JPS5811024B2 - How to measure ammonia in liquid - Google Patents
How to measure ammonia in liquidInfo
- Publication number
- JPS5811024B2 JPS5811024B2 JP9506579A JP9506579A JPS5811024B2 JP S5811024 B2 JPS5811024 B2 JP S5811024B2 JP 9506579 A JP9506579 A JP 9506579A JP 9506579 A JP9506579 A JP 9506579A JP S5811024 B2 JPS5811024 B2 JP S5811024B2
- Authority
- JP
- Japan
- Prior art keywords
- ammonia
- present
- water
- serum
- indophenol
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired
Links
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 title claims description 59
- 229910021529 ammonia Inorganic materials 0.000 title claims description 29
- 239000007788 liquid Substances 0.000 title claims description 10
- 238000000034 method Methods 0.000 claims description 51
- WTKZEGDFNFYCGP-UHFFFAOYSA-N Pyrazole Chemical compound C=1C=NNC=1 WTKZEGDFNFYCGP-UHFFFAOYSA-N 0.000 claims description 6
- 239000011734 sodium Substances 0.000 claims description 6
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 claims description 5
- 239000003054 catalyst Substances 0.000 claims description 5
- 229910052708 sodium Inorganic materials 0.000 claims description 5
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 20
- 210000002966 serum Anatomy 0.000 description 15
- PNNCWTXUWKENPE-UHFFFAOYSA-N [N].NC(N)=O Chemical compound [N].NC(N)=O PNNCWTXUWKENPE-UHFFFAOYSA-N 0.000 description 13
- RSAZYXZUJROYKR-UHFFFAOYSA-N indophenol Chemical compound C1=CC(O)=CC=C1N=C1C=CC(=O)C=C1 RSAZYXZUJROYKR-UHFFFAOYSA-N 0.000 description 10
- 239000003153 chemical reaction reagent Substances 0.000 description 8
- 238000005259 measurement Methods 0.000 description 8
- 239000000243 solution Substances 0.000 description 8
- 238000011088 calibration curve Methods 0.000 description 7
- 238000012360 testing method Methods 0.000 description 7
- 238000006243 chemical reaction Methods 0.000 description 6
- 238000011161 development Methods 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- 108010046334 Urease Proteins 0.000 description 5
- 238000002835 absorbance Methods 0.000 description 5
- 238000010521 absorption reaction Methods 0.000 description 5
- 239000007864 aqueous solution Substances 0.000 description 5
- 239000000126 substance Substances 0.000 description 4
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 3
- 239000004202 carbamide Substances 0.000 description 3
- 238000007796 conventional method Methods 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 239000012086 standard solution Substances 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 2
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- OJGMBLNIHDZDGS-UHFFFAOYSA-N N-Ethylaniline Chemical compound CCNC1=CC=CC=C1 OJGMBLNIHDZDGS-UHFFFAOYSA-N 0.000 description 2
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N Phenol Chemical compound OC1=CC=CC=C1 ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- ABBQHOQBGMUPJH-UHFFFAOYSA-M Sodium salicylate Chemical compound [Na+].OC1=CC=CC=C1C([O-])=O ABBQHOQBGMUPJH-UHFFFAOYSA-M 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 150000004982 aromatic amines Chemical class 0.000 description 2
- 239000013060 biological fluid Substances 0.000 description 2
- 239000000460 chlorine Substances 0.000 description 2
- 229910052801 chlorine Inorganic materials 0.000 description 2
- 238000005345 coagulation Methods 0.000 description 2
- 230000015271 coagulation Effects 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- -1 cyanide compound Chemical class 0.000 description 2
- 238000004821 distillation Methods 0.000 description 2
- 238000000691 measurement method Methods 0.000 description 2
- QKEOZZYXWAIQFO-UHFFFAOYSA-M mercury(1+);iodide Chemical compound [Hg]I QKEOZZYXWAIQFO-UHFFFAOYSA-M 0.000 description 2
- 231100000614 poison Toxicity 0.000 description 2
- 230000007096 poisonous effect Effects 0.000 description 2
- IOLCXVTUBQKXJR-UHFFFAOYSA-M potassium bromide Chemical compound [K+].[Br-] IOLCXVTUBQKXJR-UHFFFAOYSA-M 0.000 description 2
- 230000002265 prevention Effects 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- 238000004062 sedimentation Methods 0.000 description 2
- 229960004025 sodium salicylate Drugs 0.000 description 2
- 239000002351 wastewater Substances 0.000 description 2
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 1
- ZGTMUACCHSMWAC-UHFFFAOYSA-L EDTA disodium salt (anhydrous) Chemical compound [Na+].[Na+].OC(=O)CN(CC([O-])=O)CCN(CC(O)=O)CC([O-])=O ZGTMUACCHSMWAC-UHFFFAOYSA-L 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 239000012670 alkaline solution Substances 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 1
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 1
- 235000011130 ammonium sulphate Nutrition 0.000 description 1
- 238000004737 colorimetric analysis Methods 0.000 description 1
- 229910052802 copper Inorganic materials 0.000 description 1
- 239000010949 copper Substances 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 239000003651 drinking water Substances 0.000 description 1
- 235000020188 drinking water Nutrition 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 230000002452 interceptive effect Effects 0.000 description 1
- 238000005342 ion exchange Methods 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- 150000002731 mercury compounds Chemical class 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 229910021645 metal ion Inorganic materials 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000013535 sea water Substances 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 238000001256 steam distillation Methods 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 238000004448 titration Methods 0.000 description 1
- 238000004065 wastewater treatment Methods 0.000 description 1
Landscapes
- Investigating Or Analyzing Non-Biological Materials By The Use Of Chemical Means (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Description
【発明の詳細な説明】 本発明は液体中のアンモニアを測定する方法に関する。[Detailed description of the invention] The present invention relates to a method for measuring ammonia in a liquid.
さらに詳しくはアンモニアとフェノールを次亜塩素酸塩
と処理し生成するインドフェノールを比色定量するいわ
ゆるバースロット法（Ｊ・Ｂｅｒｔｈｅｌｏｔ：Ｒｅｐ
ｏｒｔ ｃｈｅｍ、Ａｐｐｔ １，２８４゜１８５９）
の改良に関する。More specifically, the so-called Berthelot method (J. Berthelot: Rep.
ort chem, Appt 1,284°1859)
Regarding improvements.
従って液体中のアンモニアは排水中のものであれ、空気
中のアンモニアを希硫酸（０，０２Ｎ）に溶解させたも
のであれ、食品類から抽出したものであれ、また生体液
中のものであれ、いずれの場合にも適用することができ
る。Therefore, ammonia in liquids may be in waste water, ammonia in the air dissolved in dilute sulfuric acid (0.02N), extracted from foods, or biological fluids. , can be applied in either case.
ただ本発明者等が生体液特に血清中の尿素窒素の新しい
安全な測定方法を研究する過程で到達した発明であるの
でこの分野では特にメリットが大きい。However, since this invention was achieved by the present inventors in the process of researching a new and safe method for measuring urea nitrogen in biological fluids, especially serum, it has particular merits in this field.
アンモニアの測定方法は歴史的に数多くの開発、改良が
なされ検体の種類、性質等によっているいろ使い分けら
れて来た。Historically, many methods for measuring ammonia have been developed and improved, and various methods have been used depending on the type and properties of the sample.
例えば滴定法は水蒸気蒸留あるいは微量拡散法により分
離するアンモニアに適用するが、いずれも操作が非常に
繁雑で蒸留装置や微量拡散装置などの特殊な装置が必要
でありまた測定に長時間を要しさらに再現性も悪い等の
欠点があるため現在では殆ど用いられていない。For example, the titration method is applied to ammonia separated by steam distillation or microdiffusion, but both methods are extremely complicated to operate, require special equipment such as a distillation device or microdiffusion device, and require a long time for measurement. Furthermore, it has drawbacks such as poor reproducibility, so it is hardly used at present.
これに対し比色法は比色計の普及に伴って容易に測定が
可能になったため現在広く行なわれている。On the other hand, the colorimetric method is now widely used because it has become easier to measure with the spread of colorimeter.
その中でネスラー法、バースロット法（インドフェノー
ル法）が一番広く行なわれている。Among them, the Nessler method and Barthlot method (indophenol method) are the most widely used.
ネスラー法は試薬としてヨウ化水銀と臭化カリウムを用
いこれにアンモニアを作用させ黄赤色の錯化合物を生成
させこれを比色定量する方法である。The Nessler method uses mercury iodide and potassium bromide as reagents, and reacts with ammonia to produce a yellow-red complex, which is then measured colorimetrically.
この方法は古くから用いられて来たが試薬が不安定とい
う欠陥がある上現在では水銀化合物の使用が公害の上か
ら欠点となっている。Although this method has been used for a long time, it has the disadvantage that the reagents are unstable, and the use of mercury compounds is currently a disadvantage due to the pollution.
インドフェノール法は前述の通りフェノールとアンモニ
アを酸化してインドフェノールを生成しこれを測定する
方法であるが本発明においてはフェノールの替りにサリ
チル酸ナトリウムを使用する場合も含む。As mentioned above, the indophenol method is a method of oxidizing phenol and ammonia to produce indophenol and measuring it, but the present invention also includes cases where sodium salicylate is used instead of phenol.
いずれにしても反応速度をはやめるため触媒としてニト
ロプルジッドナトリウムを加えインドフェノールの発色
を増大させている。In any case, in order to speed up the reaction rate, sodium nitropurgide is added as a catalyst to increase the color development of indophenol.
インドフェノール法は感度がネスラー法より１０倍も高
く現在微量のアンモニアの定量には最も適した方法とい
える。The indophenol method is 10 times more sensitive than the Nessler method and can be said to be the most suitable method for quantifying trace amounts of ammonia at present.
しかしこの方法もネスラー法のヨウ化水銀（毒物）使用
と同様ニトロプルジッドナトリウムというシアン化合物
（毒物）を使用するところに欠点がある。However, this method also has a drawback in that it uses a cyanide compound (poisonous substance) called sodium nitroprusid, similar to the use of mercury iodide (a poisonous substance) in the Nessler method.
アンモニアは古くから飲料水の衛生学的安全度の指標物
質として重視されているほか最近では水中アンモニアの
多寡が水中微生物の発生に重大な影響を与えるため海水
中のアンモニアの調査、また排水基準、汚水処理の目安
などにアンモニアの測定の機会及び重要性は増加の一途
にある。Ammonia has long been valued as an indicator of the sanitary safety of drinking water, and recently, the amount of ammonia in water has a significant impact on the occurrence of microorganisms in the water, so ammonia in seawater has been investigated, as well as wastewater standards. The opportunities and importance of measuring ammonia as a guideline for wastewater treatment are increasing.
このような状況下で感度の高い公害のおそれのない安全
なアンモニア測定方法の確立が強く望まれている。Under these circumstances, there is a strong desire to establish a safe ammonia measurement method that is highly sensitive and does not pose a risk of pollution.
本発明者等は血清中の尿素窒素測定の分野で酵素ウレア
ーゼを用いて血清中の尿素を分解し生成するアンモニア
をインドフェノール法で測定し尿素窒素を定量する方法
において、ニトロプルジッドナトリウムが公害上問題が
あることからこれに替る新しい触媒を研究した結果、ピ
ラゾールが最適であることを発見し本発明を完成した。In the field of measuring urea nitrogen in serum, the present inventors used the enzyme urease to decompose urea in serum and measured ammonia produced using the indophenol method. Due to the above problems, as a result of research into a new catalyst to replace it, they discovered that pyrazole was the most suitable, and completed the present invention.
本発明の方法はバースロット法（インドフェノール法）
において触媒として使用するニトロプルジッドナトリウ
ムに替えピラゾールを使用し生成するインドフェノール
を測定し検量線からアンモニアを定量する方法である。The method of the present invention is the Barthlot method (indophenol method).
In this method, pyrazole is used in place of sodium nitropurgide used as a catalyst, the indophenol produced is measured, and ammonia is determined from a calibration curve.
次に実施例により本発明の詳細な説明する。Next, the present invention will be explained in detail with reference to Examples.
実施例 １
血清中の尿素窒素の測定：
（１）原理
血清中の尿素に酵素ウレアーゼを作用させ分解し生成し
たアンモニアを本発明方法で測定する。Example 1 Measurement of urea nitrogen in serum: (1) Principle Urea in serum is decomposed by the action of the enzyme urease, and ammonia produced is measured by the method of the present invention.
（２）試薬
（■） Ａ液
ピラゾール ５１．０６ｇ（０，７５Ｍ）サ
リチル酸ナトリウム ６４．０ ｇ （０，４Ｍ）Ｎａ
２ＨＰＯ４１，６１ｇ
ＫＨ２ＰＯ４１，１８ｇ
ＥＤＴＡ・２Ｎａ ２．０ｇ水に溶解全量
を１１とする
（■）Ｂ液
Ｎａ０Ｃｌ 有効塩素量０．２％水溶液１Ｏ−５ＮＮ
ａＯＨでｐＨ１１，０に調製（■）ウレアーゼ液
ウレアーゼ ２８００ｕ
ＮａＣｌ １．９５ｇ
リン酸緩衝液（ｐＨ７，０）１ｄｌに溶解（■） 標準
液
尿素 ６４．３ｍｇ／ｄｌ（尿素窒素３０ｍｇ／ｄｌ
）水に溶解
水は全てイオン交換純水を使用する。(2) Reagent (■) Liquid A pyrazole 51.06g (0.75M) Sodium salicylate 64.0g (0.4M) Na
2HPO41.61g KH2PO41.18g EDTA・2Na 2.0g Make the total amount dissolved in water to 11 (■) Solution B Na0Cl Available chlorine amount 0.2% aqueous solution 1O-5NN
Adjust to pH 11.0 with aOH (■) Urease solution Urease 2800u NaCl 1.95g Dissolved in 1 dl of phosphate buffer (pH 7.0) (■) Standard solution Urea 64.3 mg/dl (Urea nitrogen 30 mg/dl
) Use ion-exchanged pure water for all water dissolved in water.
（３）操作
試料血清０．０２ｍ１を試験管にとりウレアーゼ液０．
０５ｍ１及びＡ液１．０ｍｌを加え３７℃で１０分間イ
ンキュベートし酵素反応を行なわせアンモニアを遊離す
る。(3) Take 0.02ml of operation sample serum into a test tube and add 0.0ml of urease solution.
Add 05ml and 1.0ml of solution A and incubate at 37°C for 10 minutes to perform an enzyme reaction and release ammonia.
次にＢ液２．０ｍｌを加え再び３７℃で１５分間インキ
ュベートし発色させて続いて５分間流水で冷却すること
によって反応を停止する。Next, 2.0 ml of solution B was added, and the mixture was incubated again at 37° C. for 15 minutes to develop color, and then the reaction was stopped by cooling with running water for 5 minutes.
波長６５５ｎｍを用い試薬ブランクを対照とし吸光度を
測定し次の計算式から尿素窒素量を求めるか、別に常法
により作製した検量線（第２図参照）から濃度を求める
。The absorbance is measured at a wavelength of 655 nm using a reagent blank as a reference, and the amount of urea nitrogen is determined from the following calculation formula, or the concentration is determined from a calibration curve (see Figure 2) prepared separately using a conventional method.
計算式
本発明方法を血清中の成分特に尿素窒素測定に適用した
場合の効果について説明すると次の通りである。Calculation formula The effects when the method of the present invention is applied to the measurement of serum components, particularly urea nitrogen, are as follows.
本発明方法の発色の波長吸収曲線は第１図に示す通りで
標準液血清共に６５５ｎｍに最大吸収を示している。The wavelength absorption curve of color development by the method of the present invention is shown in FIG. 1, and both the standard solution and serum show maximum absorption at 655 nm.
試薬にブランクに発色が見られないことは試薬の安定性
の良いことを示している。The fact that no color development is observed in the reagent blank indicates that the reagent has good stability.
反応後の呈色の安定性は第３図に示す通りで反応終了後
１時間は安定であることが明らかである。The stability of the color after the reaction is as shown in Figure 3, and it is clear that the color is stable for 1 hour after the completion of the reaction.
次に本発明方法において血清中に共存する尿素窒素以外
の窒素成分及びその他の共存物質の影響を調べた結果は
表１に示す通りであり、かなり多量を添加した場合でも
全く影響のないことが分った。Next, in the method of the present invention, we investigated the effects of nitrogen components other than urea nitrogen and other coexisting substances coexisting in serum, and the results are shown in Table 1, which shows that there is no effect at all even when a fairly large amount is added. I understand.
本発明方法の吸光度と尿素窒素濃度との関係は直線性を
示し検量線を求めてみると第２図に示す通り２００ｍｇ
／ｄｌまでの直線性が得られかなり高濃度まで直接測定
が可能であることが判明した。The relationship between the absorbance and the urea nitrogen concentration in the method of the present invention shows linearity, and when a calibration curve is obtained, as shown in Figure 2, the relationship between the absorbance and the urea nitrogen concentration is 200 mg.
It was found that linearity up to /dl was obtained and direct measurement was possible up to quite high concentrations.
従来のウレアーゼ−インドフェノール法のように最大吸
収波長をずらせて測定したり、試料の血清を前もって希
釈して測定し直す必要などはない。Unlike the conventional urease-indophenol method, there is no need to shift the maximum absorption wavelength for measurement or to dilute the sample serum in advance and remeasure it.
また本発明方法の同時再現性試験結果は表２に示す通り
で変動係数ＣＶ：２．４９％、標準偏差５Ｄ＝０．３７
と極めて高い再現性を示した。In addition, the simultaneous reproducibility test results of the method of the present invention are shown in Table 2. Coefficient of variation CV: 2.49%, standard deviation 5D = 0.37
showed extremely high reproducibility.
行なった正確度試験の結果は表３に示す通りで正常域、
異常域とも良好な結果を得た。The results of the accuracy test conducted are shown in Table 3, and are in the normal range.
Good results were obtained in both abnormal areas.
次に従来広く使用されているウレアーゼ−インドフェノ
ール法とウレアーゼ一本発明方法との相関を求めた結果
は第４図に示す通りで相関係数σ＝０．９９と極めて高
い相関を示した。Next, the correlation between the conventionally widely used urease-indophenol method and the urease-indophenol method of the present invention was determined, and the results are shown in FIG. 4, showing an extremely high correlation with a correlation coefficient σ=0.99.
以上説明した通り本発明の方法を利用する血清中の尿素
窒素測定方法は従来量も優れているとされているウレア
ーゼ−インドフェノール法（触媒としてニトロプルジッ
ドを使用）に比較して優るとも劣らない効果を有するば
かりでなくピラゾールの使用によりニトロプルジッドナ
トリウムを排除することができ公害防止の見地からより
優れた測定方法といえる。As explained above, the method for measuring urea nitrogen in serum using the method of the present invention is superior to the conventional urease-indophenol method (using nitroprusid as a catalyst), which is said to have an excellent amount. Not only does this method have a negative effect, but also the use of pyrazole eliminates nitroprusdide sodium, making it a superior measurement method from the standpoint of pollution prevention.
なお血清中の遊離アンモニアの測定を必要とするときは
上記実施例１から容易に実施することができる。Note that when it is necessary to measure free ammonia in serum, it can be easily carried out from Example 1 above.
一般の水溶液中のアンモニアの測定においてもインドフ
ェノール法が利用できる場合は殆ど本発明方法が使用で
きその効果もまた同様である。In the measurement of ammonia in general aqueous solutions, the method of the present invention can be used in most cases where the indophenol method can be used, and the effects are also the same.
実施例によって説明する。This will be explained by an example.
実施例 ２
（１） 試薬
Ａ液：
ピラゾール ２５５．３ｇ（３，７５Ｍ）Ｎａ２ＨＰ
Ｏ４１，６１ｇ
ＫＨ２ＰＯ４１，１８ｇ
ＥＤＴＡ−２Ｎａ １０．０ ｇ
水に溶解し全量を１１とする。Example 2 (1) Reagent A solution: Pyrazole 255.3g (3,75M) Na2HP
O41,61g KH2PO41,18g EDTA-2Na 10.0g Dissolve in water to bring the total amount to 11.
Ｂ液：
ＮａＣ１０有効塩素量０．２％水溶液
アンモニア標準液：
（ＮＨ４）２ＳＯ４０，６８３ｍｇ／ｄｌ水溶液（ＮＨ
４＋：０．５ｍｇ／ｄｌ、５．０ｐｐｍ）水は全てイオ
ン交換純水を使用する。Solution B: NaC10 available chlorine amount 0.2% aqueous solution Ammonia standard solution: (NH4)2SO40, 683 mg/dl aqueous solution (NH
4+: 0.5 mg/dl, 5.0 ppm) All water used is ion exchange pure water.
（２）操作法
試料検水５．０ｍｌを試験管にとり、Ａ液０．５ｍｌを
加えよく混和した後Ｂ液１．０ｍｌを加え３７℃で１５
分インキュベートして反応を行なわせる。(2) Procedure: Take 5.0ml of sample water into a test tube, add 0.5ml of liquid A, mix well, add 1.0ml of liquid B, and heat at 37°C for 15 minutes.
Incubate for 1 minute to allow the reaction to occur.
次に流水で５分間冷却し反応を停止させた後波長６５５
ｎｍを用い試薬ブランクを対照として吸光度を測定し次
の計算式からアンモニア量を求めるか、別に常法により
作製した検量線（第５図参照）から濃度を求める。Next, after cooling with running water for 5 minutes to stop the reaction, the wavelength is 655.
The absorbance is measured using a reagent blank as a reference using nm, and the amount of ammonia is determined from the following calculation formula, or the concentration is determined from a calibration curve (see FIG. 5) prepared separately by a conventional method.
計算式
一般に試料が検水の場合は試料が汚濁、着色または高濃
度の塩類を含む場合があるが、このような場合には凝集
沈澱法等によって前処理をする必要がある。Calculation formula Generally, when the sample is water test, the sample may be contaminated, colored, or contain high concentration of salts, but in such cases, it is necessary to perform pretreatment using a coagulation sedimentation method or the like.
１例を示せば、参考例
前処理としての凝集沈澱法
検水１００ｍ１を三角フラスコにとり、ＺｎＳＯ４・７
Ｈ２Ｏ水溶液（１０ｇ／ｄｌ） １．０ｍｌを加えよく
混和した後、アルカリ溶液（ＮａＯＨ１５ｇ／ｄｌ。To give an example, 100 ml of coagulation-sedimentation test water as a reference example pretreatment was placed in an Erlenmeyer flask, and ZnSO4.7
After adding 1.0 ml of H2O aqueous solution (10 g/dl) and mixing well, add an alkaline solution (NaOH 15 g/dl).
Ｎａ２ＣＯ３１２，５ｇ／ｄｌ）及び酸溶液（１１５Ｏ
ＮＨ２ＳＯ４）を使ってｐＨ７，０に調製し、しばらく
静置後３０００ｒｐｍ、１０分遠心分離するか、または
濾過により透明な液とし測定に用いる。Na2CO312,5g/dl) and acid solution (115O
Adjust the pH to 7.0 using NH2SO4), let stand for a while, and then centrifuge at 3000 rpm for 10 minutes or filter to make a clear liquid and use it for measurement.
また検水中のアンモニアが特に高い場合は予めイオン交
換純水で５．０ｐｐｍ付近になるように希釈倍率をかけ
て補正する。Furthermore, if the ammonia in the test water is particularly high, it should be corrected in advance by applying a dilution factor to bring it to around 5.0 ppm with ion-exchanged pure water.
検水中のアンモニア濃度と吸光度の関係は直線性を示し
検量線を求めて見ると第５図に示す通りで０．ｌｐｐｍ
から１０．０ｐｐｍまで直線性が得られ、感度も良好で
０．ｌｐｐｍから１０．０ｐｐｍまでの測定が可能であ
る。The relationship between the ammonia concentration in the sample water and the absorbance showed linearity, and when we calculated the calibration curve, it was 0.0 as shown in Figure 5. lppm
Linearity was obtained from 10.0 ppm to 10.0 ppm, and sensitivity was also good with 0.0 ppm. Measurement from lppm to 10.0ppm is possible.
波長吸収曲線、呈色安定性は実施例１の場合と同様であ
る。The wavelength absorption curve and color stability are the same as in Example 1.
検水中には種々の金属塩やその他組成不明の妨害物質が
含まれている場合がある。The sample water may contain various metal salts and other interfering substances of unknown composition.
鉄、銅等の金属イオンはＥＤＴＡ・２Ｎａによりその作
用をマスクしその妨害を除くことができる。The effects of metal ions such as iron and copper can be masked by EDTA/2Na and their interference can be removed.
アミノ酸、脂肪族アミンには影響を受けないが芳香族ア
ミンの一部例えば表４に示す通りＯ−トリイジン、エチ
ルアニリンには妨害を受ける。It is not affected by amino acids and aliphatic amines, but is interfered with by some aromatic amines, such as O-triidine and ethylaniline as shown in Table 4.
このような場合には例えば蒸留、凝集沈澱等の前処理を
行なわなければならないときもある。In such cases, it may be necessary to carry out pretreatments such as distillation, coagulation and precipitation.
試験方法は（ＮＨ４）２ＳＯ４（濃度４．０ｐｐｍ）を
含む検水を用い芳香族アミンの添加以外は実施例２に準
じて行なった。The test method was carried out in accordance with Example 2 using sample water containing (NH4)2SO4 (concentration 4.0 ppm) except for the addition of aromatic amine.
上述の通り本発明方法が液体中のアンモニア測定に広く
用いられ特に血清中の尿素窒素あるいは遊離アンモニア
の測定に欠くことのできないインドフェノール法の公害
防止を達成したことは産業上極めて有用なことである。As mentioned above, the method of the present invention is widely used for measuring ammonia in liquids, and is extremely useful industrially because it has achieved pollution prevention using the indophenol method, which is indispensable for measuring urea nitrogen or free ammonia in serum. be.
第１図は本発明方法の発色の波長吸収曲線を示す。
第２図は本発明方法による血清中の尿素窒素の検量線を
示す。
第３図は本発明方法の発色後の呈色の安定性を示すグラ
フである。
第４図は従来法と本発明方法との血清中の尿素窒素測定
における相関を示すグラフである。
第５図は本発明方法による検水中のアンモニアの検量線
を示す。FIG. 1 shows the wavelength absorption curve of color development by the method of the present invention. FIG. 2 shows a calibration curve for urea nitrogen in serum according to the method of the present invention. FIG. 3 is a graph showing the stability of color development after color development using the method of the present invention. FIG. 4 is a graph showing the correlation between the conventional method and the method of the present invention in measuring urea nitrogen in serum. FIG. 5 shows a calibration curve for ammonia in sample water according to the method of the present invention.
Claims (1)
おいて触媒として使用するニトロプルジッドナトリウム
の替りにピラゾールを使用することを特徴とする液体中
のアンモニアの測定方法。1. A method for measuring ammonia in a liquid, which is characterized in that pyrazole is used in place of sodium nitropurgide used as a catalyst in the bar slot method, which is one of the methods for measuring ammonia in a liquid.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP9506579A JPS5811024B2 (en) | 1979-07-27 | 1979-07-27 | How to measure ammonia in liquid |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP9506579A JPS5811024B2 (en) | 1979-07-27 | 1979-07-27 | How to measure ammonia in liquid |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS5619452A JPS5619452A (en) | 1981-02-24 |
| JPS5811024B2 true JPS5811024B2 (en) | 1983-03-01 |
Family
ID=14127599
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP9506579A Expired JPS5811024B2 (en) | 1979-07-27 | 1979-07-27 | How to measure ammonia in liquid |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS5811024B2 (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS638309U (en) * | 1986-07-03 | 1988-01-20 | ||
| JPH0354443U (en) * | 1990-09-07 | 1991-05-27 | ||
| WO2008047802A1 (en) | 2006-10-18 | 2008-04-24 | National University Corporation Nagoya University | D-serine dehydratase and use thereof |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2739484B2 (en) * | 1988-09-29 | 1998-04-15 | 株式会社ヤトロン | Urea nitrogen measurement reagent for automatic analyzer |
| GB9603031D0 (en) * | 1996-02-14 | 1996-04-10 | Boots Co Plc | Diagnostic method |
| CN100429505C (en) * | 2005-09-21 | 2008-10-29 | 广东环凯微生物科技有限公司 | Ammonia nitrogen concentration detection reagent and usage in water |
-
1979
- 1979-07-27 JP JP9506579A patent/JPS5811024B2/en not_active Expired
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS638309U (en) * | 1986-07-03 | 1988-01-20 | ||
| JPH0354443U (en) * | 1990-09-07 | 1991-05-27 | ||
| WO2008047802A1 (en) | 2006-10-18 | 2008-04-24 | National University Corporation Nagoya University | D-serine dehydratase and use thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| JPS5619452A (en) | 1981-02-24 |
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