to isolate equipment and have a specialized work force, however, large plants do isolate groups of similar equipment, and specialize the work force, which often results in higher productivity The operation of fermenters is basically the same regardless of size, but seed fermenters usually do not have sterile anti-foam and nutrient feeds piped to the tanks as the main fermenters have. Therefore, foaming in the seed fermenters can lead to infection, which is one of the reasons they need more attention. Careful inoculation procedures, sampling and sterilizing the transfer lines from the seed fermenter require alert personnel. Careful attention to these details is more important than the proximity of the seed and main fermenters The number of inoculum stages or scale-up is traditional. The rule of a tenfold volume increase per stage is followed by some companies, but is not critical. The multiplication rate of an organism is constant after the lag phase so the amount of cell mass developed to inoculate the next stage, minus the starting amount, is a matter of time, providing, of course, there is sufficient substrate and environmental conditions are reasonable. After all, the theory is that one foreign organism or spore, if not killed during sterilization, will, in time, contaminate the fermenter. Larger cell masses of inoculum can shorten the growth phase of the next larger stage. Using this concept, some companies make the inoculum volume larger than a tenth of the fermenter volume so that the number of transfers from laboratory flask to the final fermenter is minimum. This also assumes there is a higher risk of infection during transfers as well as a certain viability loss. a higher inoculum cell mass may reduce the lag time in the fermenter. This, combined with using continuous sterilization for a short"turn around" time of the fermenter can increase productivity for little or no cost
Fermentation Design 73 instructions and well trained, reliable personnel. Record keeping of batch quantities, lot numbers, pH, temperatures, etc. are necessary for quality and good manufacturing practices. 2.5 The Seed Fermenter Layout Some companies prefer to locate all the seed fermenters in one area so that a group of workmen become specialists in batch sterilizing, inoculating, and coddling the first (plant) inoculum stage to maturity. Other companies locate the seed fermenters adjacent to the fermenters. Small plants cannot afford to isolate equipment and have a specialized work force, however, large plants do isolate groups of similar equipment, and specialize the work force, which often results in higher productivity. The operation of fermenters is basically the same regardless of size, but seed fermenters usually do not have sterile anti-foam and nutrient feeds piped to the tanks as the main fermenters have. Therefore, foaming in the seed fermenters can lead to infection, which is one of the reasons they need more attention. Careful inoculation procedures, sampling and sterilizing the transfer lines from the seed fermenter require alert personnel. Careful attention to these details is more important than the proximity ofthe seed and main fermenters. The number of inoculum stages or scale-up is traditional. The rule of a tenfold volume increase per stage is followed by some companies, but is not critical. The multiplication rate ofan organism is constant after the lag phase so the amount of cell mass developed to inoculate the next stage, minus the starting amount, is a matter of time, providing, of course, there is sufficient substrate and environmental conditions are reasonable. Mer all, the theory is that one foreign organism or spore, if not killed during sterilization, will, in time, contaminate the fermenter. Larger cell masses of inoculum can shorten the growth phase of the next larger stage. Using this concept, some companies make the inoculum volume larger than a tenth of the fermenter volume so that the number of transfers from laboratory flask to the final fermenter is minimum. This also assumes there is a higher risk of infection during transfers as well as a certain viability loss. A higher inoculum cell mass may reduce the lag time in the fermenter. This, combined with using continuous sterilization for a short “turn around” time of the fermenter, can increase productivity for little or no cost
74 Fermentation and Biochemical Engineering Handbook 2.6 The main Fermenter layout For simplicity ofpiping, especially the utility piping, the fermenters are usually placed in a straight line, sometimes two or more parallel lines. In this manner the plant is easily expanded, and other tank layouts do not seem as convincing. It is desirable to have the working platform extend completely around the circumference of the top dish, and to have enough room between tanks for maintenance carts(1 to 1.5 meters). Good lighting and ventilation on the working platform should not be overlooked. Using water from hoses for cleaning is common so care must be taken to have nonskid floors with adequate drains, especially at the top of stairs. Open floor grating is not desirable. All structural steel should be well primed to prevent corrosion from the very humid atmosphere. Electronic instrumentation and computers must be placed in control rooms which run at constant(HVAC)temperature. Most fermenter buildings are between 40 and 100 feet high, making it possible to have one or more floors between the ground floor and the main fermenter working platform. The intermediate floors can be used for the utility and process piping, sterileair filters, the sterile anti-foam system, instrumentation sensors(temperature, pH, DO, etc. ) heat exchangers, motor control center, laboratories and offices. Buildings 40 feet or more high frequently have elevators installed Fermenters can be located outdoors in most countries of world working platforms usually are enclosed and heated in temperate zones only shaded in more tropical zones. In more populated areas, open fermenter buildings make too much noise for local residents. The environmental awareness,or the tolerance of the public, could preclude open fermenter buildings in the future. Odor is also offensive to the public. The environmen tal authorities are demanding that equipment be installed to eliminate the offensive odor of the off-gases. (Noise levels inside a fermenter building will be greater than 90 dBa if no preventive measures are taken. Harvest tanks can be justified as the responsibility of the fermentation or recovery department. They are economical ( carbon or stainless steel) with a shape described by(hd= 1)and should be insulated and equipped with cooling coils and agitator(s) 2.7 Nutrient Feed Tanks Essential equipment to a productive fermentation department are sterilizable tanks for nutrient feeds. Multiproduct plants usually require everal different sizes of feed tanks: i) a small volume to be transferred once
74 Fermentation and Biochemical Engineering Handbook 2.6 The Main Fermenter Layout For simplicity of piping, especially the utility piping, the fermenters are usually placed in a straight line, sometimes two or more parallel lines. In this manner the plant is easily expanded, and other tank layouts do not seem as convincing. It is desirable to have the working platform extend completely around the circumference of the top dish, and to have enough room between tanks for maintenance carts (1 to 1.5 meters). Good lighting and ventilation on the working platform should not be overlooked. Using water from hoses for cleaning is common so care must be taken to have nonskid floors with adequate drains, especially at the top of stairs. Open floor grating is not desirable. All structural steel should be well primed to prevent corrosion from the very humid atmosphere. Electronic instrumentation and computers must be placed in control rooms which run at constant (HVAC) temperature. Most fermenter buildings are between 40 and 100 feet high, making it possible to have one or more floors between the ground floor and the main fermenter working platform. The intermediate floors can be used for the utility and process piping, sterile air filters, the sterile anti-foam system, instrumentation sensors (temperature, pH, DO, etc.), heat exchangers, motor control center, laboratories and offices. Buildings 40 feet or more high frequently have elevators installed. Fermenters can be located outdoors inmost countries ofthe world. The working platforms usually are enclosed and heated in temperate zones, and only shaded in more tropical zones. In more populated areas, open fermenter buildings make too much noise for local residents. The environmental awareness, or the tolerance of the public, could preclude open fermenter buildings in the future. Odor is also offensive to the public. The environmental authorities are demanding that equipment be installed to eliminate the offensive odor ofthe off-gases. (Noise levels inside a fermenter building will be greater than 90 dBA if no preventive measures are taken.) Harvest tanks can be justified as the responsibility of the fermentation or recovery department. They are economical (carbon or stainless steel) with a shape described by (H/D z 1) and should be insulated and equipped with cooling coils and agitator(s). 2.7 Nutrient Feed Tanks Essential equipment to a productive fermentation department are sterilizable tanks for nutrient feeds. Multiproduct plants usually require several different sizes of feed tanks: (i) a small volume to be transferred once
Fermentation Design every 12 or 24 hours such as a nitrogen source;(ii) a large volume carbohydrate solution fed continuously, perhaps varying with the fermenter olume, (iii precursor feed, fed in small amounts relative to assay data; (iv) anti-foam( Some companies prefer a separate anti-foam feed system for each fermenter. A continuously sterilizing systemfor anti-foam is discussed below which is capable of servicing all the fermenters. ) /v other tanks for ac bases, salts, etc. Many companies prefer to batch sterilize a known qu and transfer the entire contents quickly. Sometimes, the feeds require programming the addition rate to achieve high productivity. In this latter case, large volume tanks are used and the contents are presterilized ( batch or continuous)or the feed is continuously sterilized between the feed tank and the fermenter. Usually feed tanks are not designed as fermenters, even though they are sterilizable, and there is no need for high volume air flow, but only sufficient air pressure for the transfer. For solvablenutrients the agitator and anti-foam system are not required Since the air requirements are needed only to transfer the feed, the air piping design is different and the sterile air filter is proportionately smaller. Instrumentation is usually limited to temperature, pressure and volume. The H/D ratio ofthe vessel can be near one foreconomy and need not be designed for the aeration/agitation requirements of a fermenter 2. 8 Sterile Filters Sterile air filtration is simple today with the commercial units readily available. However, some companies still design their own(see Aiba, Humphrey and Millis 2 )to use a variety of filter media such as carbon cotton,glass staple, etc. (For recent papers about industrial applications of artridge filters, see Bruno 3I and Perkowski. 14) The essential method to obtain sterile air, whether packed-bed or cartridge filters are used, is to reduce the humidity ofthe airafter compression so that the filter material always remains dry. The unsterilized compressed air must neverreach 100%relative humidity. Largerplants install instrumen- tation with alarms set at about 85% relative humidity Careful selection of the cartridge design or the design of packed-bed filters will result in units that can operate in excess of three years without replacement of filter media. If a fiber material is used in a packed-bed type filter, the finer the fiber diameter the shallower the bed depth needs to be for efficient filtration. Other filter media are less common and tend to have special problems and/or shorter life The bed depth of filters is only 10 to 18 inches for fibers of less than 1( microns. These filters run"clean"for 2 weeks or longer before being resterilized
Fermentation Design 75 every 12 or 24 hours such as a nitrogen source; (ii) a large volume carbohydrate solution fed continuously, perhaps varying with the fermenter volume; (iii) aprecursor feed, fed in small amounts relative to assay data; (iv) anti-foam (Some companies prefer a separate anti-foam feed system for each fermenter. A continuously sterilizing system for anti-foam is discussed below which is capable of servicing all the fermenters.); (v) other tanks for acids, bases, salts, etc. Many companies prefer to batch sterilize a known quantity and transfer the entire contents quickly. Sometimes, the feeds require programming the addition rate to achieve high productivity. In this latter case, large volume tanks are used and the contents are presterilized (batch or continuous) or the feed is continuously sterilized between the feed tank and the fermenter. Usually feed tanks are not designed as fermenters, even though they are sterilizable, and there is no need for high volume air flow, but only sufficient air pressure for the transfer. For solvable nutrients the agitator and anti-foam system are not required. Since the air requirements are needed only to transfer the feed, the air piping design is different and the sterile air filter is proportionately smaller. Instrumentation is usually limited to temperature, pressure and volume. The Hn> ratio ofthe vessel can be near one for economy and need not be designed for the aeratiodagitation requirements of a fermenter. 2.8 Sterile Filters Sterile air filtration is simple today with the commercial units readily available. However, some companies still design their own (see Aiba, Humphrey and Millisr2]) to use a variety of filter media such as carbon, cotton, glass staple, etc. (For recent papers about industrial applications of cartridge filters, see Bruno[3] and Perkowski.r4]) The essential method to obtain sterile air, whether packed-bed or cartridge filters are used, is to reduce the humidity ofthe air after compression so that the filter material always remains dry. The unsterilized compressed air must never reach 100% relative humidity. Largerplants install instrumentation with alarms set at about 85% relative humidity. Carehl selection of the cartridge design or the design of packed-bed filters will result in units that can operate in excess of three years without replacement of filter media. If a fiber material is used in a packed-bed type filter, the finer the fiber diameter the shallower the bed depth needs to be for efficient filtration. Other filter media are less common and tend to have special problems and/or shorter life. The bed depth of filters is only 10 to 18 inches for fibers of less than 10 microns. These filters run “clean” for 2 weeks or longer before being resterilized