Part 1

STANDARD METHODS FOR THE EXAMINATION OF WATER AND SEWAGE

_FOURTH EDITION_

Revised by committees of the American Public Health Association, American Chemical Society, and referees of the Association of Official Agricultural Chemists

AMERICAN PUBLIC HEALTH ASSOCIATION 169 MASSACHUSETTS AVENUE BOSTON 1920

_Copyright, 1917 and 1920_

_By the American Public Health Association_

CONTENTS.

PAGE PREFACE TO THE FOURTH EDITION vii

COLLECTION OF SAMPLES 1 QUANTITY OF WATER REQUIRED FOR ANALYSIS 1 BOTTLES 1 TIME INTERVAL BETWEEN COLLECTION AND ANALYSIS 2 REPRESENTATIVE SAMPLES 3

PHYSICAL EXAMINATION 4 TEMPERATURE 4 TURBIDITY 4 TURBIDITY STANDARD 4 PLATINUM WIRE METHOD 5 TURBIDIMETRIC METHOD 7 COEFFICIENT OF FINENESS 8 COLOR 9 COMPARISON WITH PLATINUM-COBALT STANDARDS 9 COMPARISON WITH GLASS DISKS 10 COMPARISON WITH NESSLER STANDARDS 10 LOVIBOND TINTOMETER 11 ODOR 12 COLD ODOR 12 HOT ODOR 12 EXPRESSION OF RESULTS 12

CHEMICAL EXAMINATION 14 EXPRESSION OF RESULTS 14 FORMS OF NITROGEN 15 AMMONIA NITROGEN 15 DETERMINATION BY DISTILLATION 15 MEASUREMENT OF AMMONIA NITROGEN 16 COMPARISON WITH AMMONIA STANDARDS 16 COMPARISON WITH PERMANENT STANDARDS 17 MODIFICATION FOR SEWAGE 18 DETERMINATION BY DIRECT NESSLERIZATION 19 ALBUMINOID NITROGEN 20 ORGANIC NITROGEN 21 NITRITE NITROGEN 22 NITRATE NITROGEN 23 PHENOLDISULFONIC ACID METHOD 23 REDUCTION METHOD 24 TOTAL NITROGEN 25 OXYGEN CONSUMED 25 RECOMMENDED METHOD 26 OTHER METHODS 27 RESIDUE ON EVAPORATION 29 TOTAL RESIDUE 29 FIXED RESIDUE AND LOSS ON IGNITION 29 SUSPENDED MATTER 30 DETERMINATION WITH GOOCH CRUCIBLE 30 DETERMINATION BY FILTRATION 30 DETERMINATION OF VOLUME 30 FIXED RESIDUE AND LOSS ON IGNITION 30 HARDNESS 30 TOTAL HARDNESS BY CALCULATION 31 TOTAL HARDNESS BY SOAP METHOD 31 TOTAL HARDNESS BY SODA REAGENT METHOD 34 TEMPORARY HARDNESS BY TITRATION WITH ACID 34 NON-CARBONATE HARDNESS BY SODA REAGENT METHOD 34 NON-CARBONATE HARDNESS BY SOAP METHOD 35 ALKALINITY 35 PROCEDURE WITH PHENOLPHTHALEIN 36 PROCEDURE WITH METHYL ORANGE 37 PROCEDURE WITH LACMOID 37 PROCEDURE WITH ERYTHROSINE 37 BICARBONATE 37 NORMAL CARBONATE 38 HYDROXIDE 38 ALKALI CARBONATES 39 ACIDITY 39 TOTAL ACIDITY 40 FREE CARBON DIOXIDE 40 FREE MINERAL ACIDS 41 MINERAL ACIDS AND SULFATES OF IRON AND ALUMINIUM 41 CHLORIDE 41 IRON 43 TOTAL IRON 44 COLORIMETRIC METHOD 44 COMPARISON WITH IRON STANDARDS 45 COMPARISON WITH PERMANENT STANDARDS 46 VOLUMETRIC METHOD 46 DISSOLVED IRON 47 SUSPENDED IRON 47 FERROUS IRON 47 FERRIC IRON 48 MANGANESE 48 PERSULFATE METHOD 48 BISMUTHATE METHOD 49 LEAD, ZINC, COPPER, AND TIN 50 LEAD 51 ZINC 52 COPPER 53 TIN 54 MINERAL ANALYSIS 56 RESIDUE ON EVAPORATION 56 ALKALINITY AND ACIDITY 56 CHLORIDE 56 NITRATE NITROGEN 56 SEPARATION OF SILICA, IRON, ALUMINIUM, CALCIUM, AND MAGNESIUM 56 SILICA 56 IRON AND ALUMINIUM 57 CALCIUM 57 MAGNESIUM 57 SEPARATION OF SULFATE, SODIUM, AND POTASSIUM 58 SULFATE 58 SODIUM, POTASSIUM AND LITHIUM 58 POTASSIUM 59 LITHIUM 60 BROMINE, IODINE, ARSENIC, AND BORIC ACID 61 BROMINE AND IODINE 61 ARSENIC 63 BORIC ACID 63 HYDROGEN SULFIDE 63 CHLORINE 64 DISSOLVED OXYGEN 65 ETHER-SOLUBLE MATTER 69 RELATIVE STABILITY OF EFFLUENTS 69 BIOCHEMICAL OXYGEN DEMAND OF SEWAGES AND EFFLUENTS 71 RELATIVE STABILITY METHOD 71 SODIUM NITRATE METHOD 72

ANALYSIS OF SEWAGE SLUDGE AND MUD DEPOSITS 73 COLLECTION OF SAMPLE 73 REACTION 73 SPECIFIC GRAVITY 74 MOISTURE 74 VOLATILE AND FIXED MATTER 74 TOTAL ORGANIC NITROGEN 74 ETHER-SOLUBLE MATTER 75 FERROUS SULFIDE 76 BIOCHEMICAL OXYGEN DEMAND 76

ANALYSIS OF CHEMICALS 77 REAGENTS 77 SULFATE OF ALUMINIUM 78 INSOLUBLE MATTER 78 OXIDES OF ALUMINIUM AND IRON 78 TOTAL IRON 79 FERRIC IRON 79 FERROUS IRON 80 BASICITY RATIO 80 LIME 80 SULFATE OF IRON 81 INSOLUBLE MATTER 81 IRON AS FERROUS SULFATE 81 ACIDITY 81 SODA ASH 82 INSOLUBLE MATTER 82 AVAILABLE ALKALI 82

CHEMICAL BIBLIOGRAPHY 82

MICROSCOPICAL EXAMINATION 89 MICROSCOPICAL BIBLIOGRAPHY 91

BACTERIOLOGICAL EXAMINATION 92 APPARATUS 92 SAMPLE BOTTLES 92 PIPETTES 92 DILUTION BOTTLES 92 PETRI DISHES 92 FERMENTATION TUBES 92 MATERIALS 93 WATER 93 MEAT EXTRACT 93 PEPTONE 93 SUGARS 93 AGAR 93 GELATIN 93 LITMUS 93 GENERAL CHEMICALS 93 METHODS 93 PREPARATION OF CULTURE MEDIA 93 TITRATION 93 STERILIZATION 94 NUTRIENT BROTH 95 SUGAR BROTHS 95 NUTRIENT GELATIN 95 NUTRIENT AGAR 96 LITMUS OR AZOLITMIN SOLUTION 96 LITMUS-LACTOSE-AGAR 97 ENDO’S MEDIUM 97 COLLECTION OF SAMPLE 98 STORAGE AND TRANSPORTATION OF SAMPLE 98 DILUTIONS 98 PLATING 99 INCUBATION 99 COUNTING 99 THE TEST FOR THE PRESENCE OF MEMBERS OF THE B. COLI GROUP 100 PRESUMPTIVE TEST 100 PARTIALLY CONFIRMED TEST 101 COMPLETED TEST 102 APPLICATION OF THESE TESTS 102 EXPRESSION OF RESULTS 103 SUMMARY OF THESE TESTS 104 INTERPRETATION OF RESULTS 106 DIFFERENTIATION OF FECAL FROM NON-FECAL MEMBERS OF THE B. COLI GROUP 106 METHYL RED TEST 107 VOGES-PROSKAUER TEST 108 ROUTINE PROCEDURE FOR BACTERIOLOGICAL EXAMINATION 108 BACTERIOLOGICAL BIBLIOGRAPHY 110

INDEX 113

PREFACE TO FOURTH EDITION.

The Committee on Standard Methods of Bacteriological Water Analysis was reorganized in 1918 with the following membership: F. P. Gorham, chairman, L. A. Rogers, W. G. Bissell, H. E. Hasseltine, H. W. Redfield, with M. Levine as adjunct member. This committee made a report in 1918 which was not acted on by the Laboratory Section, and in 1919 made a revised report, recommending certain changes in Standard Methods, which were adopted by the section and which are now incorporated in this present fourth edition.

Following are the more important changes:

New brands of peptone authorized.

Phenol Red Method of Hydrogen-ion Concentration.

Five-tenths per cent of sugar specified for broths instead of 1 per cent.

Sterilization of sugar is media specified in greater detail.

Preparation of Endo Medium.

Synthetic Medium for the Methyl Red Test.

There are no changes in the chemical methods in this edition.

AMERICAN PUBLIC HEALTH ASSOCIATION. _LABORATORY SECTION._ STANDARD METHODS FOR THE EXAMINATION OF WATER AND SEWAGE.

Compiled and revised by committees of the American Public Health Association and the American Chemical Society and referees of the Association of Official Agricultural Chemists.

COLLECTION OF SAMPLES.

QUANTITY REQUIRED FOR ANALYSIS.

The minimum quantity necessary for making the ordinary physical, chemical, and microscopical analyses of water or sewage is 2 liters; for the bacteriological examination, 100 cc. In special analyses larger quantities may be required.

BOTTLES.

The bottles for the collection of samples shall have glass stoppers, except when physical, mineral, or microscopical examinations only are to be made. Jugs or metal containers shall not be used.

Sample bottles shall be carefully cleansed each time before using. This may be done by treating with sulfuric acid and potassium bichromate, or with alkaline permanganate, followed by a mixture of oxalic and sulfuric acids, and by thoroughly rinsing with water and draining. The stoppers and necks of the bottles shall be protected from dirt by tying cloth, thick paper or tin foil over them.

For shipment bottles shall be packed in cases with a separate compartment for each bottle. Wooden boxes may be lined with corrugated fibre paper, felt, or similar substance, or provided with spring corner strips, to prevent breakage. Lined wicker baskets also may be used.

Bottles for bacteriological samples shall be sterilized as directed on page 98.

INTERVAL BEFORE ANALYSIS.

In general, the shorter the time elapsing between the collection and the analysis of a sample the more reliable will be the analytical results. Under many conditions analyses made in the field are to be commended, as data so obtained are frequently preferable to data obtained in a distant laboratory after the composition of the water has changed.

The time that may be allowed to elapse between the collection of a sample and the beginning of its analysis cannot be stated definitely. It depends on the character of the sample, the examinations to be made, and other conditions. The following are suggested as fairly reasonable maximum limits.

_Physical and chemical analysis._

Ground waters 72 hours Fairly pure surface waters 48 " Polluted surface waters 12 " Sewage effluents 6 " Raw sewages 6 "

_Microscopical examination._

Ground waters 72 hours Fairly pure surface waters 24 " Waters containing fragile organisms Immediate examination

_Bacteriological examination._

Samples kept at less than 10°C 24 hours

If a longer period elapses between collection and examination the time should be noted. If sterilized by the addition of chloroform, formaldehyde, mercuric chloride, or some other germicide samples for sanitary chemical examination may be allowed to stand for longer periods than those indicated, but as this is a matter which will vary according to circumstances, no definite procedure is recommended. If unsterilized samples of sewage, sewage effluents, and highly polluted surface waters are analyzed after greater intervals than those suggested caution must be used in interpreting analyses of the organic content, which frequently changes materially upon standing.

Determinations of dissolved gases, especially oxygen, hydrogen sulfide, and carbon dioxide, should be made at the time of collection in order to be reasonably accurate, in accordance with the directions given hereafter in connection with each determination.

REPRESENTATIVE SAMPLES.

Care should be taken to obtain a sample that is truly representative of the liquid to be analyzed. With sewages this is especially important because marked variations in composition occur from hour to hour. Satisfactory samples of some liquids can be obtained only by mixing together several portions collected at different times or at different places—the details as to collection and mixing depending upon local conditions.

PHYSICAL EXAMINATION.

TEMPERATURE.

The temperature of the sample, if taken, shall be taken at the time of collection, and shall be expressed preferably in degrees Centigrade, to the nearest degree, or closer if more precise data are required. The thermophone[109] is recommended for obtaining the temperature of water at various depths below the surface.

TURBIDITY.

The turbidity of water is due to suspended matter, such as clay, silt, finely divided organic matter, microscopic organisms, and similar material.

TURBIDITY STANDARD.[110]

The standard of turbidity shall be that adopted by the United States Geological Survey, namely, a water which contains 100 parts per million of silica in such a state of fineness that a bright platinum wire 1 millimeter in diameter can just be seen when the center of the wire is 100 millimeters below the surface of the water and the eye of the observer is 1.2 meters above the wire, the observation being made in the middle of the day, in the open air, but not in sunlight, and in a vessel so large that the sides do not shut out the light so as to influence the results. The turbidity of such water is arbitrarily fixed at 100 parts per million.

For preparation of the silica standard dry Pear’s “precipitated fuller’s earth” and sift it through a 200–mesh sieve. One gram of this preparation in 1 liter of distilled water makes a stock suspension which contains 1,000 parts per million of silica and which should have a turbidity of 1,000. Test this suspension, after diluting a portion of it with nine times its volume of distilled water, by the platinum wire method to ascertain if the silica has the necessary degree of fineness and if the suspension has the necessary degree of turbidity. If not, correct by adding more silica or more water as the case demands.[A]

Footnote A:

This method of correction very slightly alters the coefficient of fineness of the standard, but does not noticeably affect its use.

Standards for comparison shall be prepared from this stock suspension by dilution with distilled water. For turbidity readings below 20, standards of 0, 5, 10, 15, and 20 shall be kept in clear glass bottles of the same size as that containing the sample; for readings above 20, standards of 20, 30, 40, 50, 60, 70, 80, 90, and 100 shall be kept in 100 cc. Nessler tubes approximately 20 millimeters in diameter.

Comparison with the standards shall be made by viewing both standard and sample sidewise toward the light by looking at some object and noting the distinctness with which the margins of the object can be seen.

The standards shall be kept stoppered, and both sample and standards shall be thoroughly shaken before making the comparison.

In order to prevent any bacterial or algal growths from developing in the standards a small amount of mercury bichloride may be added to them.

PLATINUM WIRE METHOD.[42]

This method requires a rod with a platinum wire 1 mm. in diameter inserted in it about 1 inch from one end of the rod and projecting from it at a right angle at least 25 mm. Near the other end of the rod, at a distance of 1.2 meters from the platinum wire, a small ring shall be placed directly above the wire through which, with his eye directly above the ring, the observer shall look when making the examination.

The rod shall be graduated as follows: The graduation mark of 100 shall be placed on the rod at a distance of 100 mm. from the center of the wire. Other graduations shall be made according to Table 1, which is based on the best obtainable data. The distances recorded in Table 1 are intended to be such that when the water is diluted the turbidity readings will decrease in the same proportion as the percentage of the original water in the mixture. These graduations are those on what is known as the U. S. Geological Survey Turbidity Rod of 1902.[105]