The Molybdenum Blue Method

4 Spectrophotometric determination of phosphate by the molybdenum blue method 

When orthophosphates in acidified solution react with ammonium molybdate, phosphomolybdic acid is formed. When this is reduced by a suitable reductant, an intense blue complex is formed. The absorbance at the wavelength of maximum absorption is proportional to the phosphate concentration. 

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This experiment uses the molybdenum-blue method to determine the concentration of phosphate in a phosphate/sodium chloride mixture. Elow-injection analysis is used to increase the speed of analysis, allowing students to... 

Phosphorus from organophosphorus compounds, which are combusted to give mainly orthophosphate, can be absorbed by either sulphuric acid or nitric acid and readily determined spectrophotometrically either by the molybdenum blue method or as the phosphovanadomolybdate (Section 17.39). 

Table 12.13 compares results obtained by this method with those obtained by a molybdenum blue method of spectrophotometry [115, 116]. Values obtained by atomic absorption spectrometry are higher than those obtained by the molybdenum blue method and this is believed to reflect the greater inherent accuracy of the former method. A UK standard method also... 

Matrix effects in the analysis of nutrients in seawater are caused by differences in background electrolyte composition and concentration (salinity) between the standard solutions and samples. This effect causes several methodological difficulties. First, the effect of ionic strength on the kinetics of colorimetric reactions results in color intensity changes with matrix composition and electrolyte concentration. In practice, analytical sensitivity depends upon the actual sample matrix. This effect is most serious in silicate analysis using the molybdenum blue method. Second, matrix differences can also cause refractive index interference in automated continuous flow analysis, the most popular technique for routine nutrient measurement. To deal with these matrix effects, seawater of... 

The advantages of high sensitivity, rapid analysis and simplicity of equipment are discussed, and the results for both types of sample material are compared with values obtained through use of the molybdenum blue method. 

The accelerating effect of US on the determination of phosphate by the Molybdenum Blue method was ascribed to depolymerization of molybdate, which was thought to speed up its reaction with phosphate and inorease the sensitivity [32]. 

Rigler F. H. (1968) Further observations inconsistent with the hypothesis that the molybdenum blue method measured orthophosphate in lake water. Limnol. Oceanogr. 13, 7-13. 

The single-point standard addition method was used in the determination of phosphate by the molybdenum blue method. A 2.00-mL urine sample was treated with molybdenum blue reagents to produce a species absorbing at 820 nm, after which the sample was diluted to 100 mL. A 25.00-mL aliquot of this solution gave an absorbance of 0.428 (solution 1). Addition of 1.00 mL of a solution containing 0.0500 mg of phosphate to a second 25.0-mL aliquot gave an absorbance of 0.517 (solution 2). Use these data to calculate the concentration of phosphate in milligrams per milliliter of the specimen. 

On-line monitoring of phosphate in natural water and effluent stream by the molybdenum blue method using sequential injection analysis has been described [1]. The detection limit of 0.5 mg r P04 was reported. 

Standard methods for phosphates, polyphosphates, and organic phosphates in environmental samples are predominantly nonchromatographic methods, which are based upon the molybdenum blue method. Within this colorimetric method ammonium molybdate and antimony potassium tartrate react under acidic conditions with dilute solution of phosphorous to form an antimony-phospho-molybdate complex which is then reduced to an intensely blue-colored complex by ascorbic acid. U.S. EPA Methods 365.1 to 365.4 are based upon this chemistry. 

A prerequisite for the molybdenum blue method is that all the arsenic has to be present as arsenate. After digestion with oxidizing acids, such as nitric acid, all the arsenic is converted into arsenate when appropriate heating time and temperatures are applied. The principle of this determination is the reaction of arsenate with ammonium molybdate in acidic medium to form an arsenate containing molybdenum heteropolyacid that can be reduced to molybdenum blue with stannous chloride, hydrazine, or ascorbic acid. Best results are obtained with hydrazine sulfate. The absorption maximum of the blue solution is between 840-860 nm (15). The most severe interferences for this method derive from phosphates and silicates. To remove interfering ions, distillation of arsenic as AsCb or AsBrs is often recommended (12,15). 

The AOCS method Ca 12-55 (AOCS, 1978) is based on the molybdenum blue method but uses zinc oxide instead of magnesium carbonate in the ashing step. The method differs somewhat from that described under Section 6.2.15 with respect to the details of the procedure. 

Various modifications of the colorimetric determination of phosphorus by the molybdenum blue method are available. The method described above has proved to be the most suitable. 

A similar method is described by Jarabin, Vajda, and Szarvas, except that the color is measured at 660 nm (317). Another modification of the molybdenum blue method is to develop the color in perchloric acid medium, by reduction with stannous ion and ascorbic acid (318). which eliminates interferences and gives a stable color. Kahler, Betz, and Betz (309) and Milton (319) favor reduction with sulfite ion for water analysis. 

For analyzing biological materials the molybdenum blue method is invariably-used because of the very low concentrations of silica. For this purpose a method of analyzing for traces of silica in the presence of iron, phosphorus, arsenic, and reducing substances was developed by Baumann (320). 

Another example is a fully computerized MSFIA system for the spectrophotometric determination of available phosphorus in soil extracts. The molybdenum blue method is chosen for the colorimetric determination, using ascorbic acid as reducing agent, antimony to accelerate the reduction to the blue complex and applying the Egner-Riehm method to extract phosphorus from soil samples. It presents a hnear calibration curve between 0.75 and 15 mg/1. A determination frequency of 15/h may be achieved, with good repeatability for 12 consecutive injections of soil extracts (RSD <1.7%). Results obtained from 12 soil samples were statistically comparable to those attained by the usual batch method [102]. 

In an attempt to integrate all the analytical process in a single device, a hyphenated MSFIA-microcolumn semp has been assembled for automated flow-through partitioning and accurate determination of the content of bioavaUable forms of orthophosphate in soils and sediments utilizing the molybdenum blue method for extract processing [103]. 

Sugawara, K. and Kanamori, S., Spectrophotometric determination of submicromolar quantities of orthophosphate in natural waters. Bulletin of the Chemical Society of japan 34,526,1961. Sjosten, A. and Blomqvist, S., Influence of phosphate concentration and reaction temperature when using the molybdenum blue method for determination of phosphate in water. Water Research 31,1818,1997. 

FIA was used to automate the determination of silicate in natural waters by the molybdenum blue method. The rate is 80 samples/h when a continuous stream of seawater is analyzed. Thirty discrete samples can be analyzed in duplicate per hour. The relative precision was better than 1% for silicate concentrations >10 pM. A detection limit of 0.5 pM Si was achieved. The refractive index interference was eliminated. This allows a... 

Procedure To a 1 L polyethylene separatory funnel, add 1050 mL sample water, 120 mg HF, and 0.5 mL of 200 mesh Amberlite IRA 402 (OH-type). Shake the furmel for 30 min, and filter the resin through a nylon cloth. Desorb the Si with 30 mL standard H3BO3 solution, and determine Si by the molybdenum blue method. 

Measurement of phosphate concentration using the molybdenum blue method... 

Phosphate in water may be determined according to a procedure outlined in [9], known as the "molybdenum blue method" It involves the complexation of phosphate with molybdate, with subsequent reduction of the complex with ascorbic acid The result is a complex having an intense blue color The overall reaction rate is limited by the complexation step, with maximum conversion of phosphate to the reduced complex requiring about 10 minutes This analytical procedure has been adapted by many groups for phosphate analysis in flow systems (see, for instance, [2, 10]) In one instance, a system was developed to monitor phosphate concentrations in fermentation broths [11] The flow manifold employed in that application is the model for the phosphate analysis using a stacked system described m this paper... 

The phosphate analysis described here is characterized by an analysis time of four minutes, when system flushing is included This is in contrast to the conventional flow injection analysis of phosphate using the molybdenum blue method reported in [2] and [10] These two studies indicate analysis times of 30 and 80 seconds, respectively Several factors play a role in the longer analysis times for the micromachmed system First is the incorporation of a time-based injection technique in the system, as opposed to the volume-defined methods using valves described in [2] and [10] While elimination of the need for an injection valve simplifies the system, time-based injection schemes by... 

Traces of arsenic may be determined either by the molybdenum blue or Gutzeit methods. For determination of arsenic contents from T5 to 15 //g in the sample taken the molybdenum blue method is recommended by the Analytical Methods Committee of the S.A,C. the details are as follows ... 

The molybdenum-blue method may be extended for determination of amounts of arsenic down to 0 5 / g (as As) if the reagents are specially purified by extraction with diethylammonium diethyldithiocarbamate solution before use. 

In the opinion of the Analytical Methods Committee of the S,A,C. the molybdenum-blue method is considered to have inherent advantages over the Gutzeit method, which may be regarded as an estimation rather than a determination, since it depends on the evaluation of the intensity of a stain on a test-paper and the judgments of individual analysts may differ slightly. Nevertheless, within this limitation and provided that the details of the method are strictly observed, the Gutzeit method is adequate in many circumstances, particularly when it is necessary to ascertain whether or not an arsenic content exceeds a certain limit. It is also less intricate and time-consuming than the molybdenum-blue method. 

The kinetics of each silica condensation reaction were quantified using the molybdenum blue method, which is sensitive to monomeric and dimeric silicic acid. Within the initial condensation time, there was a third-order linear relationship between [Si(OH)4] and time. As a result, it was concluded that polyamines with a threefold increase in their third-order rate constant were kmetically active. C2N2-4 and the naturally occurring polyamines spermidine and spermine observed no rate enhancement AH of the propylamines and longer-chain ethyleneamines significantly influenced condensation rates, with C3N7 having a 14-fold increase in catalytic activity. 

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