The calibration of the total nitrogen analyzer should follow the logic of "preparation first, calibration later, and verification later". The core is to establish the correspondence between the instrument response value and the total nitrogen concentration through standard substances to ensure accurate detection results. The following are the specific steps (taking the common alkaline potassium persulfate digestion UV spectrophotometry principle of total nitrogen analyzer as an example):
1、 Preparation before calibration
Instrument and reagent preparation
Ensure that the instrument is in normal working condition: power on and preheat (according to the instructions for 30-60 minutes), check whether the key parameters such as the temperature of the digestion module, the intensity of the UV detector light source, and the sealing of the injection system are normal.
Preparation of standard solution: Use nationally recognized total nitrogen standard reserve solution (such as 1000mg/LKNO ∝ standard solution), and dilute it step by step with ammonia free water to form a standard series of at least 3 concentration points (the concentration range should cover the expected concentration of daily test samples, such as 0.5mg/L, 2mg/L, 5mg/L, 10mg/L, and the blank is ammonia free). Standard solutions need to be prepared and used immediately to avoid contamination.
Prepare auxiliary reagents: Prepare alkaline potassium persulfate solution, hydrochloric acid solution (for neutralizing and digesting the sample), etc. according to the instrument requirements, ensuring that the purity of the reagents meets the analytical requirements (such as superior purity).
Sample container and system cleaning
The colorimetric tube and injection bottle used for calibration should be soaked in hydrochloric acid, rinsed repeatedly with ammonia free water, and dried to avoid residual nitrogen pollution.
Run the "pipeline cleaning" program of the instrument, rinse the injection pipeline, digestion pipeline, and detection tank with ammonia free water to remove the residue from the previous test.
2、 Calibration operation steps
1. Blank calibration (zero calibration)
Take ammonia free solution as a blank sample and inject it according to the "blank determination" process of the instrument: sequentially complete injection, digestion (usually at 120-124 ℃ for 30 minutes), cooling, neutralization, and colorimetric (measure absorbance at 220nm and 275nm wavelengths).
The instrument will automatically record the absorbance value of the blank, and use it as a reference to deduct the blank interference of subsequent standard samples and actual samples, completing the "zero point calibration". If the blank value is too high (such as absorbance>0.030), it is necessary to recheck for ammonia purity, reagent contamination, or instrument pipeline cleanliness.
2. Standard curve calibration (core steps)
In order of concentration from low to high, place the standard series solution into the instrument injection position, start the "standard curve measurement" program, and the instrument will automatically digest and compare each concentration point, recording the corresponding net absorbance value (220nm absorbance -2 × 275nm absorbance, eliminating organic interference).
The built-in software of the instrument will generate a standard curve through linear regression fitting based on the "concentration net absorbance" data (requiring a correlation coefficient R ² ≥ 0.999). If a certain concentration point deviates too far from the curve (such as residual>5%), it is necessary to re prepare the standard solution of that concentration and retest.
After the curve is generated, the instrument will save the curve parameters (slope, intercept), and directly call the curve to calculate the sample concentration during subsequent testing.
3. Single point calibration (auxiliary calibration, suitable for daily rapid verification)
When the instrument has saved the standard curve and only needs to confirm the accuracy of a certain concentration range, a standard solution close to the expected concentration of the sample (such as 2mg/L or 5mg/L standard solution if the sample is expected to be 3mg/L) can be selected for single point determination.
If the relative error between the concentration value measured by the instrument and the true concentration of the standard solution is ≤ ± 5%, it indicates that the curve is valid; If the error exceeds the range, it is necessary to recalibrate the standard curve.
3、 Verification after calibration
Quality control sample verification
Select a bottle of total nitrogen quality control sample with a known concentration (the concentration should be within the standard curve range and not overlap with the standard series concentration), measure it according to the sample detection process, and calculate the relative error between the measured value and the standard value of the quality control sample. If the error is ≤ ± 5%, it proves that the calibration is qualified; Otherwise, it is necessary to investigate issues such as standard solution preparation and instrument parameter settings, and recalibrate.
Parallel sample verification
Perform 2-3 parallel measurements on the same standard concentration point and calculate the relative standard deviation (RSD). If RSD ≤ 3%, it indicates that the instrument has good precision and the calibration results are reliable.
4、 Calibration precautions
Calibration frequency: The new instrument must be fully calibrated before its first use; For daily use, it is recommended to calibrate the standard curve every 7-15 days and perform a blank calibration after each startup; If the instrument is repaired (such as replacing the light source or digestion module) or the reagent batch is replaced, it is necessary to recalibrate the entire range.
Traceability of standard substances: Standard stock solutions with "National Standard Substance Certificates" must be used to avoid the use of self prepared non traceable standard solutions and ensure the standardization of calibration.
Environmental control: During the calibration process, maintain stable laboratory temperature (20-25 ℃) and humidity (40% -70%) to avoid strong light and vibration interference with instrument detection accuracy.
Record retention: Detailed records of calibration date, standard solution information (batch number, concentration), instrument parameters, standard curve data, quality control sample verification results, etc. Establish calibration files for traceability.