JPH0743366B2 - Kjeldahl nitrogen analyzer - Google Patents

Description

The present invention relates to a Kjeldahl nitrogen determination method and device, and more particularly to a Kjeldahl nitrogen determination method and device in which substantially all of the operations and steps required for measurement are labor-saving and automated. The automatic measuring method and apparatus of the present invention can be applied very effectively even in a small amount or a minute amount measurement such as the semi-micro Kjeldahl method or the micro Kjeldahl method.

Kjeldahl's nitrogen determination method was published by Kjeldahl in 1883, and has been more than 100 years old.However, it is still extremely useful as a method for determining various nitrogen-containing substances, especially nitrogen or protein in animal and plant samples. is important. For example, the Biuret method and various other methods are known as nitrogen determination methods, but the accuracy of these other methods is determined with the Kjeldahl method as the standard, and the nitrogen determination method in the Japanese Pharmacopoeia is semi-micro. The Kjeldahl method is adopted, and the protein nitrogen determination method in the biological standard specifies the micro Kjeldahl method.

As is well known, the principle of the Gerdal method is that an organic sample containing substances is decomposed by heating with sulfuric acid in the presence of a catalyst to convert nitrogen in the sample into ammonium sulfate, and a strong alkali such as sodium hydroxide is allowed to act on this to release free ammonia. Generate
The amount of ammonia is quantified, and depending on the amount of sample used, the macro Kjeldahl method (nitrogen content: 0.5 to 1 g),
It is classified into three types: the semi-micro Kjeldahl method (nitrogen content: 2-3 mg) and the micro-Kjeldahl method (nitrogen content: 10-1000 μg). Although these methods are simple in principle, the actual operation requires many steps, which are complicated and require an extremely large heat source.

(1) Preparation and weighing of sample and each reagent (catalyst, sulfuric acid, NaOH solution, indicator, collection liquid) (2) Introduction of sample into Kjeldahl flask (3) Addition of copper sulfate and other suitable catalyst (on the flask wall surface) It is necessary to add it carefully so that it does not adhere to it, or to pour the adhered catalyst with water, etc.) (4) Add sulfuric acid little by little (5) Pyrolyze the contents (6) Cool and dilute the contents (if the flask is (Be careful not to break it.) (7) Addition of NaOH solution (8) Distillation of generated ammonia by steam (9) Absorption by ammonia trapping solution (for example, boric acid solution) (10) Indicator for ammonia absorption solution Addition of (11) Titration with sulfuric acid (12) Recording of measured values (13) Treatment of waste liquid (14) Cleaning of equipment such as flasks and beakers Therefore, if all of these steps are performed manually, It takes about 40 to 60 minutes for one sample and 20 hours for about 50 samples, which is a method strongly demanded for labor saving and automation. However, since it is too complicated, it is not known to the present application that the automated measurement method applicable to a small-quantity or micro-quantification method such as the semi-micro or micro-Kjeldahl method has been reported so far. There are only a few reported examples of automation that are only applicable in.

For example, "Food Industry, Vol.18, No.6, p.73 (1975)" was published in the June 1974 issue of the West German GORDIAN magazine by rev nat.Alfred Montag (Hamburg, National Institute of Chemical Foods). The article entitled "Quantitative determination of nitrogen by Kjeldahl method using KJEL-FOSS-AUTOMATIC apparatus" was introduced, and an automated method and apparatus for nitrogen measurement in the Macro-Kjeldahl method were disclosed. According to this, the automated measurement method consists of using a combination device consisting of a structure and a matrix as shown in FIG. 1 and FIG. 2 of the above-mentioned document, which device has a turn installed on a central waste liquid tank. From its center to the table
Six flask mounting positions are provided at an angle of 60 °, and the turntable, that is, the flask is automatically rotated every 3 minutes by a motor with a moving time of 10 to 12 seconds by 60 °, and a sample introduction position and two positions are provided. Heat decomposition position by direct fire, cooling,
It is designed so as to be sequentially circulated and fed to the dilution position, the ammonia generation distillation position, the waste liquid treatment position, and the original sample introduction position. At the top of the flask, at the ammonia generation distillation position and the waste liquid treatment position for discarding the contents, a gasket structure with a rubber stopper can be achieved so that the contents of the flask can be sealed. , Waste liquid treatment, etc. are automatically performed at each position. Further, the ammonia distilled at the ammonia generation distillation position is collected through a condenser in a beaker placed in an automatic measuring system, where the ammonia is directly titrated by sulfuric acid which is automatically discharged from the titration cylinder. The titration monitors the color change of the indicator consisting of methyl red and methylene blue with a phototube. The so-called code monitor method is used, and the titration value is obtained by knowing the movement amount of the cylinder that pushes out H ₂ SO ₄ for titration directly through a potentiometer and controlling it through a digital amplifier, and then automatically recording it as a digital display. .

However, as is apparent from the above-mentioned literature, this conventional method relates to the macro Kjeldahl method for ham, sausage, fish meat and other foods, and cannot be applied to the semi-micro or micro Kjeldahl method.
Because the intense heat such as open flame in the decomposition process used in this conventional method is a semi-micro or micro Kjeldahl that usually treats a small amount of liquid sample (solution or suspension form) in a further diluted form. This is because bumping is inevitable and cannot be used in the method. Moreover, although the above-mentioned conventional method is automated after the decomposition step, the sample and other introduction steps, that is, the addition of the sample to be measured, the catalyst and the acid are all performed manually. That is, in the above-mentioned conventional method, the weighed sample and the tableting catalyst are first put in a flask, and then sulfuric acid and hydrogen peroxide whose amounts are adjusted by a manual dial operation are added from an introduction pipe. In addition, heating with an open flame may cause damage to the decomposition flask unless careful operation is performed.

According to the present invention, there is provided a Kjeldahl nitrogen determination method and apparatus which can be applied to a semi-micro or micro Kjeldahl method and further automatically add a sample, a catalyst and sulfuric acid to a decomposition flask. In the present invention, there is provided a measuring device that does not require tableting of the catalyst and does not risk the breakage of the flask.

The present invention is achieved by additionally combining an automatic liquid transfer device, an automatic catalyst addition device, an automatic sulfuric acid addition device, and a constant temperature automatic hot air heating device as described below with a conventional turntable-type decomposition flask rotation device. It

Hereinafter, the present invention and its features will be described more specifically with reference to the accompanying drawings.

FIG. 1 is a main part 1 of the Kjeldahl nitrogen measuring device of the present invention.
FIG. 3 is a schematic plan view showing two preferable arrangements, in which a plurality of sample containers 2 containing pre-adjusted samples to be measured are placed on a series of mounting tables 3 and repeatedly moved and stopped at regular intervals. Sample container transfer device 1 for continuous circulation transfer; a plurality of, preferably, 12 decomposition flasks 11 are held at equal intervals, and the decomposition flasks 11 are sampled by rotating in the direction of the arrow corresponding to the movement / stop of the sample container 2. Introduction stop position p ₁ , catalyst addition stop position p ₂ , sulfuric acid addition stop position p ₃ , heat decomposition stop position p ₄
~ P ₉ , contents cooling / dilution stop position p ₁₀ ammonia generation distillation stop position p ₁₁ and waste liquid removal / flask cleaning position p ₁₂ ,
To be measured from the sample container 2 at the top corresponding stop position p ₁ sample container transfer device 1 'in addition sequentially circulated digestion flask turntable 10 for rotating movement, p ₁ position of the original to the sample introduction position p ₁ Liquid sample automatic transfer device 12 for transferring sample to decomposition flask 11, automatic catalyst addition device 13 for extracting a certain amount of powdery or granular catalyst at the p ₂ position and adding it to the decomposition flask 1, decomposition flask at the p ₃ position
Sulfuric acid automatic addition device for adding a fixed amount of sulfuric acid to 11, 14, p ₄ ~
A series of hot-air heating devices for heating the flask 11 by sending hot air under the decomposition flask 11 at the position of p ₉ through the blower pipe 19.
Cooling water addition device for cooling and diluting the contents of the flask 11 after pre-cooling with an air-cooling device (see Fig. 10) from the bottom at the 15 and p ₁ positions 16 and steam distillation at the p ₁₁ position to generate ammonia. Of the Kjeldahl nitrogen decomposing / distilling process section comprising an ammonia generating / distilling device 17 to be sent to the titration system and a waste liquid removing / cleaning device 18 for cleaning the inside of the flask by suctioning and removing the waste liquid in the decomposition flask 11 at the p ₁₂ position; Place the beaker 22 for titration on the mounting table 28 and rotate it in the direction of the arrow to move the beaker to the ammonia collection position Q ₁ , titration position Q ₂ , titrated liquid removal / washing position Q corresponding to the p ₁₁ position of the turntable 10. ₃ , ..., Titration cycler 20, which is sequentially rotated to the ammonia scavenger supply position Q ₆ , an automatic titrator having electrodes 26, 26 ′ and a titrant introduction pipe 27 held by a holder 25 at the Q ₂ position. 2 1, Q ₃ position ammonia absorption liquid supply device 24 in the titration-liquid removed beaker cleaning device 23 and Q ₆ position to clean the beaker interior is aspirated titration-liquid in (The supply of the collecting liquid Q ₄ Or
It can also be done at the position of Q ₅ ).

According to this arrangement of the present invention, not only the macro Kjeldahl method, but also the semi-micro and micro Kjeldahl method can be used to determine nitrogen automatically and continuously with great advantage. Hereinafter, each device constituting the present invention and its operation will be described in detail in the order of steps.

FIG. 2 shows an example of a sample container transfer device 1 used in the present invention, which comprises a sample storage conveyor 5 for moving a sample container 2 containing a sample to be measured by repeatedly rotating and stopping in the direction of the arrow. ing. Sample container transfer device 1
Is a rotating disk or other suitable means adapted to the space, and the conveyor 5 is rotated by a motor (not shown), and is repeatedly moved and stopped at an appropriate time interval according to the operation time in the subsequent disassembling step. Is set. In particular, the stop time is determined by the time required in the heating step and the ammonia generation / distillation step, as will be described later, though it depends on the sample amount, the catalyst amount, and the thermal decomposition temperature. The travel time is arbitrary, and several seconds is usually sufficient. In this way, when the sample container 2 carried by the conveyor 5 in accordance with the movement / stop of the turntable 10 comes to the p ₁ ′ position, that is, the position corresponding to the sample introduction position p ₁ of the turntable 11, The contained sample is transferred to the decomposition flask 11 at the p ₁ position by the following specific liquid sample transfer device 12. Preferred is p ₁ ′
The sample container 2 is actually placed immediately before the position, and the sample container detector 4 for detecting whether or not the sample container 2 is actually installed is installed in the conveyor 5. As shown in the figure, the sample container detector 4 is composed of a simple microswitch structure 6 and detects it when the sample container is idled or when it is no longer necessary to send the sample container after the test is completed. Therefore, it is set so that the operation in each step after the sample introduction can be stopped accordingly. In particular, in the case of idle feeding, the mechanism is such that each operation of only the flask of idle feeding can be stopped.

In the present invention, the sample specimen to be contained in the sample container is a solution or fine suspension in the form of a solution-containing white nitrogen sample to be measured in advance. For example, when quantifying the protein content of a specimen sample according to biologic standards,
The specimen sample is pretreated by the following operations.

That is, the sample should be diluted if necessary and the amount of protein nitrogen should be 10%.
Accurately take an amount corresponding to ˜500 μg, put it in a centrifugal precipitation tube, add 1/10 volume of 50 W / V% trichloroacetic acid solution, then heat to 100 ° C. for 15 minutes, and then cool to room temperature. However, for antitoxin, therapeutic serum and blood products, heat at 100 ° C for 15 minutes instead of spraying at room temperature to 37 ° C.
Incubate at ℃ for 15 minutes. Then, centrifuge at about 800g for 10 minutes. Add an appropriate amount of 5 W / V% trichloroacetic acid solution to the precipitate, shake, and centrifuge again. Dissolve the precipitate with a small amount of IN sodium hydroxide reagent. Alternatively, instead of sodium hydroxide, concentrated sulfuric acid such as 24N sulfuric acid may be used to dissolve the precipitate of the trichloroacetic acid solution, and copper sulfate and other catalysts may also be present in the sample unless they interfere with the next transfer. You may let me. However, in this case, no addition of catalyst at the p ₂ position and / or addition of sulfuric acid to the decomposition flask at the p ₃ position is required. The pretreatment can be carried out directly in the sample container 2.

The specimen sample in the sample container 2 thus prepared is the third
The liquid sample automatic transfer device 12 shown in the figure can automatically transfer the liquid sample into the decomposition flask 11.

FIG. 3 is an overall side view of the automatic sample transfer device 12 used in the present invention, showing a state immediately before sample transfer of samples. That is, the device 12 includes a shaft 51 vertically movable up and down and arms 55, 55 '.
And holders 56 and 56 'respectively hold the p ₁ ' position and the p ₁ position on the p 1'position, and are composed of a sample suction portion 57 and a sample feeding portion 58 which can be respectively moved up and down by the vertical movement of the shaft 51.

The liquid suction part 57 extends from the liquid suction pipe 52 that reaches the bottom of the sample container 2 when it descends, and the cleaning liquid spraying device 53 having a ring water nozzle structure which is mounted on the liquid suction pipe 52 and sprays the cleaning liquid toward the inner wall of the sample container 2. And the liquid feeding part 58 has a shaft 51.
When the temperature drops, that is, in the state shown in Fig. 3, the decomposition flask
It has a means 60, preferably ground glass means, which fits into the open inner wall 61 of the 11 and seals the inside of the flask.

The slicing means 60 comprises a glass member 59 having an open lower end and a hollow upper part.
It is equipped with a decompression exhaust port that allows decompression inside. The liquid suction pipe 52 and the liquid feed pipe 62 are connected by a liquid transfer pipe 54, and their respective tips are all moved when the sample container 2 and the decomposition flask 11 are moved, that is, when the shaft 51 moves up.
It is located above the container 2 and the flask 11, respectively.

In such an arrangement, the liquid transfer device 12 repeats vertical movements by, for example, a linear head motor (not shown) in accordance with the movement / stop of the conveyor 1 and the rotary table 10, and the device 12 descends to cause the decomposition flask 11 to descend. At the same time as the closed state, the inside of the flask is decompressed by a decompression device (not shown), and the sample specimen from the sample container 2 moves into the flask through the liquid suction pipe 52, the liquid transfer pipe 54, and the liquid feed pipe 62. In addition, at the same time when the liquid sample is inhaled, the cleaning liquid injection device
The washing water is sprayed from 53 onto the inner wall of the sample container 2, and the washing water is transferred to the decomposition flask 11 in the same manner as soon as the adhered sample on the wall surface of the container is washed off. At the same time as these operations are completed, the device 12 is raised to a position where it does not hinder the movement of the sample container 2 and the decomposition flask 11. These series of operations,
That is, the vertical movement of the transfer device 12, the decompression operation in the decomposition flask, and the injection of the washing water are adjusted by moving and stopping the sample container 2 and the decomposition flask 11 by a sequence controller or the like set in advance as described later. Automatically operated.

Next, when the decomposition flask 11 is moved to the p ₂ position, that is, the catalyst addition position, a certain amount of powdered or granular solid catalyst from the automatic catalyst addition device 13, for example, CuSo ₄ · K ₂ So ₄ powder mixture. (Mixing ratio 1: 9) is added into the flask. Incidentally, the catalyst is not required to be added at this position as long as it is added to the preliminarily prepared sample, as described above.

FIG. 4 is an overall side view of the automatic catalyst addition device 13 used in the present invention, and FIG. 4-a shows a state in which the catalyst A is filled in a space of a constant volume formed in the outlet passage. Show, 4th
FIG. b shows a state in which a certain amount of the catalyst filled in the space is being extracted. Further, FIG. 5 is a side view of the same catalyst addition apparatus as seen from above in the state of FIG. 4-a.

That is, as shown in the figure, the catalyst addition device used in the present invention contains a solid catalyst A to be quantitatively added to the decomposition flask 11, and a container having a catalyst A drop-in opening 72 at the bottom.
71; A discharge path 73 for the catalyst A from the opening 72, and a pair of guide paths 75, 7 for the following barrier plates that extend horizontally across the discharge path.
A block body 74 attached to a container 71 having 5 '; a reciprocating motion in the guide paths 75, 75' to block the lead-out path 73 to form a fixed volume quantitative space 76 in the guide path and stagger With holes A and 78 'for dropping catalyst A at position 1
When the hole 78 coincides with the lead-out passage 73, the cut-off plate 77 'closes the lead-out passage 73 and drops the catalyst A into the space 76 to fill the empty space 76. Then, when the hole 78 coincides with the outlet path 73, the shield plate 77 closes the outlet path 73 to drop a certain amount of the catalyst A accumulated in the space 76; And a device 90 for reciprocating the 77, 77 'at regular time intervals. The device 90 for reciprocating the shields 77, 77 'can be any device known in the art suitable for the purpose, but is preferably eccentric as shown.
An eccentric disc 91 that rotates at 93 and an eccentric disc that engages 1
It consists of a combination with a block body 95 having a pair of pulleys 92, 92 ', so that a simple and compact reciprocating device can be provided.

More specifically, the container 71 generally has an opening 72 for guiding the stored catalyst A smoothly to the opening 72 without accumulating.
It has the shape of an inverted cone or an inverted truncated cone from which the casing 81 is preferably integrated. The casing 81 is intended to support and protect the container 71, and has a peripheral protruding portion 84 at its bottom peripheral edge which can be screwed or nut-fixed 85 with a suitable support (not shown).

A block body 74 having a catalyst lead-out path 73 aligned with the opening 72 is attached to the bottom of the container 71 by an appropriate means, and this lead-out path plays an important role in the intended continuous catalyst withdrawal. That is, in the block body 74, a pair of horizontal guide paths that enable the reciprocating motion of the pair of blocking plates 77, 77 'so that a constant volume space 76 can be formed in the block body 74 across the lead-out path 73. There are 75 and 75 '. Thus, a space 76 having a constant volume determined by the distance d between the shield plates 77 and 77 'and the cross-sectional dimension 1 of the lead-out path 73 is formed inside the lead-out path 73. Therefore, holes 78, 78 'having a shape that matches the cross-sectional shape of the lead-out path 73 are provided in the shielding plates 77, 77' at alternate positions, and when the amount of the catalyst A is determined, the hole 78 of the cut-off plate 77 'is set in the lead-out path 73. The shield plate 77 'closes the lead-out path 73, and the shield plate 77 closes the lead-out path 73 at the time of extraction so that the holes 78' of the shield plate 77 'are aligned with the lead-out path 73. By reciprocating at time intervals, a fixed weight of catalyst, which is determined according to the density of the catalyst to be quantified, can be quantitatively and continuously introduced into the decomposition flask 11. Below the outlet path 73, a tapered outlet 86 is attached so that the sample is prevented from scattering or the quantified sample easily falls into the decomposition flask 11.

The block body 74 is made of a suitable material such as metal or plastic by using a suitable mold or the like, and the lead-out path 73 and the blocking plates 77, 7 are provided.
It is possible to use a block body which is integrally formed from the beginning so as to have the guide passages 75, 75 'for 7', but it is preferable to use the two members 100 and 100 having the shape shown in Fig. 6-a. It can be advantageously obtained by combining with one of the members 110 shown in Fig. 6-b. That is, with the member 110 in the middle and the two members 100 facing each other, the respective holes 111 and 10
If you assemble with screw holes 112, 102, etc. so that 1 matches, the hole
A block body 74 having a lead-out path 73 formed by 111 and 101 and a pair of blocking plate guide paths 75, 75 'formed by two recesses 103 is constructed. The holes 111 and 101 have the same size and shape, and their size and the thickness of the member 110 depend on the quantity to be quantified. The shape and dimensions of the recess 103 of the member 100 are such that the shield plates 77,
The shape and size of the shield plate are slightly larger than those of the shield plate so that the 7's can slide smoothly and reciprocate, but the shield plate and the guide passage should be prevented so that the solid catalyst is not guided along the guide passage due to the reciprocating motion of the shield plate. It is important that they fit together and have as little space between them as possible. Even by doing so, the sample derived from such a gap cannot be completely avoided. However, the amount is extremely small and does not affect the amount to be measured, that is, the amount to be added to the decomposition flask. If necessary, access positions 79, 7 of the shield plates 77, 77 'to the block body 74
A catalyst recovery receiver (not shown) may be placed under 9 '(see Fig. 4-b).

The device 90 for the reciprocating movement of the shield plates 77, 77 'may be any device known in the art suitable for the purpose, but it is preferred that the device 90 is rotated by an eccentric disc 91 which rotates about an eccentric 93 as shown. Is to do. For example, the device 90 used in the present invention, as shown in FIGS.
7'is fixed by a screw 98 or the like and a block body 97 and another block body 95 connected or integrated with this by a connecting plate 96, and the block body 95 has pulleys 92 at both ends thereof.
And 92 ', and an eccentric disc 91 that rotates about an eccentric 93 provided so as to engage with both pulleys between them to shield the shield plate 7.
7,77 'repeats reciprocating motion. Of course, the pulley and the eccentric plate may be directly attached to the block body 97. When the eccentric disc 91 is rotated by a motor (not shown) in the direction of the arrow by such an arrangement, the shield plates 77, 77 'are rotated every half turn.
Reciprocates and repeats the states of FIGS. 4-α and 4-b, and the amount of catalyst accumulated in the space 76 in the state of FIG. 4-a is equal to that of 4-b. When the state shown in the figure is reached, it flows down into the decomposition flask 11 placed under the outlet path 73. As described above, the flask 11 is sent under the discharge path 73 by the turntable 10 at regular time intervals, but the drop of the catalyst from the quantification space 76 in FIG. Therefore, while the next flask 11 is fed under the outlet channel 73, the shut-off plates 77, 77 'are set at the fourth position.
When placed in the state shown in FIG. a, sufficient dropping of the catalyst into the space 76 is achieved. Axis 99 and 99 'are block bodies 95 and 97
It is provided to stabilize and smooth the reciprocating motion of the.

The operation of the shield plates 77, 77 'is appropriately adjusted by an appropriate control means such as a sequence controller according to the feeding of the decomposition flask 11.

Further, the container 71 containing the catalyst to be quantified is usually covered with a lid 82.
More preferably, a desiccant 83 (for example, silica gel) contained in a non-woven fabric or other bag-like material is attached to the lower part of the lid, or is attached in an embedded form to prevent the catalyst inside the container from absorbing moisture.

Further, the p ₃ position after catalyst addition, if necessary, that is, in the case of using sodium hydroxide as described above for the preparation of the sample to be measured, obtain automatically performed the addition of the required amount of sulfuric acid by the device 14 . The device 14 may be a so-called general piston type dispenser, as shown in FIG. 7 which is an overall side view of the device 14, and is placed in a liquid in a supply container 121 of concentrated sulfuric acid to be added, for example, 36 NH ₂ SO _4. Extended suction pipe 122 and liquid transfer pipe 123
It is composed of a T-shaped tube 120 connecting with and. The T-tube 120 is connected to a piston means 125 by a branch tube 124 extending from its branch, which piston means is a syringe type cylinder 126.
And its inner pusher 127, the pusher 127 is in contact with the disc 128 at its lower end, and the eccentricity 129 of this disc 128 is
The structure is such that eccentric rotation about the axis makes sliding contact in the cylinder to move up and down for a certain distance. Also,
As shown in FIG. 8, check valves 133 and 133 'are provided in the T-shaped pipe 120 upstream and downstream of the branch portion, respectively.
As a result, a certain amount of sulfuric acid when the pusher 127 of the piston means moves up and down due to the eccentric rotation of the disk 128, the transfer pipe 123
It can be sent to the decomposition flask 11 via. That is,
When the pusher 127 descends, the check valve 133 closes and the check valve 133 ′ opens, and a certain amount of sulfuric acid corresponding to the distance from the supply container 121 down the pusher 127 flows into the cylinder 126, and then the disk 128 is formed. When the pusher 127 starts to move up by rotating, the valve 133 ′ is closed and the valve 133 is opened, and the cylinder 12 is opened.
A certain amount of sulfuric acid corresponding to the amount flowing into 6 is sent to the transfer pipe 123. The sulfuric acid thus sent to the transfer pipe 123 in a constant amount is dropped into the decomposition flask 11 from the transfer pipe tip 132, but at this time, the transfer pipe tip 132 prevents the scattering of the sulfuric acid falling from it. Therefore, it is moved up and down by a vertical shaft 130 that can move up and down so as to extend to the inside of the decomposition flask 11, and after dropping further, liquid that may remain attached to the tip 132 does not fall outside the decomposition flask, or for the purpose of recovery, the tip is used. It is preferable to set so that 132 is located on the collecting funnel 134 during standby. Tip 132
Is held on the vertical shaft 130 by the arm 131, and the collecting funnel 13
The collection pipe 135 is connected to the supply container 121.

In this way, the decomposition flask 11 to which the required amount of sulfuric acid has been added at the p ₃ position is then sequentially moved to the heating positions p _{4 to} p ₉ to thermally decompose the contents. That is, the time required for heating in the Kjeldahl nitrogen determination method depends on the sample amount to be measured, the heating temperature, the catalyst amount, the sulfuric acid amount, and many other factors. Alternatively, in the Micro Kjeldahl method, a temperature of about 340 to 380 ° C. for several minutes to about 7 to 8 minutes is sufficient. For example, sample volume 3-5 mg (nitrogen content 10-100
0 μg) for a Kjeldahl method measurement sample of approximately K ₂ S
When about 3 g of O ₄ / CuSO ₄ catalyst and 2 ml of 36N sulfuric acid are added, the heating temperature and the required time are about 360 ° C ± 10 ° C for about 8 minutes,
Ten minutes is more complete and 12 minutes is more complete. Therefore, in this example showing an example of nitrogen measurement by the Micro Kjeldahl method, if the stop time at each position of the decomposition flask is set to 2 minutes, 6 stop heating positions, that is, p _{4 to} p ₉ heating stop positions. This makes it possible to achieve a complete decomposition process. This heating can also be performed by direct flame with a gas burner, etc., but in order to avoid the risk of breakage of the decomposition flask, which is usually made of glass, or the bumping of the contents, the following hot air heating with an air heater is performed. Is preferred.

FIG. 9 shows an example of the air heater 15 that can be advantageously used in the Kjeldahl nitrogen measuring device of the present invention.
from p ₄ ~p ₉ bottom of the electric heater 140 and the cylindrical body 141 of the tubular body 141 housed in a housing 142 provided directly below the respective decomposition flask 11 into a position between the compressor 144. linked with blower tube 147 Become. That is, for example, in the cylindrical body 141
An electric heater 140 of 200V, 1KW is installed, which is heated to a constant temperature by an electric circuit having a variable resistor 143.
Compressor 144 to cock 145, flow meter 146
By sending pressurized air through the blower pipe 147, a fixed amount of hot air can be blown to the bottom of the decomposition flask 11, and the contents of the flask can be heated to a predetermined temperature. Since the temperature of the electric heater can be kept constant by the variable resistor 143 and the amount of air blown from the compressor 144 can be kept constant by the flow meter 146, a constant amount of heat can be always given to the bottom of the flask. Further, an exhaust duct 148 is provided above the decomposition flask, and exhaust gas generated during thermal decomposition is processed up to a safety standard by an exhaust gas processing system (not shown) and released into the atmosphere.

Thus p ₄ ~p digestion flask 11 having been subjected to heat treatment for a predetermined time at ₉ position moves to the next cooling and dilution position p _10, where the temperature of the first contents subjected to pre-cooling by air cooling of the contents is about 100 to 150 When the temperature drops to ℃, the contents, especially the sulfuric acid content, are diluted with cold water and rapidly cooled at the same time. By air-cooling the contents beforehand, bumping of the contents and damage to the flask due to rapid cooling are prevented.

FIG. 10 shows an example of the cooling / diluting device 16 for the contents of the decomposition flask of the present invention at the p ₁₀ position, and this device 16 is used in combination with the cold air blowing fan 150. After the heat treatment, the decomposition flask 11 which has been moved to the position p ₁₀ is air-cooled by the fan means 150 installed directly below the decomposition flask 11. The fan means 150 has a fan 151 driven by a motor means (not shown) and a blower port 152, and the cold air is blown from the blower port to the bottom surface of the flask so that the contents of the flask are blown for a predetermined time (usually about 1 to about 1). 100 ℃ ~ 150 ℃ for 30 seconds)
It is set according to the feeding of the flask that can be cooled to. At the same time, the vertical moving means 153 similar to the vertically movable vertical shaft 51 in FIG. 3 lowers the gas suction pipe 154 and the cooling water injection nozzle 157 so that the gas suction pipe 154 is placed in the flask and the cooling water injection nozzle 157. The devices 16 are operated so that they are positioned right above the flask mouth. That is, the gas suction pipe 154 sucks the gas component and hot air generated during the air cooling and the subsequent cooling by the jet water and discharges the hot air to the outside.

Simultaneously with the end of air cooling, the injection nozzle 157 injects the cooling / diluting water sent from a supply source (not shown) through the connecting pipe 156 into the flask, so that the content is about 50-60%.
Cool to ℃ and dilute. The injection nozzle is an annular nozzle in the illustrated embodiment, but may have any shape as long as the object is achieved. After that, the injection nozzle 157 and the gas suction pipe 154 are automatically moved to the flask 11 by the vertical movement means 153.
And the cooling process is completed. Note that the injection nozzle
157 may be fixed on the mouth of the preliminary flask without interlocking with the suction pipe 154.

After completion of the cooling step, the flask 11 moves to the next distillation step, that is, the position p ₁₁ according to a predetermined transfer cycle of the turntable 10, where ammonia is replaced (generated) by a strong alkali and ammonia distillation is performed by steam. The ammonia thus formed is sent to the next measurement cycle.

FIG. 11 shows an example of the ammonia generation / distillation device 17 used in the Kjeldahl nitrogen quantification device of the present invention, which comprises a distillation device 170 and a condenser 210 mounted on a vertically movable vertical shaft 160. Vertical axis 160 is shown in FIGS.
It has exactly the same function as the vertical shafts 51 and 153 in the figure, and is set so as to repeat vertical movement according to the movement / stop of the decomposition flask 11.

The distillation apparatus 170 is preferably a double tube structure made of glass, the bottom of which is the flask 11 when the vertical axis 160 is lowered.
For example, it has a sliding portion 178 which is tightly joined to the neck portion of the flask and seals the inside of the flask. In such a structure, when the vertical axis descends and the inside of the flask is sealed, the distillation device 170
Water vapor is fed into the space 173 between the outer wall 171 and the inner wall 172 from the feed pipe 196. The supplied steam fills the space 173 and heats the internal immediate distillation region 177 of the inner wall 172, and is sent to the contents in the flask 11 through the pipe 174, the solenoid valve 175 and the pipe 176. At the same time, a strong alkali, for example 7.5N NaOH solution, from source 183 is passed through supply line 186 to pipe 17
It is sent to 6 and added to the contents of the flask together with steam. The strong alkali supply mechanism can be exactly the same as the sulfuric acid feed mechanism shown in FIG. 7, and a predetermined amount of NaOH can be efficiently supplied into the flask 11 from the supply pipe 183 in the same manner as described with reference to FIG. That is, as in FIG. 7, the quantity T is adjusted by the piston means 184 and has a check valve.
A fixed amount of NaOH is sent from the inlet tube 186 into the flask by the tube 185. Further, in the figure, reference numeral 179 is a drain for condensed water generated in the space 173.

Thus, the NaOH fed into the decomposition flask 11 acts with the ammonium sulfate generated in the contents of the flask to generate free ammonia, which is distilled together with the steam fed at the same time as a fraction passing through the distillation region 177. It is sent to the condenser 210 through the branch pipe 180, and from this condenser, Q _{1 of} the titration cycle corresponding to the p ₁₁ position.
It is collected in the collecting liquid in the beaker 22 at the position, that is, the ammonia collecting position, as described later. Distillation equipment branch pipe
The end of 180 has, for example, a sliding structure 181 so that it can be sealed with the neck of the condenser. Further, the distillation apparatus 170 may have a structure as shown in FIG.

As a steam supply device for supplying steam to the distillation device 170, an automatic steam generator 190 as shown in FIG.
The present invention can be advantageously practiced by using That is,
The steam generator 190 used in the present invention is an electrode heater 192.
A container 191 having a water inside is heated by a heater 192 to fill water up to a constant water level L, and the generated steam can be sent to a distillation apparatus 170 through a feed pipe 196. Further, the container 191 has a pipe 193 for makeup water (distilled water) and a drain port 195, and this pipe has a solenoid valve 194. 197 is a thermocouple, 198 is a thermometer, and 199 is a variable resistor for adjusting the temperature of the heater. The container 191 is supported by a ring 206 and a clamp 207, and is supported by a fulcrum 201 so as to balance with the weight 202, that is, when the water in the container is at a constant water level L. 203 is a detection end of a proximity switch that detects the proximity of the weight 202, 204 is a safety device that detects the descent of the weight, and 205 and 205 'are stoppers.

In such an arrangement, in the steam generator 190, the filling water decreases and the water level falls below the water level L as the steam is generated by starting the heater 192 and the generated steam is fed to the distillation device 170 through the pipe 196. At the same time as the water level starts to descend, the weight 202 starts to descend, and the detection end 203 detects the proximity of the weight.
Since this detection end 203 is set so as to send a signal to the electromagnetic valve 194 in the makeup water pipe 193, the solenoid valve 194 opens and the makeup water is supplied from the pipe 193 into the container. Then, when the water in the container 191 reaches a predetermined water level, that is, the water level L, the weight rises and the proximity signal of the detection end 203 is cut off, so the solenoid valve 194 is closed. That is, the fluctuation of the water level is converted to ON / OFF of the proximity switch via the balance, and this is connected to the opening / closing signal of the solenoid valve, so that the water level can be finely adjusted and the container 191 can always maintain a constant water level. Since a sharp inclination of the balance is not desirable, stoppers 205, 205 'for controlling the inclination are provided. Further steam generator 190
The water level is lowered due to an unexpected accident such as water interruption of operation, and the safety device 204 detects and stops the device entirely. This is to further protect the device. Also, the temperature control of the heating element can be freely reached by the variable resistor.

Since the steam feed pipe 196 is connected to the distillation device 170 that moves up and down, a flexible pipe or tube is usually used. Thus, the supply of steam to the ammonia distillation step of the present invention can be carried out extremely safely and efficiently.

After the distillation process, the decomposition flask 11 is
_{Moved to} position _{12 for} content disposal and flask wash.

The waste liquid treatment apparatus 18 of FIG. 13 is set to be vertically movable by a linear motor (not shown) or the like like the vertical shaft 51 of FIG. 3, the vertical shaft 153 of FIG. 10 or the vertical shaft 160 of FIG. A liquid suction pipe 224 held by an arm 222 of a vertical shaft 221 and a holder 223, a liquid transfer pipe 225 connected to or integrated with the liquid suction pipe 224, and a cleaning liquid jetting device 226.

That is, when the decomposition flask 11 stops at the position p ₁₂ , the tip of the suction pipe 224 descends to the bottom of the flask, and at the same time, the inside of the waste liquid tank 228 is decompressed from the pipe 229 by a vacuum pump (not shown) and the waste liquid is discharged. Is sucked from the suction pipe tip port through the liquid transfer pipe 225 connected to the suction pipe and moves into the waste liquid tank 228. At the same time, the cleaning water is sprayed from the cleaning water spraying device 226 to the inner wall surface of the flask through the cleaning water supply pipe 227, and this sprayed cleaning water is also sucked by the liquid suction pipe 225 and moved to the waste liquid tank similarly to the steam waste liquid.

The cleaning water spraying device 226 cleans the inner wall surface of the disassembling flask 11 uniformly so as not to interfere with the preparation of the next measurement liquid, and in general, it is a liquid suction pipe so that it comes directly above the flask opening when spraying. 224 or any other suitable location, preferably using an annular nozzle structure with nozzle holes facing the inner wall of the flask. However, other shapes may be used as long as the object of the present invention is achieved.

As described above, the decomposition flask 11 which has completed the respective operations by sequentially setting the p ₁ to p ₁₂ positions returns to the p ₁ position again, and the same operation is repeated thereafter, and a series of sample treatments in the Kjeldahl nitrogen determination is performed. Can be carried out continuously and automatically.

In particular, operating position of the 12 described with respect to FIG. 1-FIG. 13, i.e., if p ₁ ~p ₁₂ operating time at the position (stop time) was set in each 2 minute interval, as described above, the sample digestion The heating time for the sample is sufficient, and the processing time per sample is 24 minutes. However, it becomes possible to continuously and automatically process a large number of samples at intervals of 2 minutes thereafter. In particular, it is quite surprising that this continuous operation could be achieved in the Micro Kjeldahl method, which requires a strict and complicated operation.

Then, titration of the ammonia generated in the distillation step at position p ₁₁ can also be entirely automated according to the invention. That is, the eleventh
As shown in the figure, the ammonia from the condenser 210 is collected in the beaker 22 on the Q ₁ position of the titration cycler 20, and the titration operation is automatically repeated as follows.

First, the titration cycler 20 and the automatic titration device 21 constituting the titration structure of the present invention shown in FIG. 1 may be a commercially available automatic cycle titration device, preferably an automatic titration recording device. For example, FIG. 14 illustrates an essential part of an automatic cycle titrator with an automatic recording device that can be advantageously used in the present invention. That is, the automatic titrator shown at 21 in FIG. 1 has a PH electrode 26, a titration sulfuric acid dropping pipe 235, a holder 25 for holding the dropping pipe and the electrode, and a vertical movement according to the movement / stop of the titration cycler 20. Equipment for making 23
6, also including an annular washer 241, an Auburet 231, a titration sulfuric acid source storage container 232, a titration control unit 237, and a printer 238, which are also attached to the device 236 for electrode washing, and a titration sulfuric acid source 232 and an auto view. Lett 231, the auto burette and the drip pipe 235 are pipe 233, respectively.
And 234 are connected. Further, a magnetic stirrer device 239 is provided below the beaker stop position for titration of the titration cycler 20, that is, the Q ₂ position, and the magnet 240 in the beaker is rotated to stir the liquid to obtain a more uniform measurement value. To Here, it should be noted that the stop / transfer cycle of the titration cycler 20 needs to be adjusted to match the move / stop cycle of the decomposition flask turntable 10.

Hereinafter, the measurement step of the present invention will be described from the operation from Q ₁ in which ammonia is collected from the p ₁₁ position of the decomposition step, and the beaker 22 at the Q ₁ position is a suitable position (Q _{4 to} Q ₆ ) in front of it. A predetermined amount of introduced ammonia scavenging liquid, for example, 3% boric acid liquid, which will be described later, is included. The ammonia-containing distillate from the condenser 210 is collected by the collected liquid in this beaker, and at that time, as shown in FIG. 11, the level detection sensors 211, 211 'for detecting the liquid level in the beaker 22 are used. It is preferable to put.

This sensor can estimate the amount of ammonia distillate from the sample to be measured within a certain range depending on the sample amount of the sample group, that is, the micro, semi-micro, or macro Kjeldahl method. The final liquid level in the beaker that increases due to the collection of minutes is sensed, thereby controlling the distillation process and making the automatic operation of the present invention more complete.

In this way, the beaker 22 that has collected all of the ammonia to be measured moves to the next Q ₂ position, that is, the titration position according to the movement cycle of the titration cycler 20, and the titration is performed by the automatic titrator 21 shown in FIG. 14. It is performed on an automatic drop and its measurement is automatically recorded by an automatic recording device, i.e. printer 238.

Beaker finished titration subsequently Q ₃ position, i.e. moved to a beaker cleaning position, removal and beaker wash droplet Teisumi liquid is performed. As the device 23 for removing the waste liquid and cleaning the beaker, the same device as the waste liquid removing / decomposing flask cleaning device 18 in FIG. 13 can be used, and therefore the details thereof will be omitted.

It suffices to simply change the position or shape of the cleaning liquid injection annular nozzle in the device 18 of FIG. 13 so as to be suitable for beaker cleaning.

Thus beaker finishing the waste removal and cleaning the next Q ₄ ~Q ₆
It moves sequentially to the position, and at any of the positions (the other positions of Q _{4 to} Q ₆ are play positions in this example), the required amount of ammonia trapping liquid, for example, 3% boric acid liquid is supplied. Since the supply of the collected liquid can also be performed by using the device for adding the sulfuric acid or alkali liquid illustrated in FIGS. 7, 8 and 11 and the device which is completely used, the details of the mechanism are described. Is omitted. FIG. 15 is only a schematic diagram thereof.

As described above, a series of devices from the sample preparation to the quantification of the Kjeldahl nitrogen determination method according to the present invention, the waste liquid treatment, and the instrument cleaning and the operation thereof have been described. These operations, that is, the decomposition flask turntable and the titration cycler Rotation / stopping; vertical movement at each position; addition / injection operation of various reagents, cooling water, cleaning liquid, etc. at each position; depressurization operation at p ₁ , p ₁₀ , p ₁₂ , Q ₃ positions; p _{5 to} p ₉ Lighting of heater at position;
………… etc. A series of all operations in the method and apparatus of the present invention are automatically controlled by a preset sequence controller or the like with respect to the sequence, timing, time and the like. A normal control method can be applied to the control method, and thus the detailed description will not be given.

Therefore, according to the present invention, it is possible to automate all the operations in the Gerdall nitrogen determination method except for the preparation (pretreatment) of the measurement sample, and especially in the micro or macro Kjeldahl method which has conventionally been done only manually. Can be achieved.

In particular, the use of the sample container detector (see FIG. 1) provided on the sample sledge conveyor and the level detector (see FIG. 11) at the time of ammonia collection, which is a preferred embodiment of the present invention, eliminates erroneous operation of this device. Safe and accurate measurement operation can be achieved. Moreover, in the past, even a skilled person required about 40 to 60 minutes per sample and about 20 hours even before and after 50 samples, but according to the present invention, the stop time of the turntable and the titration cycler is as described above. If the (operating time of each step) is set to 2 minutes, the time required for titration per sample is about 28 minutes, and the subsequent samples can be continuously processed at about 2 minute intervals. It is possible to process 26 specimens per hour. Therefore, the labor saving in the Kjeldahl nitrogen determination method according to the present invention is immeasurable. Moreover, as will be described below, it is possible to obtain an accurate measured value exactly at the same time as when it is manually performed by a skilled person.

Comparative Example For various protein samples, the automatic measurement according to the present invention and the conventional manual measurement of the amount of contained protein nitrogen were compared.

Each sample was pretreated according to the standard of biological preparation, that is, treated with trichloroacetic acid, centrifuged, and dissolved with 1N NaOH solution. The measurement is performed 10 times for each sample, and the result is an average value thereof.

Further, since the pretreatment of the sample was dissolution with NaOH, the automatic measurement according to the present invention included the sulfuric acid automatic addition step at the p ₃ position. The results are as follows, and no difference was found in the variance and average between the method according to the present invention and the conventional method.

[Brief description of drawings]

FIG. 1 is an overall schematic plan view showing an example of the Kjeldahl nitrogen automatic measuring device of the present invention. FIG. 2 is a perspective view showing an example of the sample container transfer device used in the Kjeldahl nitrogen automatic measuring device of the present invention. FIG. 3 is an overall side view of the sample transfer device used in the Kjeldahl nitrogen automatic measuring device of the present invention during operation. FIG. 4 shows an example of the catalyst addition device used in the Kjeldahl nitrogen automatic measuring device of the present invention. FIG. 5 is a plan view of the device shown in FIG. 4 as seen from above. FIG. 6 is an exploded view of a block body 74 which is a main part of the apparatus shown in FIG. FIG. 7 shows an example of the sulfuric acid addition device used in the Kjeldahl nitrogen automatic measuring device of the present invention. FIG. 8 is a detailed view of a cylinder dispenser which is a main part of the apparatus shown in FIG. FIG. 9 shows an example of a heating device used in the Kjeldahl nitrogen determination device of the present invention. FIG. 10 shows an example of a cooling / diluting device used in the Kjeldahl nitrogen determination device of the present invention. FIG. 11 is an overall side view of the ammonia distillation apparatus used in the Kjeldahl nitrogen determination apparatus of the present invention. FIG. 12 shows another example of a distillation tube which can be used in the distillation apparatus shown in FIG. FIG. 13 is an example of a waste liquid treatment / equipment cleaning device used in the Kjeldahl nitrogen determination device of the present invention. FIG. 14 shows an example of an automatic titrator assembly used in the Kjeldahl nitrogen determination device of the present invention. FIG. 15 shows one embodiment of the ammonia scavenging liquid supply device of the Kjeldahl nitrogen determination device of the present invention. 1 ... Sample container transfer device, 10 ... turntable 11 ... decomposition flask, 12 ... sample transfer device 13 ... catalyst addition device, 14 ... sulfuric acid addition device 15 ... heating device, 16 ... cooling Diluter 17 …… Distiller, 18 …… Waste liquid treatment / flask washing device 20 …… Titration cycler, 21 …… Automatic titration device 22 …… Beaker, 23 …… Waste liquid treatment / beaker washing device 24 …… Collection liquid supply Device 170 ... Steam generator

Claims (4)

[Claims] 1. A plurality of decomposition flasks are installed, and these decomposition flasks are repeatedly stopped and moved to a sample introduction position, a plurality of thermal decomposition positions, a cooling / diluting position, an ammonia distillation position and a waste liquid removing / washing position. While rotating and moving, the decomposition flask rotating table that was able to repeatedly achieve the predetermined steps when stopped at each position and the ammonia from the ammonia distillation position were collected, and the titration was stopped and moved on the rotating table. In a continuous Kjeldahl nitrogen quantification device consisting of a combination with an automatic cycle titrator that automatically performs corresponding to, in accordance with the stop / movement of the decomposition flask rotary table,
A sample container transfer device that continuously feeds a plurality of sample containers containing protein nitrogen-containing preparations to be measured near the sample introduction position; and, in accordance with the stop of the rotary table, continuously sent near the sample introduction position. A device for transferring the prepared sample from the stopped sample container to the decomposition flask at the sample introduction position, (a) a sample on a sample container that is held by a vertically movable shaft and can move up and down at the same time. The sample suction part and the sample feeding part on the decomposition flask are as follows: (b) The sample suction part and the sample feeding part respectively move the sample container and the decomposition flask according to the vertical movement of the shaft when the sample container and the decomposition flask are moved. It is located above the flask and is configured to descend at the same time when the sample container and the decomposition flask are stopped. Liquid suction pipe that reaches the bottom of the instrument, a cleaning liquid ejection device that is attached to this liquid suction pipe and that spouts the cleaning liquid toward the inner wall of the sample container immediately after the liquid sample is sucked by the liquid suction pipe, and an integrated or connected liquid suction pipe A liquid transfer pipe attached, which has a liquid transfer pipe extending to the inside of the decomposition flask through the sample feeding part that descends at the same time, (d) the sample feeding part is a sealing member for the decomposition flask, And a Kjeldahl nitrogen continuous including a continuous sample transfer device from the sample container to the decomposition flask, which comprises a decompression means for automatically decompressing the inside of the decomposition flask and automatically transferring the liquid and washing solution in the sample container to the decomposition flask. Quantitative device. 2. When the prepared sample does not contain a catalyst, a catalyst addition position is provided between the sample introduction position and the thermal decomposition position, and the powdery or granular catalyst to be added is accommodated at the position, and the catalyst is added to the bottom. A container having an opening for dropping the catalyst; a catalyst discharge path from the opening and a pair of guide paths for the following barrier plates extending in parallel across the discharge path, and attached to the catalyst container A block body; a pair of parallel parallel plates each having a catalyst dropping hole at alternate positions by reciprocating in each of the above-mentioned guide passages to block the lead-out passages to form a fixed volume in the guide passages. In the case of the blocking plate, when the hole of the upper blocking plate coincides with the discharge path, the lower blocking plate closes the discharge path and drops the catalyst into the space to fill the space, and the hole of the lower blocking plate is When the lead-out paths match And a pair of the shielding plates for closing a certain amount of the catalyst so as to drop a certain amount of the catalyst in the space; and a reciprocating motion of the pair of the shielding plates at a constant time interval in accordance with the stop / movement of the rotary table. The Kjeldahl nitrogen linked quantitative apparatus according to claim (1), further comprising a continuous quantitative addition apparatus to the decomposition flask of the catalyst, which comprises a stirring apparatus. 3. When the prepared sample does not contain sulfuric acid, a sulfuric acid addition position is provided between the sample introduction position and the thermal decomposition position,
The Kjeldahl nitrogen according to claim (1), further comprising sulfuric acid addition comprising a piston type dispenser which operates at the stoppage and movement of the rotary table and adds a fixed amount of sulfuric acid to the decomposition flask at the position. Continuous quantitation device. 4. The continuous Kjeldahl nitrogen quantitative determination according to claim 1, wherein the heating device at each thermal decomposition position comprises a pressurized air heater comprising an electric heater provided at the bottom of the decomposition flask and a compressed air blowing means. apparatus.