Patent Application
20220315966
The present invention pertains to a method for producing fermentation products, and in particular to a fermentation product manufacturing method and sensor device for producing fermentation products by the fermenting operation of a fermentation vessel into which bubbles are mixed, and in which crystals of an average particle size of 5 μm or larger are produced in a liquid.
The production of various fermentation products requires measurement of various properties such as the electrical conductivity and turbidity of liquid in a fermentation vessel, and the concentration of specific components in the liquid. Japanese Patent No. 3074781 (Patent Document 1) describes a method for manufacturing L-lysine. In this production method, a carbon source is added to maintain a carbon source concentration at 5 g/L or less in the culture liquid. I.e., in this method, carbon source concentration is measured by sampling the culture liquid in a timely manner and directly analyzing carbon source concentration, or by measuring pH and dissolved oxygen concentration to sense a carbon source deficiency from changes therein, so as to control feeding of the medium.
Although it is relatively easy to extract a portion of liquid to be measured from the manufacturing process and measure properties of the liquid, as in the invention described in Patent Document 1, inline measurement without extracting liquid from the manufacturing process is preferable from a manufacturing efficiency standpoint. However, it can be difficult to measure liquid properties inline, particularly if bubbles are present in the liquid to be measured, or crystals are formed in the liquid. For example, bubbles of supplied oxygen or air, or bubbles of the carbon dioxide gas metabolic product of microorganisms themselves in culture may be mixed into the culture liquid during aerated culture, introducing noise into the readings, or increasing measurement errors.
Japanese Patent No. 4420168 (Patent Document 2) describes a turbidity sensor. The turbidity sensor has a hollow semicylindrical member made of stainless steel, with a test solution inlet and an automatically opening and closing swing valve at the bottom, and a hole at the top for venting bubbles. In addition, a wetted photometric portion of a laser turbidimeter is disposed at the tip position on the inside of the hollow semicylinder. When measuring with this turbidity sensor, a swing valve is first opened to replace the test solution inside the hollow semicylinder. The swing valve is then closed, bubbles in the hollow semicylinder are discharged through bubble vent holes and, after the detected turbidity is allowed to stabilize, turbidity is measured. In the Patent Document 2 invention, the effect of bubbles in a liquid being tested is thus reduced.
However, the turbidity sensor of Patent Document 2 requires waiting for the detection value to stabilize after the swing valve is closed, making it difficult to perform real-time detection. If the turbidity sensor is applied to a liquid in which crystals form in the liquid, there is a risk that crystals may accumulate on moving parts such as the swing valve, causing failures. In addition, the requirement in the Patent Document 2 turbidity sensor for a swing valve which is opened and closed by remote control complicates the structure, leading to the problem of time-consuming maintenance to achieve stable operation over long durations.
On the other hand, when a sensor is applied to a liquid in which crystals are produced, there is a need not only to suppress the effect of the crystals, but also to analyze, simultaneously with the liquid phase, the amount of solids precipitated when the concentration of the fermentation product in the liquid phase exceeds its solubility. Here, when a sensor cover is provided on the sensor to suppress the effect of bubbles, crystals are deposited near the sensor and inside the sensor cover, making it difficult to accurately measure the concentration of products. Measures such as discharging crystals precipitated inside the sensor cover as needed are therefore required to keep the crystal concentration in the fermentation vessel and the crystal concentration around the sensor at the same levels, so as to prevent such deposition.
The present invention therefore has the object of providing a fermentation product manufacturing method and sensor device for same with which, using a simple structure, the effects on measurement results by bubbles mixed into a liquid are suppressed, deposition of crystals produced in the liquid around the sensor and within the sensor cover is prevented, properties of liquid and solids within a fermenting vessel are measured and fermentation operations can be conducted based on those measurements.
To solve the above-described problems, the present invention is a method for manufacturing fermentation products in which the fermentation products are produced by fermentation operation of a fermentation vessel in which bubbles are mixed and crystals with an average particle size of 5 μm or greater are formed in a liquid, comprising steps of; preparing a fermentation vessel and a sensor device for measuring properties of liquid inside the fermentation vessel; introducing liquid to be fermented into the fermentation vessel; and operating the fermentation vessel for fermentation, wherein properties of liquid in the fermentation vessel are measured by the sensor device, and conditions of fermentation operation are adjusted based on results of the measurement; wherein the sensor device includes a sensor for measuring liquid properties and a cover body for the sensor disposed to surround the sensor; wherein the cover body includes a bottom side permeable portion for passing at least a portion of the liquid and crystals in the liquid disposed on at least a portion of the bottom surface of the cover body and a top side permeable portion for passing at least a portion of the liquid and crystals in the liquid disposed on at least a portion of the top surface of the cover body; and wherein the bottom side permeable portion and the top side permeable portion have numerous micropores for passing liquid respectively, and the micropores disposed on the top side permeable portion are sized to be the same as or larger than the micropores disposed on the bottom side permeable portion.
In the invention thus constituted, the sensor device for measuring properties of liquid in the fermentation vessel has a cover body disposed so as to surround the sensor. Bottom and top permeable portions through which a liquid and at least some of the crystals in the liquid can pass is provided on this cover body, therefore liquid to be measured is able to constantly flow in and out of the cover body. Liquid inside the cover body is therefore constantly replaced, and sensors disposed on the cover body can continuously measure liquid properties in real time. Because the bottom and top permeable portions of the cover body allow at least some of the crystals in the liquid to pass through, crystals formed in the liquid are less likely to accumulate in the cover body, therefore fermentation operations can continue for a long duration without performing maintenance such as cleaning the cover body. Since micropores in the top permeable portion have a size equal to or greater than micropores disposed on the bottom permeable portion, bubbles floating up from below the cover body have difficulty passing through the micropores in the bottom permeable portion, thus enabling penetration by bubbles into the cover body to be effectively suppressed. On the other hand micropores in the top permeable portion are the same or larger than those in the bottom permeable portion, therefore bubbles penetrating into the cover body can easily float up within the cover body and pass through the top permeable portion to be discharged. Bubbles penetrating into the cover body can thus be prevented from accumulating therein, so adverse effects on sensor readings can be effectively suppressed.
In the present invention the average diameter of the bubbles mixed into the fermentation vessel liquid is preferably 50 μm or greater.
In the invention thus constituted, the average diameter of the bubbles mixed into the fermentation vessel liquid is 50 μm or greater, therefore the cover body can suppress penetration of bubbles while allowing passage of crystals produced in the liquid with an average particle size of 5μ or greater. Stable readings can thus be obtained by the sensor. Note that average bubble diameter in this Specification denotes the mean value of the weighted chord length (square-weighted cord length) obtained by measurement using focused beam reflectometry (FBRM), corresponding to what is known as the volumetric average value.
In the present invention the fermentation vessel preferably has a ventilation tube for feeding gas into the fermentation vessel, with a hole diameter at the outlet of the ventilation tube of 1 μm or greater.
In the invention thus constituted, the hole diameter at the outlet of the ventilation tube for feeding gases into the fermentation vessel is 1 μm or greater, thus increasing the average diameter of bubbles mixed into in the fermentation vessel liquid. The cover body can therefore suppress penetration by bubbles while allowing crystals produced in the liquid with an average particle size of 5μ or greater to pass through. Stable readings can thus be obtained by the sensor.
In the present invention the fermentation vessel preferably has a ventilation tube for feeding gas into the fermentation vessel, and the volume of gas fed into the fermentation vessel per hour through this ventilation tube is less than or equal to twice the culture medium volume at the start of fermentation in the fermentation vessel.
In the invention thus constituted, the volume of gas fed into the fermentation vessel per hour through the ventilation tube is less than or equal to twice the volume of the culture liquid at the start of fermentation in the fermentation vessel, therefore gas can be made to dissolve into the liquid without overly fragmenting the gas bubbles fed into the fermentation vessel. As a result, penetration of bubbles into the cover body can be easily suppressed.
In the present invention the cover body is preferably formed in an approximately cylindrical shape, with the sensor extending in an axial direction therein.
In the invention thus constituted, the cover body is formed in an approximately cylindrical shape, therefore bubbles floating up from the bottom of the cover body can easily flow upward along the bottom surface of the cover body, further suppressing penetration by bubbles into the cover body. Since the cover body is formed in an approximately cylindrical shape, bubbles penetrating into the cover body and floating to the surface are collected in the highest part of the cover body, where they are less likely to contact the sensor. This enables adverse effects on measurement to be minimized even if bubbles do penetrate into the cover body.
In the present invention, preferably, the bottom permeable portion is disposed on the entire surface of the lower semicircular portion of the approximately semi-cylindrical cover body, and the top permeable portion is disposed on the entire surface of the upper semicircular portion of the cover body.
In the invention thus constituted, bottom and top permeable portions are respectively disposed over the entire surfaces of the lower and upper semicircular portions, therefore the portion of the cover body through which liquid and crystals pass can be made extremely large. Crystals which have entered into the cover body can as a result be easily discharged to the outside, and accumulation of crystals inside the cover body can be suppressed.
In the present invention the cover body is preferably formed from a thin sheet of metal, and micropores disposed in the bottom and top permeable portions are approximately circular holes formed in the thin sheet of metal.
In the invention thus constituted, micropores in the bottom and top permeable portions are formed by holes disposed in a thin metal plate, therefore compared to forming the bottom and top permeable portions with a mesh made by weaving strands, crystals are less likely to adhere to and deposit on the bottom and top permeable portions, and maintainability of the sensor cover can be improved. By forming micropores of the bottom and top permeable portions using holes in a thin metal plate, the cover body can be constituted to be less susceptible to damage and more durable than a mesh object.
In the present invention the diameter of micropores in the top permeable portion is preferably one to five times the diameter of the micropores in the bottom permeable portion.
In the invention thus constituted, selection of a diameter for the micropores in the top permeable part of one to five times the diameter of the micropores of the bottom permeable part enables an appropriate balance to be struck between suppressing the penetration of bubbles into the cover body and discharging bubbles that do end up penetrating into the cover, so that the effect of bubbles on the sensor can be effectively suppressed.
In the present invention the diameter of micropores disposed in the bottom permeable portion is preferably between 540 μm and 750 μm.
In the invention thus constituted, the diameter of micropores disposed in the bottom permeable portion is between 540 μm and 750 μm, therefore penetration of bubbles large enough to easily adversely affect sensor readings can be effectively suppressed, while liquid and crystals are allowed to flow into the cover body.
In the present invention, the fermentation product produced by the fermentation operation of the fermentation vessel is preferably cysteine, and oxidation of at least part of the cysteine results in the accumulation of cysteine in the fermentation vessel.
In the invention thus constituted, in the fermentation vessel, cysteine is produced as fermentation products, and cysteine is formed by the oxidation of at least part of the cysteine. Therefore, in the case the diameter of micropores disposed in the bottom permeable portion is between 540 μm and 750 μm, inflow and outflow of the crystalized cysteine and cysteine in the liquid through the cover body are allowed, while the entry of bubbles into cover body is effectively suppressed, the cysteine concentration can be accurately measured by the sensor.
In the present invention, the cover body is preferably disposed to project diagonally downward from the sidewall surface of the fermentation vessel containing the liquid, and the measuring portion of the sensor is positioned near the tip of the cover body.
In the invention thus constituted, the cover body projects diagonally downward from the sidewall surface of the fermentation vessel containing the liquid, therefore bubbles floating up from below and reaching the cover body can easily flow upward along the bottom surface of the cover body, effectively suppressing their penetration into the cover body. Bubbles penetrating into the cover body collect at the base portion of the cover body positioned above, thus moving away from the measurement portion of the sensor positioned near the tip of the cover body, so that negative effects on measurement can be reduced.
The invention is a sensor device for measuring properties of a liquid in a fermentation vessel into which bubbles are mixed, and crystals with an average particle size of 5 μm or larger are formed in a liquid, having a sensor for measuring liquid properties, and a sensor cover body disposed to surround the sensor, whereby a bottom permeable portion for passing the liquid and at least a portion of the crystals in the liquid is disposed on at least a portion of the bottom of the cover body, and a top permeable portion for passing the liquid and at least a portion of the crystals in the liquid is disposed on at least a portion of the top of the cover body, multiple micropores for passing liquid are respectively formed on the bottom permeable portion and the top permeable portion, and the micropores disposed on the top permeable portion are the same or larger than the micropores disposed on the bottom permeable portion.
According to the method for manufacturing fermentation products and the sensor device used for same of the present invention, a simple structure is used to suppress the effect of bubbles mixed into the liquid on sensor readings and to prevent crystals formed in the liquid from accumulating near the sensor or in the sensor cover, so that properties of the liquid and solids in the fermentation vessel can be measured, and the fermentation operation can be performed based on those measurements.
Below, referring to the attached figures, we explain preferred embodiments of the invention.
As shown in
In the present embodiment a sparger 8, being a ventilation tube, is provided on the bottom portion of fermentation vessel 2 to aerobically culture the liquid in fermentation vessel 2, and oxygen or air is introduced from supply device 8a through sparger 8 into fermentation vessel 2. Therefore fine bubbles of supplied oxygen or air or fine bubbles of carbon dioxide gas produced by the microorganisms being cultured are mixed into the culture liquid L. In the present embodiment the diameter of the hole at the outlet of gas-discharging sparger 8 is approximately 6 mm, which results in bubbles with an average diameter of 250 μm being mixed into the culture liquid in fermentation vessel 2. The hole diameter for discharging gas in the sparger 8, etc. is set at approximately 5 μm to approximately 36 μm, and bubbles with an average diameter of approximately 150 μm to approximately 350 μm are mixed into the culture liquid in fermentation vessel 2. Alternatively, a FAD (Fine Air Diffuser) can be used as an aerating tube to mix bubbles into the culture liquid. The FAD has a large number of holes ranging from approximately 1 μm to approximately 20 μm in diameter; this enables bubbles with an average diameter of approximately 50 μm to approximately 250 μm to be mixed into the culture liquid.
Referring next to
As shown in
Sensor cover 10 has an approximately cylindrical cover body 14 and a flange portion 16, disposed at the base end of this cover body 14 and affixed to the side wall surface 2a of fermentation vessel 2.
Cover body 14 is formed of a thin sheet of stainless steel with an approximately cylindrical shape, closed at the end, and is disposed to surround sensor 12. As described below, numerous micropores are disposed over the entire surface of cover body 14, and the liquid and at least some crystals in fermentation vessel 2 can pass through these micropores to flow into the cover body 14. Note that in addition to a cylinder, the cover body can also be constituted in any desired shape, such as a rectangle. The cover body can also be made of metals other than stainless steel, or resins such as polytetrafluoroethylene; it is best made of a material not easily damaged by the flow of liquid caused by stirring.
Flange portion 16 is a stainless steel disk, to the center portion of which a cover body 14 is welded to form an integrated single unit. Cover body 14 is mounted perpendicular to the flat surface of flange portion 16. Bolt holes 16a are disposed on flange portion 16, and flange portion 16 is affixed to the outside of sidewall surface 2a by affixing bolts 18a. In addition, a packing 18b is disposed between flange portion 16 and side wall surface 2a, assuring watertightness between flange portion 16 and sidewall surface 2a. Furthermore, as shown in
In the present embodiment a transmission-reflectance type near-infrared spectroscopic sensor (NIR sensor) is adopted as sensor 12 to measure the concentration of amino acids in liquid L by near infrared analysis. However, in addition to NIR sensors, the sensor cover 10 in the present embodiment can be applied to various sensors, such as spectroscopic sensors using ultraviolet or visible light, other optical sensors, sensors using focused beam reflectometry (FBRM) to measure crystal particle size, and electromagnetic sensors to measure dielectric constant or electrical conductivity, and can be combined with these sensors to constitute a sensor device.
As shown in
Next, referring again to
As described above, numerous micropores are formed over the entire surface of cover body 14 of a sensor device 1 according to an embodiment of the present invention, and the surrounding liquid as well as at least some crystals can flow into the interior of cover body 14. Here, in sensor device 1 of the present embodiment, the size of each micropore formed on the top side of cover body 14 should be set to be equal to or larger than the size of each micropore formed on the bottom side thereof. I.e., a bottom permeable portion 14a which allows liquid to pass through is formed on the bottom surface of cover body 14, and a top permeable portion 14b which allows liquid to pass through is formed on the top surface thereof, whereby the micropores formed in top permeable portion 14b are formed to be the same size or larger than the micropores formed in bottom permeable portion 14a.
As shown in
In the present Specification, the cover body 14 “bottom surface” means the side illuminated when light is projected vertically downward from sensor cover 10 installed in a usage state, and “top surface” means the side illuminated when light is projected in the vertically upward direction therefrom. In the present embodiment numerous micropores are formed over the entire “bottom surface” and “top surface” of cover body 14, but micropores do not necessarily have to be formed over the entire surface; it is sufficient that they be formed in a portion thereof. Furthermore, in the present embodiment micropores are also formed on the tip surface 14c of cover body 14, but it is also acceptable not to form micropores on tip surface 14c.
Next, as shown in
Here, bottom permeable portion 14a and top permeable portion 14b, formed by arraying circular holes with a diameter D=approximately 750 μm at a pitch P=approximately 1070 μm results in an opening area of approximately 44.4%, roughly corresponding to a #20 mesh (#20 mesh: a mesh in which 20 strands per inch are longitudinally and transversely disposed). In the present Specification, “micropore size” in the bottom permeable portion 14a and top permeable portion 14b refers to the circular hole diameter.
Note that if circular holes with a diameter D=approximately 540 μm are arrayed at a pitch P=approximately 770 μm, the open area ratio is about 44.5%, roughly corresponding to a #30 mesh. In addition, circular holes of diameter D=approximately 350 μm arrayed at a pitch P=approximately 630 μm would result in an aperture ratio of approximately 28%, roughly corresponding to a #40 mesh. Circular holes of diameter D=approximately 250 μm arrayed at a pitch P=approximately 360 μm would result in an aperture ratio of approximately 43.6%, roughly corresponding to a #60 mesh. Circular holes of diameter D=approximately 180 μm arrayed at a pitch P=approximately 320 μm would result in an aperture ratio of approximately 28.7%, roughly corresponding to a #80 mesh. Circular holes of diameter D=approximately 150 μm arrayed at a pitch P=approximately 250 μm would result in an aperture ratio of approximately 32.5%, roughly corresponding to a #100 mesh, and circular holes of diameter D=approximately 100 μm arrayed at a pitch P=approximately 210 μm would result in an aperture ratio of approximately 20.5%, roughly corresponding to a #150 mesh.
In the present embodiment the bottom permeable portion 14a and top permeable portion 14b are constituted by forming numerous micropores in a thin plate, but a bottom permeable portion 14a or top permeable portion 14b can also be constituted from a mesh-shaped object formed by combining fine strands of wire, such as by weaving, knitting, or the like. In such cases, “micropores” are formed as the spaces between strands constituting the mesh object, and “micropore size” indicates the distance between adjacent strands. The diameter of micropores formed in bottom permeable portion 14a is preferably set to between approximately 180 μm and approximately 750 μm. More preferably, the diameter of micropores in bottom permeable portion 14a is set to between approximately 250 μm and approximately 750 μm. Even more preferably, the diameter of micropores in bottom permeable portion 14a is set to between approximately 350 μm and approximately 750 μm. Even more preferably, the diameter of micropores in bottom permeable portion 14a is set to between approximately 540 μm and approximately 750 μm. I.e., it is preferable to set the diameter of micropores in bottom permeable portion 14a to a size that suppresses the penetration of bubbles into culture liquid L and also allows at least some of the crystals of the culture liquid L to pass through. The bottom permeable portion 14a and top permeable portion 14b, in which numerous micropores are formed in a thin plate, reduce adherence and deposition of crystals, and improve maintainability of the sensor cover.
To facilitate the discharge of bubbles which have entered into the cover body, it is preferable to form the micropores disposed in top permeable portion 14b in a size larger than the micropores disposed in bottom permeable portion 14a. However, the diameter of micropores in top permeable portion 14b should be set to a size fully capable of suppressing the inflow of bubbles through top permeable portion 14b caused by the downward flow of liquid. The diameter of the micropores formed in top permeable portion 14b is preferably about 1 to 5 times the diameter of micropores formed in bottom permeable portion 14a. This allows for an appropriate balance between the suppression of bubbles penetrating into cover body 14 and the discharge of bubbles which do penetrate into cover body 14.
Next, referring to
First, it has been known for some time that when measuring the properties of a liquid into which bubbles are mixed, the effect of the bubbles can be reduced by covering the sensor with a mesh-shaped strainer. However, as shown in
On the other hand, as shown in
More preferably, as shown in
On the other hand, small bubbles approaching from below pass through bottom permeable portion 14a and penetrate into cover body 14. However, by forming large-diameter micropores in top permeable area 14b, small bubbles passing through bottom permeable portion 14a and penetrating will float up inside cover body 14 and can be easily discharged to the outside through top permeable portion 14b. Penetrating bubbles can therefore be prevented from accumulating for long periods inside cover body 14 and growing there into large bubbles.
In addition, while not all bubbles approaching sensor cover 10 approach from the bottom side, a buoyancy force from the liquid is always acting on the bubbles, therefore the fraction of bubbles approaching sensor cover 10 from below is high, and their penetration is greatly suppressed by bottom permeable portion 14a. Moreover, top permeable portion 14b more effectively stops the penetration of some bubbles approaching sensor cover 10 from above or from the side, etc. as the result of liquid flow caused by stirring in the fermentation vessel than if the top portion of the cover were completely open. Furthermore, forming larger micropores in top permeable portion 14b allows crystals C to more easily penetrate into the interior of sensor cover 10 so that the crystal component can be more accurately detected.
By disposing cover body 14 so that it projects diagonally downward, bubbles that have penetrated into cover body 14 will move upward toward the base portion of cover body 14. On the other hand, measurement portion 12b on sensor probe 12a is disposed close to the tip of cover body 14, therefore bubbles inside cover body 14 are moved away from sensor 12 measurement portion 12b. This enables the effect on measurement of bubbles entering into cover body 14 to be even further reduced.
Using sensor cover 10 of the present embodiment, these actions effectively reduce the influence of bubbles on measurement.
Next, referring to
First, in step S1 of
Next, in step S2, liquid to be fermented in fermentation vessel 2 is introduced into fermentation vessel 2. In the example shown here, cysteine is produced as the fermentation product by fermenting the culture liquid L introduced into fermentation vessel 2; at least a portion of this cysteine is oxidized in the culture liquid to produce cysteine (Cys2). As an example, culture liquid L contains a carbon source, a nitrogen source, a sulfur source, and inorganic ions.
Sugars such as glucose, fructose, sucrose, molasses, and starch hydrolysates, and organic acids such as fumaric acid, citric acid, succinic acid, and the like can be used as carbon sources.
Inorganic ammonium salts such as ammonium sulfate, ammonium chloride, and ammonium phosphate, organic nitrogen such as soybean hydrolysate, ammonia gas, ammonia water, and the like can be used as nitrogen sources.
Sulfur sources can include inorganic sulfur compounds such as sulfates, sulfites, sulfides, hypo sulfites, thiosulfates, and the like.
It is desirable to include appropriate amounts of required substances such as vitamin B1 or yeast extract as an organic micronutrient source. In addition to these, small amounts of potassium phosphate, magnesium sulfate, iron ions, manganese ions, and the like may be added as needed.
In step S3, fermentation vessel 2, into which culture liquid L is introduced, is operated for fermentation, and properties of the liquid in fermentation vessel 2 are measured by sensor device 1. During the fermentation operation, the culture liquid L in the fermentation vessel 2 is agitated by operating an agitator 6, and by operating a feeder 8a; bubbles are mixed into culture liquid L via sparger 8. In the present embodiment the average diameter of bubbles mixed into culture liquid L by sparger 8 is approximately 250 μm. This volume introduced per hour of this air corresponds to about 0.1 to 1.2 times the volume of the medium in fermentation vessel 2 at the start of fermentation. The volume of air fed into fermentation vessel 2 per hour is preferably less than or equal to 1 to 1.2 times the volume of the medium at the start of fermentation in fermentation vessel 2, so that air can be dissolved in the liquid without excessive fragmentation of the bubbles.
In addition, fermentation operating conditions are adjusted based on measurement results from sensor device 1. In the present embodiment, cysteine is produced by the fermentation operation; as the amount of cysteine dissolved in culture liquid L increases, it precipitates to form cysteine crystals. Properties of culture liquid L are measured from time to time during the fermentation operation by sensor device 1. In the present embodiment, the properties of culture liquid L measured by sensor device 1 include cysteine concentration [g/L], sugar concentration [g/L] (R.S.), ammonia nitrogen concentration (AN), S2O3 concentration [g/L], turbidity (OD) of culture liquid L, and the like.
In addition, fermentation operating conditions are adjusted based on the properties of culture liquid L measured by sensor device 1. Conditions of the fermentation operation to be adjusted include the culture liquid L temperature, the culture liquid L pH, the amount of aeration fed through sparger 8, the amount of sugar solution added, the amount of each component such as phosphorus added, and the like. Adjustments to fermentation operating conditions based on these measured properties of culture liquid L are performed as needed during the fermentation operation.
We now explain the measurement of cysteine concentration by sensor device 1.
As noted above, sensor device 1 comprises a sensor 12 (
Also, when compared to the
I.e., this is believed to be due to the fact that for a fermentation operation in which cysteine is produced in a culture liquid L in fermentation vessel 2, the micropores are too fine in a sensor cover with micropores equivalent to a #60 mesh, so that crystals precipitated from the liquid phase cannot be discharged from the cover, resulting in crystal precipitation and deposition. In contrast, in the
In the examples shown in
Next, referring to
As shown in
First, as shown in
Next, when sensor covers 10 with the #20 mesh equivalent of condition (2) and the #20 mesh equivalent of condition (3) were used, no adhesion was found on sensor 12 or the outer surface of sensor cover 10, nor was any crystal accumulation inside sensor cover 10 found. There was also no significant fluctuation in estimated cysteine concentrations, and data could be stably measured. I.e., when sensor cover 10 is set to a #20 mesh equivalent or a #30 mesh equivalent, the micropore size is sufficiently larger than the average particle size of cysteine crystals, and cysteine crystals can sufficiently penetrate into sensor cover 10. Highly reliable estimates of cysteine concentration by detection of crystals penetrating into sensor cover 10 using sensor 12 can thus be obtained, and crystals that do penetrate into sensor cover 10 can be easily discharged out of sensor cover 10. On the other hand, most of the bubbles in culture liquid L are prevented from penetrating into sensor cover 10, and the adverse effects of bubbles on sensor 12 are suppressed.
Next, when a sensor cover 10 with a #40 mesh equivalent was used in condition (4), no adhesion of crystals to the outer surface of sensor cover 10 was found, but a small accumulation of crystals was found on the inside of sensor cover 10. Estimated cysteine concentrations did not fluctuate significantly during the experimental period, and data could be stably measured. Under these conditions, however, there is a risk that when a fermentation operation is conducted for an extended period, crystal accumulation inside sensor cover 10 may increase, such that proper readings cannot be acquired. When the micropores disposed in sensor cover 10 are small, the penetration of bubbles into sensor cover 10 can in this way be more strongly suppressed, which is advantageous for eliminating the adverse effects of bubbles. On the other hand when micropores are small, cysteine crystals which have entered sensor cover 10 and crystals formed inside the cover are difficult to discharge to the outside, and are more prone to accumulate inside sensor cover 10.
Furthermore, when a condition (5) #60 mesh equivalent sensor cover 10 was used, adhesion of crystals to the outer surface of sensor 12 and sensor cover 10, and a large accumulation of crystals inside sensor cover 10, were found. Also, it became impossible to obtain proper data during the experimental period due to the effects of crystals accumulated in sensor cover 10. Effectiveness in suppressing the penetration of bubbles is thus strengthened when a sensor cover 10 with even finer micropores than condition (4) is used, but the amount of cysteine crystals accumulating inside sensor cover 10 increases dramatically. When a large quantity of cysteine crystals accumulates in sensor cover 10 in this way, cysteine concentration inside sensor cover 10 rises dramatically, making it difficult to accurately measure the cysteine concentration in the culture liquid L inside fermentation vessel 2.
Finally, no adhesion of crystals to the outside surface of sensor 12 or sensor cover 10 was found when an #80 mesh equivalent was used for bottom permeable portion 14a and a #40 mesh equivalent was used for top permeable portion 14b of sensor cover 10 under condition (6), but a small accumulation of crystals was found inside sensor cover 10. The estimated cysteine concentrations did not fluctuate significantly during the experimental period, and data could be stably measured. Under these conditions, however, there is a risk that when the fermentation operation is conducted for an extended period, crystal accumulation inside sensor cover 10 may increase, such that proper readings cannot be acquired. Also, a better result was obtained in condition (6) than in condition (5), even though micropores in bottom permeable portion 14a were set at an #80 mesh equivalent, which is finer than in condition (5). I.e., it is believed that because the micropores in top permeable portion 14b are larger in condition (6) than in condition (5), crystal accumulation inside sensor cover 10 is less likely to occur. Therefore a sensor cover 10 with which data can be stably measured over a long time period, while also further reducing the adverse effects of bubbles can, by fine tuning the size of the micropores in the sensor cover 10 under condition (6), be constructed using a bottom permeable portion 14a with fine micropores.
From the results shown in
According to the method of producing fermentation products of the present embodiment of the invention, a sensor device 1 for measuring properties of culture liquid L, which is the liquid in fermentation vessel 2, has a cover body 14 disposed to surround sensor 12. A bottom permeable portion 14a and top permeable portion 14b which pass culture liquid L and at least part of the crystals in culture liquid L are disposed on cover body 14, therefore the culture liquid L to be measured can continuously flow into and out of cover body 14. As a result, the culture liquid L in cover body 14 is constantly replaced, and the sensor 12 disposed inside cover body 14 can measure properties of the culture liquid L containing crystals continuously and in real time.
In addition, because bottom permeable portion 14a and top permeable portion 14b of cover body 14 pass at least a portion of crystals in culture liquid L, those crystals which form in culture liquid L are less likely to accumulate in cover body 14, and the fermentation operation can be continued for a long without the need for maintenance such as cleaning the cover body 14. The micropores formed in top permeable portion 14b are larger than those formed in bottom permeable portion 14a, therefore bubbles which float up from the bottom of cover 14 are less prone to pass through the micropores in bottom permeable part 14a, and penetration of bubbles into cover body 14 can be effectively suppressed. On the other hand the micropores in top permeable portion 14b are formed to be the same or larger than those in bottom permeable portion 14a, therefore bubbles which penetrate into cover body 14 can easily float up in cover body 14 to be discharged through top permeable portion 14b. As a result, bubbles penetrating cover body 14 accumulate inside cover body 14, so their adverse effects on sensor 12 readings can be suppressed.
According to the fermentation product manufacturing method of the present embodiment, the average diameter of bubbles mixed into culture liquid L in fermentation vessel 2 is 250 μm, therefore cover body 14 can suppress the penetration of bubbles while also allowing the passage of crystals produced in the liquid with an average particle size of 5 μm or greater. Stable measurements can thus be obtained by sensor 12.
In addition, according to the fermentation product manufacturing method of the present embodiment, the hole diameter at the outlet of the ventilation tube for feeding gas into fermentation vessel 2 is 6 mm, therefore the average diameter of bubbles mixed into culture liquid L in fermentation vessel 2 increases. Therefore cover body 14 suppresses the penetration of bubbles while allowing crystals produced in culture liquid L with an average particle size of 5 μm or greater to pass through. Stable measurements can thus be obtained by sensor 12.
According to the fermentation product manufacturing method of the present embodiment, the volume per hour of air, which is the gas being fed into fermentation vessel 2 through sparger 8, a ventilation tube, is between 0.1 to 1.2 times the culture medium volume at the start of fermentation, therefore the air can be dissolved into culture liquid L without excessive fragmentation of the air bubbles fed into fermentation vessel 2. As a result, bubbles can be easily suppressed from penetrating into cover body 14.
According to the fermentation product manufacturing method of the present embodiment, cover body 14 is furthermore formed in an approximately cylindrical shape (
According to the fermentation product manufacturing method of the present embodiment, a bottom permeable portion 14a and top permeable portion 14a are respectively located over the entire surfaces of the lower and upper semicircular portions, therefore the portion of cover body 14 through which the culture liquid L and crystals can permeate can be made extremely large. As a result, crystals which have penetrated into cover body 14 can be easily discharged to the outside, and accumulation of crystals in the cover body 14 can be suppressed.
Furthermore, according to the method for manufacturing fermentation products of the present embodiment, micropores are formed in bottom permeable portion 14a and top permeable portion 14b by the holes provided in a thin metal plate (
In the method for manufacturing fermentation products of the present embodiment, when the diameter of micropores in top permeable portion 14b is larger than the diameter of micropores in bottom permeable portion 14a, the discharge of crystals and bubbles within the cover body can be promoted, while the entry of bubbles into cover body 14 is effectively suppressed, therefore the influence of accumulated crystals and bubbles on sensor 12 can also be effectively suppressed.
Furthermore, according to the method for manufacturing fermentation products of the present embodiment, the diameter of micropores formed on bottom permeable portion 14a is from 180 μm to 750 μm, therefore the penetration of large bubbles which are prone to adversely affect sensor 12 readings can be effectively suppressed, while the inflow of liquid and crystals into cover body 14 is allowed.
According to the fermentation product manufacturing method of the present invention, cover body 14 projects diagonally downward (
We have explained preferred embodiments of the present invention above, however various changes may be applied to the above-described embodiments. In particular, in the above-described embodiment of the invention, cysteine was produced as the fermentation product in a culture liquid L into which bubbles were mixed, but the invention may be applied to the manufacture of any other desired fermentation product in which crystals are produced in a liquid into which bubbles are mixed. For example, the invention may be applied to fermentation vessels where amino acids, nucleic acids, or peptides, which have relatively low solubility and from which crystals can easily precipitate, are produced in a culture liquid L in which fermentation products, fermentation carbon sources, nitrogen sources, phosphorus sources, oxygen sources, and/or sulfur sources are dissolved.
Here fermentation products include, for example, amino acids; fermentation carbon sources include, for example, sugars, monosaccharides, disaccharides, and polysaccharides; nitrogen sources include, for example, ammonia and ammonium sulfate; phosphorus sources include, for example, phosphate ions; and sulfur sources include, for example, thiosulfate ions. Examples of amino acids, nucleic acids, and peptides whose crystals are relatively easy to precipitate during fermentation include glutamic acid (Glu), tryptophan (Trp), phenylalanine (Phe), threonine (Thr), tyrosine (Tyr), cysteine (Cys), cysteine (Cys2), glutamine (Gln), aspartic acid (Asp), leucine (Leu), isoleucine valine (Val), inosine, guanosine, adenine, and others.
In the embodiments described above, measurement was made of the concentration of components precipitated in the liquid, but it is also possible to measure the properties of the liquid itself, such as its dielectric constant or electrical conductivity, or to measure bacteria in the liquid.
Furthermore, in the above-described embodiment of the invention, a single sensor device was installed in fermentation vessel 2 to measure various properties of the liquid, but it is also possible to install multiple sensor devices 1 in fermentation vessel 2. In such cases, different covers for sensors may be applied to each sensor device according to the characteristic measured by each sensor device. For example, one sensor device can measure properties of precipitated crystals, while another sensor measures properties of components dissolved in the liquid phase.
| Number | Date | Country | Kind |
|---|---|---|---|
| 2019-233846 | Dec 2019 | JP | national |
| Number | Date | Country | |
|---|---|---|---|
| Parent | PCT/JP2020/044859 | Dec 2020 | US |
| Child | 17807998 | US |
