3.2.9.3. Turbidimetry

Method and Instrumentation

There are two differences between the instrumental setups for nephelometry and turbidimetry:

1) The flow cell for turbidimetry (K, left) is constructed from 10cm long 0.8 mm I.D. straight tubing made of green PEEK. It serves as a collimator directing a beam of light from the tungsten halogen lamp (THL) atop the tubing into the CCD detector on the bottom of the flow cell. In this way, 50 μL of suspension within the flow cell is monitored in absorbance mode analogous to spectrophotometry. Therefore, the “visible light” (400 to 800 nm), absorbed by the material in the flow cell yields an absorbance A related, in this work, to NTU via calibration with the standard. (Construction of this Long Light Path flow cell is in Sec. 3.2.8.) [1]

2) The flow protocol (K, right) comprises three parts: autocalibration [2] with NTU standard solutions, transport of NTU suspension into the flow cell, and monitoring of turbidity during a 15 sec STOP flow period, on a selected section of bead suspension that has been arrested within the flow cell. Therefore, volume and flow rate of carrier solution (pump 2) during the second step, are chosen to position the centroid of the sample zone into the middle of flow cell, to maximize sensitivity of determination (Detailed explanation is in Sec. 3.2.3.). The flow protocol is executed by software program (L) that also performs data collection and processing.

 

 

 

Since the spectrophotometer will monitor radiation absorbed by suspended particles, it has to be set up to function in absorbance mode (M).

Results

Data obtained by autocalibration of 20 NTU Formazin standard prepared in DI water are presented in three formats: spectra versus concentration (N), time versus concentration (O), and resulting calibration graph (P).

 

Over the entire spectrum range, the absorbance A is directly proportional to the concentration reaching maximum at 450 nm (N), when the signal becomes noisy due to the diminishing intensity of the THL source in this range. Interestingly, this arbitrary peak is not specific for Formazin since it is also present on turbidity spectra of particles of Suspended Matter. We decided to monitor absorbance at 550 nm because the THL light is most intense at this wavelength and because this wavelength complies with the EPA norm, however wavelengths up to 850nm would be suitable.

It follows that ,in variance with common practice (EPA, ISO) conditions of turbidimetric determination should NOT be specified by wavelength produced by a light source, but in analogy with spectrophotometry, should be always specified by the wavelength of monitored radiation.

Absorbance values versus time (O) increase sharply from baseline (BS) at the 13 seconds mark as the leading edge of suspension enters the flow cell from below. Upon reaching a maximum at 22 seconds, the flow is stopped for 15 seconds during which the suspension is arrested in the flow cell. Since during this period the absorbance (A) seems to decrease, the data collecting window (WIN) used to construct the calibration graph (P) is placed towards the end of the stop flow period. As the flow resumes, suspension is flushed from the flow cell and absorbance reaches baseline.

The calibration graph (P) in the range 0 to 20 NTU is strictly linear, of excellent reproducibility resulting in low limit of detection (LOD) and low blank values making stop flow turbidimetry well suited for the determination of suspended matter (SPM) in sea water. However, the influence of salinity of SW must now be investigated.

Sea Water calibrations for turbidimetry were obtained by using Simulated Sea Water made of 3.5% NaCl in DI, which approximates salinity in open ocean water (35 PSU). Therefore, the 3.5% NaCl was used to prepare a 20 NTU standard and was used as the carrier supplied by pump 2. In this way, the SSW based formazin standard was stepwise diluted with SSW. The data obtained by SSW calibration are presented in three formats: 1) spectra versus concentration (R), 2) time versus concentration (S), and 3) resulting calibration graph (T).

The spectra of SSW based calibration (R left) looks identical to the spectra obtained with DI water (N), while closer examinations of SSW and DI spectra (R right) show no difference in A at 550 nm.. 

There is, however, a large difference between time/concentration plots, recorded with formazin standards prepared in DI (O) and SSW (S). These differences are caused by difference in Refractive Index between DI (carrier pump 1) and SW (carrier pump 2 and sample), that distorts absorbance A in response to the schlieren effect. However, by placing the data collection window WIN at a 30 to 33 seconds positions, well away from the refractive index position, its interference is eliminated. Therefore, DI and SW based calibration data (P and T) are identical, while the slopes of DI/SW calibration lines differ only by 2%.

TAKEOUT

Turbidimetry in stop flow programmable format yields, with Formazin standards, well reproducible data, strictly linear calibration lines which differ by only by 2% between fresh water of 0 PSU and the highest salinity water of 35 PSU in the open ocean. Therefore in DI calibration can be used for analysis of SW samples collected in estuaries where salinity varies daily and seasonally. 
While both turbidimetry and nephelometry are, surprisingly, equally sensitive (compare LOD and BLANK of Figs. P with H), the ultimate test will be linearity, reproducibility and LOD of assays of as diverse materials as Suspended Matter in sea water, drinking water or of heterogenous solutions such as PhosphoMolybdenum Blue or SilicaMolybdenum Green on which spectrfotometric analysis of soluble phosphate [3] and silicate [4] are based.

References:

[1] Mariko Hatta, Jaromir Ruzicka, Christopher I. Measures “The performance of a new linear light path flow cell is compared with a liquid core waveguide and the linear cell is used for spectrophotometric determination of nitrite in sea water at nanomolar concentrations. “ Talanta 219 (2020) 1212
[2] Mariko Hatta, Jaromir Ruzicka, Christopher I. Measures, Madeline Davis “Autocalibration using a single standard  solution prepared in deionized water is applied to determination of nutrients in sea water by programmable flow injection “ Talanta 253(2023) 124041
[3] E.A. Nagul, I.A. McKevie, P. Worsfold, S.. Kolev “The molybdenum blue reaction for the determination of orthophosphate revisited: Opening the black box” Anal. Chim. Acta. 890 (2015) 60-82
[4]  J. Ruzicka, G. D. Marshall, Ch. Measures, M. Hatta“ Flow injection programmed to function in batch mode is used to determine molar absorptivity and to investigate the phosphomolybdenum blue method “Talanta 201 (2019) 519–526

 

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