Application Note No. 191/2015 TKN determination in water and waste water
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1 Application Note No. 191/2015 TKN determination in water and waste water KjelDigester K-449, KjelMaster K-375 with KjelSampler K-376: Colorimetric determination of TKN (Total Kjeldahl Nitrogen) in water and waste water according to the Kjeldahl method
2 1 Introduction A reliable method for the determination of total Kjeldahl nitrogen (TKN) in water and waste water, according to ISO 5663, DIN EN , AOAC and EPA 351.3, is introduced [1] [2] [3]. The samples are digested using the KjelDigester K-449. The distillation and boric acid titration are performed with the KjelMaster K-375 with KjelSampler K-376. Equivalent to the international norms, the detection principle of the boric acid titration is colorimetric. I.e. a mixed indicator according to Sher is added to the boric acid solution and the KjelMaster K-375 is equipped with the colorimetric sensor. The fast heating and cooling of the KjelDigester K-449, the synchronized process steps of the KjelMaster system K-375 / K-376 combined with the titration mode Online increases the sample throughput per work day. 2 Equipment KjelDigester K-449 (the parameters used are also valid for the K-446) User protection shield (BUCHI ) Scrubber K-415 TripleScrub ECO with TKN Set (BUCHI ) KjelMaster K-375 with colorimetric sensor KjelSampler K-376 (the parameters used are also valid for the K-377) Analytical balance (accuracy ± 0.1 mg) Digestion rods, boiling aid (BUCHI ) Volumetric pipettes 3 Chemicals and Materials Chemicals: Sulfuric acid conc. 98 %, analytical reagent, Beijing Chemical Works Titanium, BUCHI Kjeldahl Tablet ( ) Sodium hydroxide 32 %, analytical reagent, Sinoharm chemical reagent Boric acid 2 %: 100 g boric acid, (analytical reagent, Tianjin guangfu Fine chemical research institute) diluted to 5 L with deionized water, dissolve completely, add 12.5 ml of mixed Sher indicator, BUCHI (003512), adjust ph to 4.65 Sulfuric acid 0.01 mol/l, Aladdin (S L) Neutralization solution for the Scrubber: 600 g sodium carbonate (analytical reagent, Sinoharm chemical reagent) about 2 ml ethanol and a spatula tip of bromthymol blue, (analytical reagent, Tianjin Kemiou Chemical Reagent) diluted to 3 L with distilled water Urea, assay %, Aladdin (U g) For a safe handling please pay attention to all corresponding MSDS. Samples: Urea stock solution, g Urea diluted to 1000 ml with deionized water mg N/mL Surface water of a river (city moat, slightly turbid), Beijing China Application Note 191/2015 June /9
3 4 Procedure The determination of nitrogen in water and waste water includes the following steps: Acidify the water sample with 0.5 ml sulfuric acid (conc.) per liter for preservation Digestion of the sample, using the KjelDigester K-449 (K-446 respectively) Distillation and colorimetric titration of the sample, using KjelMaster system K-375 / K Digestion method urea stock solution (verification of the method) 1. Start the KjelDigester K-449 according to the parameters listed in Table 1 and 2 2. Place different volumes of the urea stock solution in a 300 ml sample tube and dilute with deionized water to the required volume (e.g. 4 ml of stock solution diluted with 21 ml deionized water to total volume of 25 ml) 3. Add 1 Titanium Kjeldahl Tablet, 8 ml of sulfuric acid (conc. 98 %) and 1 digestion rod to each tube (immerge the end with the void into the sample) to avoid bumping (!) 4. Prepare additional blanks, chemicals without sample, with the corresponding volume of deionized water 5. Attach the user protection shield to the rack (it protects the user in case of boiling delays) 6. Connect the Scrubber K-415 to the K-449 for absorbing the acid fumes created during digestion 7. Insert the rack with the samples into the cooling position and mount the suction module onto the samples, immediately start the digestion according to the parameters listed in Table Let the samples cool down when the digestion is completed. 4.2 Digestion method samples 1. Start the KjelDigester K-449 according to the parameters listed in Table 2 2. Place each sample in a 300 ml sample tube as described in Table 1 Table 1: Nitrogen content depending sample volume TKN [mg/l] Volume [ml] Digestion Time [min] Add 1 Titanium Kjeldahl Tablet, 8 ml of sulfuric acid (conc. 98%) and 1 digestion rod to each tube (immerge the end with the void into the sample) to avoid bumping (!) 4. Prepare additional blanks, chemicals without sample, with the corresponding volume of deionized water 5. Attach the user protection shield to the rack (it protects the user in case of boiling delays) 6. Connect the Scrubber K-415 to the K-449 for absorbing acid fumes created during digestion 7. Insert the rack with the samples into the cooling position and mount the suction module onto the samples, immediately start the digestion according to the parameters listed in Table Let the samples cool down when the digestion is completed. Application Note 191/2015 June /9
4 Table 2: Temperature profile for digestion with the K-449 Step Temperature [ C] Time [min] See Table 1 Cooling 35 NOTE: If the liquid inside the sample tube is not clear and blue-green just after the digestion finished, digest for additional 15 min at 420 C. 4.3 Distillation and titration For colorimetric titration it is necessary to determine the setpoint of the boric acid solution in advance to the blank and sample determinations. It is necessary to determine the setpoint every day before starting sample determinations, and when the method is changed or fresh chemicals are used to adjust the device to the current conditions. The detailed procedure including the preparation of the sensor is described in the Technical Note 179/2015 Colorimetric titration procedure using Sher indicator [4]. The setpoint was measured three times. 1 st setpoint preheating 2 nd setpoint 1 st measurement 3 rd setpoint 2 nd measurement, confirms the 1 st measurement The last setpoint measurement is used as endpoint for all following determinations including priming, blanks and samples. 1. Determine the setpoint and check it`s range and deviation: Select all paramters for the setpoint determination according to Table 3. Table 3: Parameters for setpoint determination Parameter Setting Preheating before setpoint yes Setpoint runs 3 Setpoint cycle Via sampler Boric acid 2 % Indicator Sher Method Select the same method as for sample determination NOTE: The selected method, boric acid and indicator for setpoint determination must be identical to the method used for sample determination! 2. Check the setpoint range and deviation The determined setpoints should be in a range of mv The deviation between the two last measured setpoints should be 20 mv 3. Perform a priming to remove all residues. 4. Determine blanks according to the parameters listed in Table Determine samples according to the parameters listed in Table 5. Table 4: Setpoint measurements and deviation Setpoint 1 Setpoint 2 Deviation mv mv 8.5 mv Table 5: Distillation and titration with the KjelMaster system K-375 / K-376 Application Note 191/2015 June /9
5 H2O volume 50 ml Titration solution H2SO mol/l NaOH volume 60 ml Sensor type Colorimetric Reaction time 5 s Titration mode Online Distillation mode Fixed time Titration start time 90 s Distillation time 180 s Measuring mode Setpoint Stirrer speed distillation 5 Stirrer speed titration 10 Steam output 100 % Titration start volume 0 ml Titration type Boric acid Titration algorithm Optimal Receiving solution vol. 60 ml NOTE: The sample throughput for this application was increased by using the Online titration mode: By applying the Online titration the time for the distillation and titration process is reduced to about 5 minutes per analysis because titration starts during the distillation is still in progress. NOTE: The titration start time must not be less than 60 seconds 4.4 Calculation The results are calculated as TKN (Total Kjeldahl Nitrogen). For the urea stock solution, the recovery was caslculated (2). TKN (V - V m ) Sample z c f MN 1000 Sample Blank actual (1) TKN actual Recovery *100 (2) TKN theoretical TKNactual : Total Kjeldahl nitrogen [mg/l] VSample VBlank z c : amount of titrant for the sample [ml] : mean amount of titrant for the blank [ml] : molar valence factor (1 for HCl, 2 for H2SO4) : titrant concentration [mol/l] f : titrant factor (for commercial solutions normally 1.000) MN msample : molecular weight of nitrogen ( g/mol) : sample volume [ml] 1000 : conversion factor [ml/l] Recovery : of the urea stock solution [%] Application Note 191/2015 June /9
6 4.5 Limit of Detection (LOD) and Limit of Quantification (LOQ) The blank method from DIN [5] was used for the determination of the detection limit (LOD) and the quantification limit (LOQ). Based on these limits the performance of the method can be evaluated. First, ten blanks were determined using 200 ml deionised water, one Titanium Kjeldahl Tablet and 8 ml sulfuric acid. The blanks were digested and determined according to the parameters listed in Table 2 and 3, the results are presented in Table 6. Table 6: Results of the blank determination (Volume of deionized water 200 ml) VBlank [ml] Average [ml] SD [ml] RSD [%] The following equation (3) was used to calculate the limit of detection (LOD). ᶲn;α SD LOD = ᶲn;α SD (3) LOD = ml : factor 3.0; depending on the number of blanks (n=10) and the level of significance (α=0.01) : standard deviation of the blank determination (SD=0.019 ml) Depending on the LOD, the limit of quantification (LOQ) can be calculated, see equotation (4). LOQ = k ᶲn;α SD (4) LOQ = ml k : factor 3 (for more information refer to DIN 32645) According to DIN 32645, the LOD =0.081 mg/l and LOQ = 0.25 mg/l for 200 ml of sample could be estimated. Application Note 191/2015 June /9
7 5 Results 5.1. Recovery of urea stock solution The results of TKN determination and recovery for urea stock solution with different sample volumes are presented in Tables 7-8. Table 7: Results of the recovery of TKN in urea stock solution with a total sample volume of 25 ml 25 ml VSample [ml] TKNtheoretical [mg/l] TKNactual [mg/l] Recovery [%] Sample Sample Sample Average [%] RSD [%] The mean blank volume (VBlank) was ml (n = 3). Sample Sample Sample Average [%] RSD [%] The mean blank volume (VBlank) was ml (n = 3). Table 8: Results of the recovery of TKN in urea stock solution with a total sample volume of 200 ml 200 ml VSample [ml] TKNtheoretical [mg/l] TKNactual [mg/l] Recovery [%] Sample Sample Sample Average [%] RSD [%] The mean blank volume (VBlank) was ml (n = 3). Sample Sample Sample Average [%] RSD [%] The mean blank volume (VBlank) was ml (n = 3). 5.2 TKN determination in water samples The results of the TKN determination in samples of river surface water (city moat, slightly turbid, Beijing, China), are presented in Table 9. Table 9: Results of the TKN determination in surface water of a pond (Beijing, China) with total sample volume of 200 ml VSample [ml] TKNactual [mg/l] Sample Sample Sample Average [mg/l] 0.97 Rsd [%] 5.0 The mean blank volume (VBlank) was ml (n = 3). Acording to the AOAC and EPA regulations, these results correspond well to the low content of Total Kjeldahl Nitrogen (TKN) in water and waste water with low relative standard deviations (Rsd). Application Note 191/2015 June /9
8 6 Comparison to Standard Methods The differences between this application and the official methods ISO 5663, AOAC and EPA are shown in Table 10. The AOAC and EPA regulations are similar with minor differences as indicated in Table 10. Table 10: Differences to ISO 5663 and AOAC Application note ISO 5663 DIN EN Catalyst 3.7 g Tablets Composition: 94.4 % K2SO4 2.8 % TiO2 2.8 % CuSO4*5H2O 5 g mixture Composition: 99.0 % K2SO4 1.0 % Se AOAC EPA ml digestion solution cont. 13 g K2SO4 0.2 g Hg Notes / Impact The choice of catalyst does not influence the result. No use of toxix Se/Hg! Easy handling using tablets. Sulfuric acid 8 ml 10 ml 10 ml No impact, same ration of sulfuric acid/catalyst. Water 50 ml 250 ±50 ml 300 ml The K-375 generates steam in a separated vessel; therefore it is not necessary to add such a high amount of water Sodium hydroxide 60 ml (conc. 32 %) 50 ml (conc. 36 %) 60 ml (conc. 50 %) (EPA: 100 ml) No impact, same ratio of sodium hydroxide/sulfuric acid. Na2S2O3 no no yes Only necessary when Hg is used as catalyst Titration measurement method Indicator Titration solution colorimetric colorimetric colorimetric (EPA: colorimetric & potentiometic) Mixed indicator according to Sher H2SO mol/l Methyl red / methylene blue HCl 0.02 mol/l Methyl red / methylene blue H2SO mol/l No impact, the results are equal. No impact, Sher indicator has even a more accurate transition point Application Note 191/2015 June /9
9 7 Conclusion The determination of TKN (Total Kjeldahl Nitrogen) in water using the KjelDigester K-449, the KjelMaster system K-375 / K-376 and the colorimetric sensor provides reliable and reproducible results, even for nitrogen contents close to the limit of quantification. The recovery with urea stock solution was very good with low standard deviation. The LOD is mg/l and the LOQ is 0.25 mg/l for 200 ml sample volume according to DIN With the KjelDigester K-449 the digestion process (including preheating, digestion and cooling) is very fast and is fully automated. Together with the fully-automatic KjelMaster system K-375/K-376 and the novel Online titration mode, the time to results is significantly reduced and it offers fully walk-away convenience. 8 References [1] ISO 5663 Water quality Determination of Kjeldahl nitrogen Method after mineralization with selenium [2] AOAC Nitrogen (Total) in Water [3] EPA Nitrogen, Kjeldahl, Total (colorimetric or titrimetric or potentiometric) [4] Technical Note No.179/2015 Colorimetric titration procedure using Sher indicator [5] DIN Nachweis-, Erfassungs- und Bestimmungsgrenze Application Note 191/2015 June /9
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