HJ 1216-2021 English PDFUS$379.00 · In stock
Delivery: <= 4 days. True-PDF full-copy in English will be manually translated and delivered via email. HJ 1216-2021: Water quality - Determination of phytoplankton - 0.1 ml chamber - Microscope counting method Status: Valid
Basic dataStandard ID: HJ 1216-2021 (HJ1216-2021)Description (Translated English): Water quality - Determination of phytoplankton - 0.1 ml chamber - Microscope counting method Sector / Industry: Environmental Protection Industry Standard Word Count Estimation: 16,136 Issuing agency(ies): Ministry of Ecology and Environment HJ 1216-2021: Water quality - Determination of phytoplankton - 0.1 ml chamber - Microscope counting method---This is a DRAFT version for illustration, not a final translation. Full copy of true-PDF in English version (including equations, symbols, images, flow-chart, tables, and figures etc.) will be manually/carefully translated upon your order. (Water quality Determination of phytoplankton 0.1 ml counting box-microscopic counting method) National Ecological Environment Standard of the People's Republic of China Determination of Phytoplankton in Water Quality 0.1 ml Counting Box- Microscopic counting Water quality - Determination of phytoplankton - 0.1 ml chamber - Microscope counting method This electronic version is the official standard text, which is reviewed and typeset by the Environmental Standards Institute of the Ministry of Ecology and Environment. Published on 2021-11-29 2022-06-01 Implementation Released by the Ministry of Ecology and Environment directory Foreword...ii 1 Scope...1 2 Normative references...1 3 Terms and Definitions...1 4 Principles of the method...1 5 Reagents and materials...2 6 Instruments and equipment...2 7 Samples...2 8 Analysis steps...3 9 Result calculation and representation...5 10 Precision...6 11 Quality Assurance and Quality Control...6 12 Notes...6 13 Waste Disposal...7 Appendix A (Normative Appendix) Calculation of the detection limit of the random field of view method...8 Appendix B (Informative Appendix) Measurement and Calculation of Microscope Field of View Area...9 Appendix C (informative appendix) Phytoplankton count original record table...12 References...13 Determination of Phytoplankton in Water Quality 0.1 ml Counting Box - Microscope Counting Method 1 Scope of applicationThis standard specifies the 0.1 ml counting box-microscopic counting method for the determination of phytoplankton in water. This standard applies to the density determination of phytoplankton in surface water. When the sample is concentrated 50 times, the detection limit of the diagonal counting method is 9.2×103 cells/L; the detection limit of the row counting method is 3.0×103 cells/L; the detection limit of the whole film counting method is 9.2×102 cells/L; the detection limit and observation of the random field counting method It is related to the number of fields of view and the field of view area of the microscope, and is calculated according to Appendix A.2 Normative referencesThis standard refers to the following documents or clauses thereof. For dated references, only the dated version applies to this standard. For undated references, the latest edition (including all amendments) applies to this standard. GB/T 14581 Technical Guidelines for Sampling of Water Quality in Lakes and Reservoirs HJ/T 91 Technical Specification for Surface Water and Sewage Monitoring HJ 494 Water Quality Sampling Technical Guide3 Terms and DefinitionsThe following terms and definitions apply to this standard. 3.1 phytoplankton The tiny algae that live planktonic life in water, usually phytoplankton are planktonic algae, including prokaryotic cyanobacteria and other types of eukaryotes algae. 3.2 microscope counting field A defined area in the microscope field of view used to quantitatively count phytoplankton. 3.3 detection limit During a single count, the probability of discovery is not lower than the lowest phytoplankton density at 99%.4 Principles of the methodUnder the microscope, manually classify and count the phytoplankton in the sample using a 0.1 ml counting box, and calculate the unit volume of the sample. Cell numbers of various phytoplankton species.5 Reagents and MaterialsUnless otherwise stated, analytical reagents that meet national standards were used in the analysis, and the experimental water was freshly prepared deionized water or distilled water. 5.1 Iodine (I2). 5.2 Potassium iodide (KI). 5.3 Formaldehyde solution. w(HCHO)=37%～40%. 5.4 Glycerol (HOCH2CHOHCH2OH). 5.5 Lugol's iodine solution. Weigh 60 g of potassium iodide (5.2), dissolve it in 100 ml of water, add 40 g of iodine (5.1), stir well to dissolve it completely, add water Dilute to 1000 ml, transfer to a brown ground-mouth glass bottle, and store at room temperature away from light.6 Instruments and equipment6.1 No. 25 plankton net. the diameter of the mesh is 0.064 mm, the net is conical, the net mouth is set on the copper ring, and there is a water outlet switch at the bottom of the net. plug. 6.2 Qualitative sampling bottle. 30 ml ~ 100 ml wide-mouth polyethylene bottle. 6.3 Water collector. stainless steel or plexiglass material, cylindrical. Capacity and depth specifications to meet sampling requirements. 6.4 Quantitative sampling bottle. 1L～2L wide-mouth polyethylene bottle. 6.5 Upright or inverted biological microscope. objective lens 4×, 10×, 20×, 40×, eyepiece 10× or 15×. 6.6 Concentrating device. 1 L ~ 2 L cylindrical separatory funnel or measuring cylinder. 6.7 Vials. 50 ml amber glass jars with stoppers. 6.8 Ultrasonic generating device. working frequency 40 kHz, water bath mode. 6.9 Micropipette. 100 μl. 6.10 0.1 ml phytoplankton counting frame. the area is 20 mm×20 mm, and the frame is divided into 10 horizontal and vertical grids, with a total of 100 small grids. 6.11 Cover glass. the area is 22 mm × 22 mm, and the thickness is less than 0.2 mm. 6.12 Stage micrometer. also known as stage micrometer, 1 mm/100 DIV, with a division value of 0.01 mm. Note. DIV refers to equal division, 1 mm/100 DIV means that 1 mm is divided into 100 divisions. 6.13 Eyepiece reticle. also known as eyepiece micrometer, 5 mm/50 DIV, with a division value of 0.1 mm. 6.14 Counters. 6.15 Common laboratory instruments and equipment.7 samples7.1 Collection of samples 7.1.1 Qualitative samples The point layout and sampling frequency shall be implemented in accordance with the relevant regulations of GB/T 14581, HJ/T 91 and HJ 494.can also be based on research ok. Collect qualitative samples using plankton net 25 (6.1). Turn off the water outlet piston switch at the bottom of the plankton net, at the surface of the water to 0.5 m In the depths, reciprocate in an "∞" shape at a speed of 20 cm/s to 30 cm/s, and drag slowly for about 1 to 3 minutes, until there are obvious phytoplankton in the net. After entering, lift the plankton net (6.1) out of the water surface, the water in the net is naturally filtered out through the mesh holes, and there is a little water sample (5 ml ~ 10 ml) left at the bottom. , move the bottom outlet into the qualitative sampling bottle (6.2), and open the bottom piston switch to collect qualitative samples. When collecting layered samples, use 25 No. plankton net (6.1) to filter specific water layer samples, and other steps are the same as collecting surface samples. After the qualitative sample collection is completed, the planktonic Clean the bio-net. The samples were refrigerated and transported away from light after collection. If there are technical specifications for qualitative sample collection, follow the relevant requirements of the technical specifications. 7.1.2 Quantitation of samples Quantitative sample collection shall be carried out in accordance with the relevant provisions of GB/T 14581, HJ/T 91 and HJ 494. Use the water collector (6.3) to collect the sample into the quantitative sampling bottle (6.4), generally not less than 500 ml of sample. If the water body is more transparent When the number of phytoplankton is high and the number of phytoplankton is small, the sampling volume should be increased as appropriate. After quantitative sample collection, the vial should not be full so that it can be shaken well. Note 1.Some phytoplankton (such as blue-green algae) often float on the water surface or are distributed in sheets and strips, so sampling can be taken as a peak reference in the dense area of algal blooms. Note 2.Quantitative sample collection should precede qualitative sample collection. Fixed time period sampling should be maintained so that results can be compared with each other. 7.2 Storage of samples 7.2.1 Qualitative samples Immediately after the qualitative samples were collected, Lugol's iodine solution (5.5) was added in an amount of 1.0% to 1.5% of the volume of the water sample. Microscopic biopsy samples are not added Lugol's iodine solution. Qualitative samples can be stored for 3 weeks at room temperature in the dark; 12 months in the refrigerator at 1 ℃～5 ℃ in the dark. In vivo samples can be stored for 36 h at 4 ℃～10 ℃ in the dark. 7.2.2 Quantitation of samples Immediately after the quantitative sample collection, add Lugol's iodine solution (5.5) to fix it, and the dosage is 1.0% to 1.5% of the volume of the water sample. Lugol's The iodine solution (5.5) is added to the quantitative sampling bottle (6.4) in advance and brought to the site for use. Quantitative samples can be stored for 3 weeks at room temperature in the dark; 1 ℃～ It can be stored for 12 months at 5 ℃ in the dark. During the storage process of the sample, the oxidation degree of Lugol's iodine solution (5.5) should be checked every week. Add an appropriate amount of Lugol's iodine solution (5.5) until the color of the sample returns to yellow-brown. Note. If the sample needs to be stored for a long time, formaldehyde solution (5.3) should be added in an amount of 4% of the volume of the water sample.8 Analysis steps8.1 Mixing the sample Before each sampling, thoroughly mix the sample by inverting it upside down at least 30 times, and the mixing action should be light. 8.2 Analysis of qualitative samples Observe the qualitative sample (7.1.1) under the microscope (6.5) to identify the phytoplankton species. Predominant species identified to species, other species to The genus is rarely identified. Some species identification references can be found in ref. Note. In addition to qualitative samples for species identification, quantitative samples that have been counted can also be used for observation. 8.3 Analysis of quantitative samples 8.3.1 Specimen preparation 8.3.1.1 Preflight Bring the sample to room temperature, take 0.1 ml of the sample with a micropipette (6.9) and mix it, and pour it into the 0.1 ml phytoplankton counting box (6.10) , completely cover the counting frame (6.10) with a cover glass (6.11), let it stand for a while, and the sample can be observed without air bubbles. Sample. Randomly select a number of counting cells or fields of view to preliminarily estimate the number of phytoplankton. For phytoplankton samples containing aggregated cells, ultrasonic analysis should be performed when either of the following two conditions are not met. Scatter processing. a) Individual phytoplankton cells in the colony can be easily identified, and the cells in the colony can be counted; b) When the number of cells contained in the colony has a fixed ratio to the volume or length of the colony, such as algae, algae, filamentous algae, etc., it can be Taking the population as the counting object, the number of phytoplankton cells was obtained according to the ratio. 8.3.1.2 Adjusting phytoplankton density The suitable phytoplankton density is 107 cells/L～108 cells/L. If the phytoplankton cell density in the quantitative sample (7.1.2) is low At 107 cells/L, the sample should be concentrated; if the phytoplankton cell density in the quantitative sample (7.1.2) is higher than 108 cells/L, the sample should be diluted. Finally, the 0.1 ml sample added to the counting box contains about 500 to 10,000 phytoplankton cells. Sample concentration. Shake all the quantitative samples and pour them into the concentration device (6.6), avoid direct sunlight, and let stand for 48 hours. use Transfer the supernatant to a beaker with a fine siphon until the phytoplankton sediment volume is about 20 ml. Unscrew the bottom piston of the concentrator (6.6), Place the phytoplankton sediment in a 100 ml graduated cylinder. Rinse the concentration device (6.6) 1 to 3 times with a little supernatant, and put the rinse water into the In a graduated cylinder, use the supernatant to make up the volume to the desired concentration multiple. In order to reduce the adsorption of phytoplankton on the wall of the concentration device, in the initial stage of standing, The walls of the concentrator should be tapped at appropriate times. During the siphoning process, the distance between the suction port and the phytoplankton sediment should be greater than 3 cm. floating in water The plant density is extremely low, and when the sampling volume is 1 L or more, it can be concentrated several times, that is, after each concentration, it can be left standing for 24 h to 48 h, and the concentration operation can be repeated. Adjust to the desired concentration fold volume. Concentrated samples can be counted after sonication as needed. Sample dilution. According to the dilution ratio, select a volumetric flask of the corresponding volume, and measure a quantitative sample of no less than 25 ml after mixing or ultrasonically. Disperse the treated sample and make up to the mark with water. If you want to save the diluted sample, you should add Lugol's iodine solution (5.5) to make the dilution The concentration of Lugol's iodine solution in the samples after dilution was the same as before dilution. Note. The specific steps of ultrasonic dispersion treatment are to take the mixed quantitative sample into the sample bottle (6.7), and use the ultrasonic generator (6.8) for about 10 minutes. Observe under the microscope (6.5), if there are still a large number of undispersed cell clusters, the ultrasonic treatment time should be prolonged until accurate counting can be achieved. overtake In the process of sonic dispersion treatment, attention should be paid to the water temperature to prevent water evaporation and damage to the phytoplankton cell structure caused by overheating. 8.3.2 Microscope counting 8.3.2.1 Loading Load the slides in the same way as in 8.3.1.1.If necessary, use a dropper to pipette a little glycerol (5.4) to spread evenly around the coverslip to prevent Stop the water in the counting box from evaporating to form air bubbles. When applying, avoid bleeding into the counting box. 8.3.2.2 Select counting method According to the density of phytoplankton in the adjusted sample (8.3.1.2), select an appropriate counting method to make the phytoplankton in the measurement process The total number of cells is 500 to 1500 cells. Table 1 shows the recommended counting methods. Table 1 Recommended counting methods Number of phytoplankton cells (cells) contained in a 0.1 ml sample in the counting box Recommended method of counting 500～1500 Whole slice count 1500～5000 row count 5000～10000 diagonal count, random field of view 8.3.2.3 Counting 8.3.2.3.1 Whole slice counting method Under the 40× objective lens, observe all 100 small squares in the phytoplankton counting frame one by one, and classify and count all the plankton in each small square. Plant cells, and record the differential count results for each row. If the phytoplankton cell volume is large, the objective lens magnification can be reduced. 8.3.2.3.2 Row counting method Under a 40× objective lens, observe the 2nd, 5th, and 8th rows of the phytoplankton counting box one by one, a total of 30 small squares, and count each small square by classification All phytoplankton cells in the cell were recorded, and the results of the classification count of each cell were recorded. If the phytoplankton cell volume is large, the objective lens magnification can be reduced. 8.3.2.3.3 Diagonal counting method Under the 40× objective lens, observe the 10 small squares located on the diagonal position of the phytoplankton counting frame one by one, and count each small square by classification All phytoplankton cells in the cell were recorded, and the results of the classification count of each cell were recorded. If the phytoplankton cell volume is large, the objective lens magnification can be reduced. 8.3.2.3.4 Random field of view method Under a 40× objective lens, randomly select a certain number of fields of view, classify and count all phytoplankton cells in each field of view, and record each Classification count results of visual fields. If the phytoplankton cell volume is large, the objective lens magnification can be reduced. The microscope field of view should be measured or calculated before counting See Appendix B for area, measurement and calculation methods. 8.3.2.3.5 Counting requirements Each sample was loaded and counted twice. The relative deviation of the total number of phytoplankton cells in the two times should be within ±15%, otherwise the calculation should be increased. Count once until the result of two counts meets this requirement. The measurement result is the result of two counts with a relative deviation within ±15% average value.9 Result calculation and presentation9.1 Result calculation The cell density of phytoplankton in the sample is calculated according to formula (1). N= Ac V1 V0 × 1000 (1) In the formula. N—the density of phytoplankton in the sample, cells/L; A--Counting frame area, mm2; Ac--counting area, when the counting method is diagonal, row and full slice, the counting area is A/10, 3A/10 and A, respectively. When the counting method is random field of view, the counting area is the total field of view area, mm2; n--the number of phytoplankton cells counted by microscopy, cells; V--count box volume, ml; V1--diluted or concentrated sample volume, ml; V0--sample volume before dilution or concentration, ml; 1000--volume conversion factor, ml/L. 9.2 Result representation The measurement results are expressed in scientific notation, with 2 significant figures reserved. 10 Precision The phytoplankton density levels were determined by whole-sheet count, row count, diagonal count, and random field count in 6 laboratories, respectively. Lake samples of 1 × 107 cells/L, 3 × 107 cells/L, 5 × 107 cells/L, and 1 × 108 cells/L were tested in 7 replicates, laboratory The intra-laboratory relative standard deviations were 2.4%-11%, 2.9%-10%, 2.7%-11%, 4.8%-8.2%; the inter-laboratory relative standard deviations The differences were 22%, 5.6%, 7.1%, and 21%, respectively. Calculate the 95% confidence interval of the logarithm of the measurement result, and then take the logarithm to obtain the laboratory The 95% confidence interval between them is shown in Table 2. 11 Quality Assurance and Quality Control 11.1 Phytoplankton uniformity The uniformity of phytoplankton distribution in the counting frame should be confirmed before starting the microscopic count. Using a low magnification objective to observe phytoplankton in The distribution of the entire counting box, if the distribution is not uniform, should be resampled. 11.2 Minimum count In a single assay, the total number of phytoplankton cells is not less than 500.If the measurement accuracy is difficult to meet the requirements, an appropriate increase in each The total count of phytoplankton cells in each assay. 12 Notes 12.1 If a part of a phytoplankton cell is in the row or field of view and the other part is outside the row or field of view, then the Cells in the border and left border or upper half of the field of view were not counted, and cells in the lower and right border of the row or the lower half of the field of view were counted. damaged cells Or cell residues are not counted. 12.2 During the counting process, if the sample water evaporates and bubbles are formed in the counting frame, discard the film and re-sample and co......Tips & Frequently Asked Questions:Question 1: How long will the true-PDF of HJ 1216-2021_English be delivered?Answer: Upon your order, we will start to translate HJ 1216-2021_English as soon as possible, and keep you informed of the progress. The lead time is typically 2 ~ 4 working days. The lengthier the document the longer the lead time.Question 2: Can I share the purchased PDF of HJ 1216-2021_English with my colleagues?Answer: Yes. The purchased PDF of HJ 1216-2021_English will be deemed to be sold to your employer/organization who actually pays for it, including your colleagues and your employer's intranet.Question 3: Does the price include tax/VAT?Answer: Yes. Our tax invoice, downloaded/delivered in 9 seconds, includes all tax/VAT and complies with 100+ countries' tax regulations (tax exempted in 100+ countries) -- See Avoidance of Double Taxation Agreements (DTAs): List of DTAs signed between Singapore and 100+ countriesQuestion 4: Do you accept my currency other than USD?Answer: Yes. If you need your currency to be printed on the invoice, please write an email to Sales@ChineseStandard.net. In 2 working-hours, we will create a special link for you to pay in any currencies. Otherwise, follow the normal steps: Add to Cart -- Checkout -- Select your currency to pay. |
