Effects of agitation and aeration in mixing time determination for viscous suspensions using Double Indicator System

 

P. Satya Madhuri, B. Sri Moukthika, N. Sumanth, K. Sri Vinusha,V. S. Rama Krishna Ganduri^*

Department of Biotechnology, K. L. University, Green Fields, Vaddeswaram- 522 502, Andhra Pradesh, India.

 

ABSTRACT:

Mixing time is the time taken to achieve a predefined level of homogeneity of the mixture in a reaction vessel using a tracer. It is one of the key parameters to evaluate the mixing efficiency in agitated systems. In order to make this definition valid, the tracer should be in the same physical phase (e.g. liquid) as the bulk material. In this study, determination of mixing time was done by Double Indicator System for Mixing Time (DISMT), which is a colorimetric method that indicates homogeneity in the reaction mixture composition. Here, we attempted the determination of mixing parameters by varying energy input configurations, i.e., with only mechanical agitation, with only aeration, and combination of agitation and aeration using different liquid broths, viz., water, 5%(w/v) of Sucrose,Starch,Polyethylene glycol, and Glycerol. DISMT experiments were conducted using food colors as indicators and confirmed the use of these food colors as tracers. In addition, the dependence of mixing time on gas (air) hold up was also elucidated experimentally.

 

 

 

INTRODUCTION:

Mixing is considered as fundamental evolution time of spatially dependent concentrations toward achieving final homogeneous state. Mixing processes have an access to concentration maps of relevant species in entire vessel as function of time. This mixing time, as an empirical parameter, define the overall behavior of stirred tanks¹. For its determination, colorimetric probes or planar laser induced fluorescence measurements are being widely used. In colorimetric methods, a tracer pulse is injected into the liquid batch and the concentration transient is detected at a selected point. Several tracer properties and measuring techniques have been used for this purpose^2,3,4.

 

However, for the definition of mixing time as macroscopic parameter deduced objectively from quantitative concentrations measured non-intrusively for the whole volume of the mixing vessel, available experiment techniques may be considered as a partial route of initial purpose.

 

Mixing time can be determined either with experiments or numerical modeling. The experimental methods to determine the mixing time in liquid include conductivity method and discoloration method⁵. The conductivity method requires a conductivity probe to present in the target system, which make it an intrusive method because the existence of the probe might change the mixing efficiency of the mixing device. Discoloration method does not require any probe which makes it a non-intrusive method. However, the color detection device (sometimes the human eye) needs to be calibrated against the conductivity method. Both methods are usually applied to monitor the concentration of the tracer in the most difficult to mix locations such as the area adjacent to the impeller shaft. The benefit of numerical modeling is that once the modeling is completed, the blend time of any predetermined level of homogeneity of any location within the mixing system can be predicted, which is impossible to accomplish by experimental methods. However, numerical modeling needs to be validated by experimental methods^6,7.

 

Double Indicator System for Mixing Time (DISMT) method is being widely applied to highly viscous media that reduces the range of concentration and encodes in terms of colors of evolving mixture. DISMT uses two standard acid-base indicators, acidic methyl red (red to yellow) and basic thymol blue (yellow to blue) which turn, the regions whose mixing fractions are within 5% of the mixing fraction at infinite times, into yellow^2,8,9,10. Using a transparent mixing vessel, an observer may notice the distinct red and blue regions and their emergence as yellow regions. Several researchers have successfully used this method for determining 95% mixing time for entire vessel, i.e., the time taken for entire liquid volume to become yellow. DISMT system is cheap in price and simple in operation which uses low toxic reagents and mixed liquids can be drained back into reservoirs^10,11,12.

 

In the present study, two food colors were used to study the mixing times for different viscous solutions as function of impeller rotational speed and air flow rates, using pulse injection based DISMT system. The dye system concentration, for it to act as an indicator of homogeneous mixing, was standardized. The relationship of different energy input modes (mechanical mixing or agitation, aeration and combination of both) with mixing time of the broth was deduced experimentally.

 

MATERIAL AND METHODS:

Chemicals and reagents:

DISMT analysis was performed using1% (w/v) tracer solutions of ‘Allura’ red (E129) and Fast green (FCF) food colors. De-ionized water was used in all procedures. In this study, water and fixed concentration (5% w/v) of Sucrose, Starch, PolyEthylene Glycol (PEG) and Glycerol were used as simulated fermentation broths.

 

Experimental Setup:

The experiments were carried out in 5 L (3 L working volume, hemispherical bottom) laboratory bioreactor (Lark Innovative Fine Teknowledge), in which mixing system consists of 6-flat blade turbine impeller and three equi-positioned baffles. The bioreactor and its agitation system characteristics are given Table 1.The entire set up facilitated aeration with a vertical sparger and agitation promoted by Rushton turbine impeller integrated with an electrical motor. Schematic representation of bioreactor setup for conducting DISMT analysis is shown in Fig.1.

 

Table 1. Characteristics of bioreactor and impeller

 

Fig. 1: Schematic representation of the reactor set up for carrying out DISMT analysis.

 

Determination of Mixing time and (dimensionless) mixing factor:

Both the food colors were released into the vessel at the same time (pulse input) and the change in color was observed. Mixing time was considered precisely when the mixing of two colors rigorously identified (as complete or homogeneous mixing resulted into pale yellow zone) as ‘mixed within tolerance’. With DISMT, the 95% of mixing time was defined as the time taken for complete liquid volume to turn into yellow.The mixing process was captured by a color video camera located in front of the vessel, images were shot frame by frame simultaneously and further used for analysis.Dimensionless mixing factor, F_t was calculated using the following eqn. (1),

 

                                                (1)

 

where,    t_Mis the mixing time,

                Nis the impeller speed,rps

                H is the height of the liquid level in the vessel (12.6cm),

                D_iis impeller diameter, cm, and

                D_tis tank diameter, cm

               

Determination of Gas Holdup:

Gas hold up was measured by visual method. A graduated graph paper was pasted on the outside of the vessel in between two baffles. A dimensionless fractional gas hold-up (ε_g) was estimated using eqn. (2)¹³:

                                                                                (2)

 

where, H_g is the height of liquid after aeration, cm,

                H_b is the height of clear liquid (broth) without aeration =12.6cm.

 

DISMT analysis when the broth is mechanically agitated:

Each of the experimental viscous solutions was mechanically agitated at varied impeller speeds of 100, 200 and 300 rpm and the corresponding mixing time was determined based on the time frames in the video followed by determination of respective mixing factor.

 

DISMT analysis when the broth isaerated:

Sparging of filtered air was done at different volumetric flow rates of 2, 4, 6 and 8 L/min. for all the viscous solutions. Their mixing times, liquid rise heights and gas holdup were estimated.

 

 

DISMT analysis when the reactor contents are aerated and mechanically agitated:

Mixing times and gas holdup were determined by varying air flowrates (2, 4, 6 and 8 L/min.) at each impeller rotational speed (100, 200, and 300 rpm) for all the solutions.Each experiment has been carried out thrice, for identical conditions, the average value of mixing times was reported.

 

RESULTS AND DISCUSSIONS:

Any bioreactor equipped with one or more impellers and operated under aerobic conditions, gets complicated as the combined action of mechanical and pneumatic mixing has to be considered. The analysis of mixing efficiencies for aerated systems uses the principles of non-aerated systems and it was assumed that the gas phase does not influence the flow of liquid phase in mechanically stirred systems^14,15. In this context, the experiments were carried out to study the effects of variations in the impeller rotational speed, air flow rates on mixing time for water and other simulated broths having different viscosities.

 

Influence of Impeller rotational speed:

Contrarily to the non-aerated systems, for which the mixing time is reduced by increasing the impeller rotational speed value, for aerated broths the influence of this parameter is different and must be related to the apparent viscosity and air flow rate. The mixing times were observed as the change in the color of whole solution by injecting of dual color solutions as tracers (Fig.2). For all the solutions, the mixing times were decreased with increase in rotational speed and (dimensionless) mixing factors were increased with increase in speed. This evolution could be the result of modification of mixing mechanism with increase in rotational speed (Fig. 3). The value of the rotation speed which corresponds to the minimum of mixing time is called critical rotation speed¹⁶.The observations were tabulated in Table 2.

 

Fig.2. Frames extracted from a sequence of images captured during mixing process with only mechanical agitation.

 

Fig.3. Influence of impeller rotational speed on (a) mixing time and (b) on dimensionless mixing factor.

 

Table 2. Influence of Impeller speed (only agitation) on mixing parameters

 

Influence of air volumetric flow rate:

Sparging of air into liquid broth strongly influence the mixing intensity and depends on the apparent viscosity of liquid phase. In general, for water and other liquids, the mixing time continuously decreases with air flow rate, the magnitude of this effect can be a function of impeller rotational speed. Fig. 4 and 5 shows the variation of mixing efficiency with aeration rate for different viscous media. At lower aeration rates, it was observed that size of air bubble was large, which made heterogeneous distribution of air in the liquid phase and this caused less air hold-ups, rise of bubbles through preferential central routes, in turn resulted in higher values of mixing times (Table 3). Further, for the increase of aeration rate induced the decrease of mixing time. This variation was due to the formation of smaller bubbles having lower rise velocity, which led the increase in gas hold-up values.

 

Combined influence of impeller rotational speed and air volumetric flow rate:

To study the influence of combined effects of both rotational speed and aeration rate, another set of experiments were carried out for the same set of liquids, i.e., water and fixed concentration (5%) of sucrose, starch, PEG and glycerol. The results indicated that the increase in the aeration rate with impeller speed had decreased the mixing time value and increased gas hold-up values for all the viscous liquids (Table 4). This was due to integrated effect of both aeration and agitation which made the quick mixing distribution possible inside the liquid phase (Fig.6 and 7).Gas hold up within the reaction mixture influences the mixing time, though not to a great extent.

 

Fig.4. Frames extracted from a sequence of images captured during mixing process with only aeration.

 

Fig.5. Influence of (a) air flow rate and (b) gas hold-up on mixing time.

 

Table 3. Influence of Air flow rate (only aeration) on mixing parameters.

 

Fig.6. Frames extracted from a sequence of images captured during mixing process with integrated effect of aeration and agitation.

 

Fig.7. Integrated influence of aeration and agitation for impeller speed (a) at 100 rpm, (b) at 200 rpm, and (c) at 300 rpm on mixing time.

 

CONCLUSION:

The flow mechanisms of aerated and non-aerated systems are very different as they have been influenced by impeller rotational speed and volumetric flow rate of air on mixing time. In case of mechanically agitated and equipped with DISMT system, the increase of impeller speed had reduced the mixing times and increased dimensionless mixing factor, for water and other simulated viscous liquids. For only aerated system, the increase of airflow rate, decreased the mixing time with increased gas hold-up values. Finally, in case of both aerated and agitated systems, increase in impeller speed and airflow rate decreased the mixing time values, while increase in the gas hold-up was also observed. Thus, the observed phenomena in this study could be a consequence of operational variation in pumping capacity of stirrer assembly, due to cavity formation, compartmentalization in regions around the impeller, coalescence and dispersion of bubbles and flooding.

 

Table 4. Integrated influence of impeller speed and airflow rate on mixing parameters.

Liquid broth	Impeller rotational speed, rpm	Air Flow Rate, Lpm	Mixing Time, sec	Gas Hold-up
Water	100	2	3.13	0.0667
		4	3.05	0.0735
		6	2.98	0.0803
	200	2	2.72	0.0735
		4	2.51	0.0803
		6	2.06	0.0935
	300	2	1.96	0.0803
		4	1.48	0.0870
		6	1.03	0.1000
5% Sucrose	100	2	3.99	0.0526
		4	3.71	0.0597
		6	3.46	0.0667
	200	2	3.35	0.0526
		4	3.01	0.0526
		6	2.62	0.0667
	300	2	1.98	0.0597
		4	1.72	0.0667
		6	1.03	0.0735
5% Starch	100	2	4.09	0.0382
		4	3.88	0.0455
		6	3.79	0.0455
	200	2	3.77	0.0455
		4	3.56	0.0526
		6	3.02	0.0526
	300	2	2.45	0.0597
		4	2.20	0.0667
		6	1.71	0.0735
5% Poly Ethylene Glycol	100	2	4.79	0.0308
		4	4.58	0.0455
		6	4.06	0.0526
	200	2	3.92	0.0382
		4	3.71	0.0455
		6	2.94	0.0597
	300	2	2.69	0.0526
		4	2.40	0.0597
		6	1.98	0.0667
5% Glycerol	100	2	4.97	0.0156
		4	4.72	0.0233
		6	4.63	0.0382
	200	2	4.58	0.0233
		4	4.01	0.0308
		6	3.83	0.0455
	300	2	3.07	0.0382
		4	2.69	0.0455
		6	2.05	0.0597

 

ACKNOWLEDGEMENTS:

The authors are very much thankful to the management of K. L. University, Vaddeswaram, Guntur, for providing the facilities to carry out this work and also thank Ms. K. Swathi, Lab technician for extending her support towards accomplishment of this experimental study.

 

 

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

 

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Accepted on 09.09.2016           © RJPT All right reserved

Research J. Pharm. and Tech 2016; 9(11): 1971-1977.