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Dispersed phase hold-up in a Graesser raining bucket contactor using aqueous two-phase systems
Journal of Food Engineering 72 (2006) 302–309 www.elsevier.com/locate/jfoodeng
Research note
Dispersed phase hold-up in a Graesser raining bucket contactor using aqueous two-phase systems Abraham D. Giraldo-Zuniga a,*, Jane S.R. Coimbra b, Luis A. Minim b, Edwin E. Garcia Rojas b b
a Curso de Engenharia de Alimentos, Universidade Federal do Tocantins (UFT), 77020-210, Palmas, TO, Brasil Laborato´rio de Processos de Separac¸a˜o (LPS), Departamento de Tecnologia de Alimentos, Universidade Federal de Vic¸osa (UFV), 36571-000, Vic¸osa, MG, Brasil
Received 20 January 2004; accepted 23 November 2004 Available online 17 March 2005
Abstract This study presents dispersed hold-up data for an aqueous two-phase system containing polyethylene glycol (PEG) 1500 Da and potassium phosphate (PPP) at pH 7 and 25 C in a Graesser raining bucket contactor. The influences of the operational variables rotor speed and phase flow ratio on the hold-up were analyzed. Densities, viscosities and interfacial tension were measured. The results obtained were correlated by the equation proposed by Richardson and Zaki, for the description of the slip velocities. 2005 Elsevier Ltd. All rights reserved. Keywords: Hold-up; Graesser contactor; Aqueous two-phase systems; Bioseparation
1. Introduction Data of dispersed phase hold-up and slip velocity are necessary for the design of liquid–liquid extraction columns. The hold-up is needed to calculate the interfacial area per unit volume and the phase velocities. It is an important factor as it not only determines mass transfer but also dictates the onset of flooding (Kumar & Hartland, 1995; Aravamudan & Baird, 1999; Misek, 1994). The performance of an extraction unit operated continuously is dependent on the amount of solvent present in the extractor. If the solvent quantities are high compared to the feed, the solute concentration gradients are favorable to mass transfer. The hold-up represents the amount of solvent available in the extractor to extract the feed solute (Coimbra, Mojola, & Meirelles, 1998). *
Corresponding author. Tel.: +55 63 218 8086/2923; fax: +55 63 218 8020/2922. E-mail address: [email protected] (A.D. Giraldo-Zuniga). 0260-8774/$ - see front matter 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.jfoodeng.2004.11.037
Aqueous two-phase systems (ATPS) have been used as a biomolecule purification method over the last three decades (Albertsson, 1986; Zaslavsky, 1995). The most frequently used systems are the polyethylene glycol (PEG)–dextran and the PEG–salt system. ATPS will be formed above certain concentrations of the two components in water. In a polymer–polymer system each phase is enriched in one of the polymers, while in a polymer–salt system one polymer-rich phase is in equilibrium with a salt-rich phase. Both systems contain (60–90)% of water in each phase, what makes them suitable for separation of biological materials (Husted, Kroner, Menge, & Kula, 1985; Wu, Pereira, Venaˆncio, & Teixeira, 2001). Aqueous two-phase extraction may be used most efficiently it the beginning of the purification process (Rojas et al., 2004). It offers distinct advantages over classical methods of protein purification, such as protein fractional precipitation and centrifugation (Papamichael, Boerner, & Husted, 1992). Some ATPS advantages include, low material costs, good reproducibility, minimal
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303
Nomenclature ATPS Hd Vt V (H) Vslip
aqueous two-phase systems fractional hold-up of the dispersed phase volume of the dispersed phase total volume of the extraction region hold-up slip velocity
protein denaturation and easy scale-up (Kula, Kroner, & Husted, 1982). Equipment used in the organic liquid–liquid extraction can be applied for extraction process using ATPS. Husted, Kroner, Menge, and Kula (1988) studied the continuous process carried out in a Kuhni column, a Graesser contactor, a mixer-settler unit and a Podbielniak extractor. Rostami-Jafarabad, Patil, Sawant, and Joshi (1992) investigated the mass transfer in a YorkSheibel column using ATPS. Recently, Jarudilokkul, Paulsen, and Stuckey (2000) used a Graesser contactor to extract lysozyme from egg. The Graesser raining bucket differential contactor was originally developed to handle difficult settling systems encountered in the coal tar industry (Coleby, 1962). However, its simple mechanical design, relatively low cost and other advantages have made it attractive for much broader industrial applications. The Graesser contactor is a useful tool for the treatment of two-phase systems with low density difference, low interfacial tension and tendency for emulsification, which are physical ATPS characteristics. This special design leads to some specific hydrodynamic characteristics of the Graesser contactor, which makes it especially suitable for ATPS processing (Coimbra, Thommes, & Kula, 1994). The equipment seems to fit the demands of continuously operated separation of proteins using ATPS. This was predicted by Coimbra et al. (1994) and Jarudilokkul et al. (2000) who used a Graesser contactor for continuous separation of whey proteins and lysozyme, respectively. Dispersed phase hold-up characteristics of Graesser contactor and of several liquid–liquid extraction columns have been studied experimentally using mixtures of water and organic solvents. Spray columns (Raghav-Rao, Szlag, Sikdar, Joshi, & Sawant, 1991), Yor-Scheibel column (Rostami-Jafarabad et al., 1992), Graesser extractor (Coimbra et al., 1994; Husted et al., 1988) and another column types have been used successfully a great variety of extraction process. Besides, a limited number of studies on dispersed phase hold-up behavior have been carried out using in a Graesser contactor operated with ATPS. The present study reports dispersed phase hold-up, in a Graesser contactor using
c V0 Qp Qs
parameter calculated by the least-squares method characteristic velocity polymeric phase flow rate saline phase flow rate
ATPS composed by PEG 1500–potassium phosphate at 25 C and pH 7.0.
2. Experimental 2.1. Equipment The Graesser contactor is composed of a horizontal cylindrical shell containing an internal rotor assembly consisting of a series of circular discs mounted on a central shaft. Between each pair of discs, a series of buckets are supported by tie rods extending the length of the apparatus. Under operation, the interface level of the two liquids is controlled at the central line of the vessel. Gentle mixing is provided by the movement of the buckets, transporting portions of each phase to the other phase. Phase separation occurs in the end zones on the right and left sides of the cylindrical vessel. For more details of construction and operational principles also see Coleby (1962). The Graesser contactor (QVF Glastechnik, Germany) consists of a glass cylinder with an internal rotor containing 35 compartments formed by 37 circular stainless steel boards. Each of them contains six stainless steel buckets. A 0.25-kW motor was used to drive the rotor. The rotor movement direction had been reversed to obtain more stable operational conditions (Coimbra et al., 1994). The extractor dimensions are listed in Table 1. The rotation speed varied from 6.6 rpm to 15.5 rpm. 2.2. Preparation of the aqueous two-phase system The optimization of the extraction phase system has been described in a previous paper (Giraldo-Zuniga, Coimbra, & Minim, 2002). The systems used here were composed by 18% (w/w) PEG 1500, 18% (w/w, pH 7.0) potassium phosphate (PPP) and 64% (w/w) water. The systems were prepared by adding directly 3.6 kg of PEG 1500 (Labsynth, Brazil), 1.276 kg of potassium phosphate monobasic (Vetec, Brazil), 2.324 kg of phosphate potassium dibasic (Vetec, Brazil) and 12.8 kg of distilled water, totalling 20 kg. The components were agitated for 2 h. After interrupting the agitation, the
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Table 1 Graesser extractor dimensions
3. Results and discussion
Diameter [cm] Internal Discs Bucket tubes Input and output tubes Rotor
10.0 9.40 2.54 1.30 1.30
Length [cm] Total Mixing zone Settling zone Between discs
101 92 9.0 2.54
3.1. Phase diagram for the aqueous two-phase system studied
system was left to decant for 12 h for phase separation and to achieve the equilibrium. The bottom and the top phases were separated and introduced into the column by peristaltic pumps. In all experiments, the saltrich phase was the bottom phase and the PEG-rich phase was the top. Phase densities were measured by pycnometry. Phase viscosities were determined by using an Ostwald rheometer and the interfacial tension using a spinning drop tensiometer (SITE 04, Kru¨ss, Germany). 2.3. Operational conditions Hold-up was determined at rotation speeds of 6.6, 10, 12.5 and 15.5 rpm, at phase flow rate ratios (bottom:top) of 1.0, 1.3, 2.0 and 4.0, corresponding to the ratios (bottom:top) of 80:80, 80:60, 80:40 and 80:20 ml min1 for bottom phase fixed. When the polymeric phase was maintained fixed the flow rate (bottom:top) were 1.0, 0.75, 0.5 and 0.025, corresponding to the (bottom:top) ratio of 80:80, 60:80, 40:80 and 20:80 ml min1. The upper and lower phases were pumped (peristaltic pumps, Cole Parmer) into the extractor above and below the rotor axis at opposite ends of the contactor. A digital tachometer was used, to measure the rotation speed. 2.4. Hold-up experiments The fractional hold-up of the PEG-rich phase was measured after steady state was reached, or either after approximately 120 min for each operation condition. This was performed by the simultaneous interruption of the rotation and of the inlet and outlet streams. The extractor was emptied and the total volume and the volume of both phases were measured. This method leads to an average hold-up calculated by H d ¼ V t =V
ð1Þ
where Hd is the fractional hold-up of the dispersed phase; Vt is the volume of the dispersed phase and V is the total volume of the extraction region.
In Fig. 1 is shown the phase diagram of potassium phosphate–PEG 1500 system at 25 C, indicating phase component concentrations at which separation occurs. Two-phase systems form the component concentrations above the binodal curve. This curve shows the required concentrations for the phase separation. After phase separation, the top phase becomes PEG-rich and the bottom phase becomes PPP-rich. As shown in Fig. 1, the PPP–PEG 1500 systems required lower PEG concentrations and high PPP concentrations to form the two-phase systems. This effect was also observed by Albertsson (1986) using PEG of different molecular masses. 3.2. Influences of the rotor speed and flow rate ratio on the hold-up The hold-up (H) values are the volumetric concentrations of the disperse phase in dispersion, corresponding to each of the measured data points for all operational as shown in Table 2. Fig. 2 shows the polymeric phase hold-up as a function of the rotation speed for different flow rate ratios with the flow rate of the saline phase maintained constant (80 ml min1). A very smooth variation of polymeric phase hold-up on the increase of the rotation speed was observed for four different flow rate ratio. A discreet tendency of hold-up reduction was observed at higher rotation speed. The rotation speeds affected the size and upward velocity of the drops, with high rotation speeds resulting in an intense drop breakage. Small
40
PEG (% w/w)
304
30
20 M
B
10
0 0
5
10
15
20
25
30
Potassium Phosphate (% w/w)
Fig. 1. Phase diagram for potassium phosphate–PEG 1500 system at 25 C and pH 7: (. . . . . .): binodal curve, (d): initial composition, (n): top phase, (h) bottom phase, (M): monophasic mixture, (B) biphasic region.
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Table 2 Hold-up values for the aqueous two-phases systems Flow rates
Rotation speed 1
1
Qp (ml min )
Qs (ml min )
6.6
10
12.5
15.5
80 60 40 20 80 80 80 80 60 70 40 80
80 80 80 80 80 60 40 20 60 50 80 40
0.5604 0.4553 0.4110 0.2526 0.5604 0.5635 0.5756 0.6225 0.4870 0.4788 0.4115 0.5757
0.5566 0.5440 0.4528 0.2463 0.5566 0.5644 0.6200 0.7023 0.4617 0.4300 0.4518 0.6205
0.5756 0.4781 0.4110 0.2716 0.5756 0.5844 0.6048 0.7783 0.5288 0.5180 0.4123 0.6039
0.4971 0.5034 0.3793 0.2400 0.4971 0.5067 0.6060 0.7276 0.5212 0.5250 0.3789 0.6061
0.8
1.0
0.8 Polymeric phase hold-up
Polymeric phase hold-up
0.6
0.4
0.2
Qs/Qp = 1 Qs/Qp = 1.3 Qs/Qp = 2 Qs/Qp = 4
0.6
0.4 Qs/Qp = 1.0 Qs/Qp = 0.75 Qs/Qp = 0.50 Qs/Qp = 0.25
0.2
0.0
0.0 6
8
10 12 rotation speed (rpm)
14
16
6
8
10
12
14
16
rotation speed (rpm)
Fig. 2. Dependence of polymeric phase hold-up on the rotation speed (Qs: flow rate of the saline phase [80 ml min1—fixed]); Qp: flow rate of the polymeric phase).
Fig. 3. Dependence of polymeric phase hold-up on the rotation speed (Qs: flow rate of the saline phase; Qp: flow rate of the polymeric phase [80 ml min1—fixed]).
drops tend to present higher upward velocity reducing the residence time in the extractor and consequently the hold-up, although insignificant variation in polymeric hold-up was observed. The hold-up decreased with increase of the flow rate relationship and with the decreased flow rate of the polymeric phase. This behavior is probably generated by the presence of smaller rates of polymeric phase in the equipment. Eq. (1) shows that the saline phase hold-up under the same operational conditions would be larger than that in the polymeric phase, also presenting inverse behavior when this phase is maintained fixed. Fig. 3 shows the effect of the rotation speed on the polymeric phase hold-up for different flow rate values of the saline phase at the polymeric phase flow rate of 80 mL min1. In this case, the polymeric phase holdup decreased on the increase of the saline phase volume inside the extractor. Fig. 3 shows that the hold-up was practically constant at the different rotation speeds tested here, except when the flow rates ratio was
very low. An example is the relationship of 0.25 (20/80) ml min1, for which the hold-up values between 0.62 and 0.73, showing a variation of 18%. Fig. 4 shows that the variation of the polymeric phase hold-up was maintained constant when the sum of the flow rates of the inlet streams in the equipment was kept constant. Changes in the rotation speeds did not affect the hold-up substantially. For different flow rate phases ratios, the hold-up remained in a narrow band. The rotation speed had a little influence on the holdup for the systems studied in this work. This behavior can be attributed to the fact that the Graesser contactor contains two independent flowing phases with an interface line that can be controlled in the central line of the contactor, establishing a small band of variation of the hold-up values. This behavior was also observed by Coimbra et al. (1994). Sheikh, Ingham, and Hanson (1972) described a behavior for the hold-up similar to that observed in this study. The authors analyzed the influence of a Graesser
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the continuous separation of a-lactalbumin and b-lactoglobulin from whey with ATPS, obtaining satisfactory yields at a rotor speed of 2 rpm. Recently, Jarudilokkul et al. (2000) employed a Graesser contactor to separate lysozyme from egg white using reverse micelles, also obtaining the optimal conditions for the extraction of lysozyme at a rotor speed of 5 rpm, and a phase flow relation of 60:20 ml min1, indicating low rotor speeds as good operational condition for this equipment.
Polymeric phase hold-up
0.8
0.6
0.4
0.2
60/60 70/50 80/40 40/80
3.3. Correlation
0.0 6
8
10 12 rotation speed (rpm)
14
16
Fig. 4. Dependence of polymeric phase hold-up on the rotation speed. Total flow rate constant (saline phase + polymeric phase) = 120 ml min1.
extractor (75 cm of length for 15 cm of diameter) rotation speed, using a system composed by water–n-butylamine– kerosene. An increased disperse phase hold-up with increased flow rate relations was verified, which is an opposite tendency to our results. This behavior can be expected since the hold-up is very dependent on this type of extractive system, physical properties, type of equipment and operational conditions. The physical properties of the ATPS studied are very different from the organic liquid–liquid extraction systems, mainly regarding viscosity and interfacial tension values. In the vertical columns, the rise of hold-up with the increase of the rotation speed and flow rate relation has been previously observed. For example, Coimbra et al. (1998), studying the hydrodynamic of a perforated rotating disc contactor (PRDC) using ATPS formed by PEG 6000 and dibasic potassium phosphate, observed an increase in hold-up with increased rotating disc speed and phases flow relation. Hamidi, Van Berlo, Luyben, and Wiwlen (1999) found polymeric phase hold-up values ranging from 0.1 to 0.9 and also observed an increase in hold-up with increased phases flow relation for a countercurrent sieve-plate column with ATPS composed by PEG 400 and Na2SO4. For a Karr extractor, Kader (1985) and Aravamudan and Baird (1999) verified the increase in hold-up with increased agitation speed for the systems acetone–toluene– water, kerosene oil–water and isopropyl alcohol–water. Among the different flow rate of phase tested, the interface in the extraction zone was not clearly observed at 15 rpm. The interface that should be controlled in the central line of the contactor could only be maintained in the separation zones. Wang, Ingham, and Hanson (1977) studied axial mixing and mass transfer in a Graesser contactor (15 cm diameter, 75 cm long) changing the rotor speed from 4 to 13.3 rpm, low rotor speed and low (solvent/aqueous) flow ratio being carried out to best overall mass transfer performance. Coimbra et al. (1994), also using a Graesser contactor, studied
Aiming to compare our hold-up data with those predicted by correlations developed for conventional liquid–liquid systems, we used the Richardson and Zaki (1954) correlation. This equation is applied to various types of extractors, except the Graesser contactor. The hold-up values obtained experimentally were used to determine the slip velocity (Vslip) according to Eq. (2). The slip velocity is the single most important variable controlling the mass transfer coefficients when a solute is transferred between the two phases (Misek, 1994). The direct measurement of slip velocity in the extraction columns is difficult, so it may be obtained from the flow rates of the continuous and dispersed phases and dispersed-phase hold-up. According to Slater (1985), a better slip velocity equation, first formulated by Richardson and Zaki (1954) is V slip ¼ V 0 ð1 H Þ
c
ð2Þ
V0 and exponent c are parameters calculated by the least-squares method. Within limits of 0 < H < 0.3, it was demonstrated that the slip velocity in various types of extraction equipment can be described by this empirical equation. In this work, Eq. (2) was correlated to the experimental data. Figs. 5–9 shows the experimental and correlation results for all the tested conditions.
0.001
6.6 rpm 10 rpm 12.5 rpm 15.5 rpm predicted 6.6 rpm predicted 10 rpm predicted 12.5 rpm predicted 15.5 rpm
0.0008
Vslip (m s-1)
306
0.0006
0.0004
0.0002 0.3
0.4
0.5
0.6
0.7
0.8
1—H
Fig. 5. Correlation of Eq. (2) to the experimental data for the polymeric phase (fixed).
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0.001 0.0008
0.0006
Vslip exp (m s-1)
0.0012
Vslip (m s-1)
0.0007
6.6 rpm 10 rpm 12.5 rpm 15.5 rpm predicted 6.6 rpm predicted 10 rpm predicted 12.5 rpm predicted 15.5 rpm
0.0014
307
0.0006
0.0005
0.0004 0.0002 0.0004 0 0.1
0.2
0.3
0.4
0.5
1—H 0.0004
0.0005
Fig. 6. Correlation of Eq. (2) to the experimental data for the saline phase (fixed).
0.0006
0.0007
Vslip cal (m s-1)
Fig. 8. Slip velocity: observed and correlated (Eq. (2)) values for polymeric phase—fixed (R2 = 0.77).
0.0007
Vslip exp (m s-1)
Vslip exp (m s-1)
0.000495
0.0006
0.000490
0.0005
0.000485
0.0004 0.0004
0.0005
0.0006
0.00050
0.0007
0.00052
0.00054
Vslip cal (m s-1)
Vslip cal (m s-1)
Fig. 7. Slip velocity: observed and correlated (Eq. (2)) values for saline phase—fixed (R2 = 0.95).
Fig. 9. Slip velocity: observed and correlated (Eq. (2)) values for the sum of the flow rate constant (R2 = 0.75).
According to Figs. 4 and 7, the validity of Eq. (2) to represent our experimental data has been confirmed for small hold-up values, such as the saline phase-fixed hold-up values, whose average value of hold-up was of 0.42. The mean value was obtained from an arithmetic average of measurement at each condition. The good agreement between calculated and experimental values is shown in Fig. 7, with the residual analysis for Vslip resulting in R2 higher than 0.95. For hold-up value means of 0.60 and 0.50 corresponding to polymeric phase-fixed and the sum of the
flow rate constant, respectively, the validity of the eq. (2) to represent our experimental data has not been confirmed, as can be seen in Figs. 6, 7 and 9. In Figs. 8 and 9 can be observed the residual analysis for Vslip with R2 less than 0.77 and 0.75, respectively. The difference between experimental data and correlated data is probably due to the fact that the hold-up values for our systems are not in the region of validity of Eq. (4). Also the systems used in this work are quite different from the water–organic solvent systems used to develop the correlations given in this work, such as very small
Table 3 Values of exponent c and characteristic velocity V0 Condition
Flow rate 6.6 rpm
Qs—fixed Qp—fixed Qs + Qp = 120
10 rpm 1
12.5 rpm 1
15.5 rpm 1
c
V0 (m s )
c
V0 (m s )
c
V0 (m s )
c
V0 (m s1)
0.9910 3.6224 0.0347
0.0003 0.0123 0.0005
0.9133 1.4530 0.0347
0.0003 0.0022 0.0005
0.9654 3.3617 0.0347
0.0003 0.0122 0.0005
1.0701 0.8123 0.0347
0.0003 0.0011 0.0005
308
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interfacial tension, small density and a high viscosity difference between the dispersed and continuous phases. This is uncommon in the majority of the organic liquid–liquid extraction systems. Differences in their hydrodynamic behaviors are thus likely to be expected. The exponents c and V0 resulting from the correlation of all conditions are given in Table 3. Kumar and Hartland (1994) found values of Vslip ranging from 0.0022 to 0.256 m s1 for spray columns, 0.0037 to 0.3178 m s1 for rotating discs and asymmetric rotating disc columns, 0.0025 to 0.110 m s1 for Kunhi columns, 0.0032 to 0.1464 m s1 for pulsed perforated-plate columns and 0.0084 to 0.1156 m s1 for Karr reciprocating columns. Godfrey and Slater (1991) found values for c ranging from 0.3 to 1.5 for packed columns, 0 to 4 for rotary agitated columns and 3 to 1 for sieve plate columns. These values were obtained for conventional liquid–liquid systems (water–organic solvent). Hamidi et al. (1999) found values for c ranging from 0.27 to 0.16 for countercurrent sieve plate column for aqueous two-phase systems composed by PEG 4000 and Na2SO4 and 0.31 for the V0 value. The values obtained in this work for Vslip ranging from 3.88 · 104 ml min1 to 6.93 88 · 104 ml min1 and for c values the range obtained was from 0.9133 to 3.6224. The differences in the Vslip and c values can be expected due to the different difference of type of equipments and physical properties of the extraction systems used in this work.
4. Conclusions For a constant flow rate of the saline phase, a tendency of the hold-up decrease on the increase of phase flow ratio was observed. When the polymeric phase was maintained fixed, hold-up was also decreased with the increase of the phase flow ratio. The rotation speed of 6.6 rpm and the flow rate ratio of 80/40 ml showed optimal conditions for the extractor operation regarding the best interface control. The correlation of slip velocity and hold-up using Eq. (2) is possible for the Graesser contactor using aqueous two-phase systems for small hold-up values, with the residual analysis for Vslip resulting in R2 higher than 0.95.
Acknowledgement The authors thank FAPEMIG, PADCT/CNPq (62.0167/97-1) and CAPES, for the financial support.
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Research note
Dispersed phase hold-up in a Graesser raining bucket contactor using aqueous two-phase systems Abraham D. Giraldo-Zuniga a,*, Jane S.R. Coimbra b, Luis A. Minim b, Edwin E. Garcia Rojas b b
a Curso de Engenharia de Alimentos, Universidade Federal do Tocantins (UFT), 77020-210, Palmas, TO, Brasil Laborato´rio de Processos de Separac¸a˜o (LPS), Departamento de Tecnologia de Alimentos, Universidade Federal de Vic¸osa (UFV), 36571-000, Vic¸osa, MG, Brasil
Received 20 January 2004; accepted 23 November 2004 Available online 17 March 2005
Abstract This study presents dispersed hold-up data for an aqueous two-phase system containing polyethylene glycol (PEG) 1500 Da and potassium phosphate (PPP) at pH 7 and 25 C in a Graesser raining bucket contactor. The influences of the operational variables rotor speed and phase flow ratio on the hold-up were analyzed. Densities, viscosities and interfacial tension were measured. The results obtained were correlated by the equation proposed by Richardson and Zaki, for the description of the slip velocities. 2005 Elsevier Ltd. All rights reserved. Keywords: Hold-up; Graesser contactor; Aqueous two-phase systems; Bioseparation
1. Introduction Data of dispersed phase hold-up and slip velocity are necessary for the design of liquid–liquid extraction columns. The hold-up is needed to calculate the interfacial area per unit volume and the phase velocities. It is an important factor as it not only determines mass transfer but also dictates the onset of flooding (Kumar & Hartland, 1995; Aravamudan & Baird, 1999; Misek, 1994). The performance of an extraction unit operated continuously is dependent on the amount of solvent present in the extractor. If the solvent quantities are high compared to the feed, the solute concentration gradients are favorable to mass transfer. The hold-up represents the amount of solvent available in the extractor to extract the feed solute (Coimbra, Mojola, & Meirelles, 1998). *
Corresponding author. Tel.: +55 63 218 8086/2923; fax: +55 63 218 8020/2922. E-mail address: [email protected] (A.D. Giraldo-Zuniga). 0260-8774/$ - see front matter 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.jfoodeng.2004.11.037
Aqueous two-phase systems (ATPS) have been used as a biomolecule purification method over the last three decades (Albertsson, 1986; Zaslavsky, 1995). The most frequently used systems are the polyethylene glycol (PEG)–dextran and the PEG–salt system. ATPS will be formed above certain concentrations of the two components in water. In a polymer–polymer system each phase is enriched in one of the polymers, while in a polymer–salt system one polymer-rich phase is in equilibrium with a salt-rich phase. Both systems contain (60–90)% of water in each phase, what makes them suitable for separation of biological materials (Husted, Kroner, Menge, & Kula, 1985; Wu, Pereira, Venaˆncio, & Teixeira, 2001). Aqueous two-phase extraction may be used most efficiently it the beginning of the purification process (Rojas et al., 2004). It offers distinct advantages over classical methods of protein purification, such as protein fractional precipitation and centrifugation (Papamichael, Boerner, & Husted, 1992). Some ATPS advantages include, low material costs, good reproducibility, minimal
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303
Nomenclature ATPS Hd Vt V (H) Vslip
aqueous two-phase systems fractional hold-up of the dispersed phase volume of the dispersed phase total volume of the extraction region hold-up slip velocity
protein denaturation and easy scale-up (Kula, Kroner, & Husted, 1982). Equipment used in the organic liquid–liquid extraction can be applied for extraction process using ATPS. Husted, Kroner, Menge, and Kula (1988) studied the continuous process carried out in a Kuhni column, a Graesser contactor, a mixer-settler unit and a Podbielniak extractor. Rostami-Jafarabad, Patil, Sawant, and Joshi (1992) investigated the mass transfer in a YorkSheibel column using ATPS. Recently, Jarudilokkul, Paulsen, and Stuckey (2000) used a Graesser contactor to extract lysozyme from egg. The Graesser raining bucket differential contactor was originally developed to handle difficult settling systems encountered in the coal tar industry (Coleby, 1962). However, its simple mechanical design, relatively low cost and other advantages have made it attractive for much broader industrial applications. The Graesser contactor is a useful tool for the treatment of two-phase systems with low density difference, low interfacial tension and tendency for emulsification, which are physical ATPS characteristics. This special design leads to some specific hydrodynamic characteristics of the Graesser contactor, which makes it especially suitable for ATPS processing (Coimbra, Thommes, & Kula, 1994). The equipment seems to fit the demands of continuously operated separation of proteins using ATPS. This was predicted by Coimbra et al. (1994) and Jarudilokkul et al. (2000) who used a Graesser contactor for continuous separation of whey proteins and lysozyme, respectively. Dispersed phase hold-up characteristics of Graesser contactor and of several liquid–liquid extraction columns have been studied experimentally using mixtures of water and organic solvents. Spray columns (Raghav-Rao, Szlag, Sikdar, Joshi, & Sawant, 1991), Yor-Scheibel column (Rostami-Jafarabad et al., 1992), Graesser extractor (Coimbra et al., 1994; Husted et al., 1988) and another column types have been used successfully a great variety of extraction process. Besides, a limited number of studies on dispersed phase hold-up behavior have been carried out using in a Graesser contactor operated with ATPS. The present study reports dispersed phase hold-up, in a Graesser contactor using
c V0 Qp Qs
parameter calculated by the least-squares method characteristic velocity polymeric phase flow rate saline phase flow rate
ATPS composed by PEG 1500–potassium phosphate at 25 C and pH 7.0.
2. Experimental 2.1. Equipment The Graesser contactor is composed of a horizontal cylindrical shell containing an internal rotor assembly consisting of a series of circular discs mounted on a central shaft. Between each pair of discs, a series of buckets are supported by tie rods extending the length of the apparatus. Under operation, the interface level of the two liquids is controlled at the central line of the vessel. Gentle mixing is provided by the movement of the buckets, transporting portions of each phase to the other phase. Phase separation occurs in the end zones on the right and left sides of the cylindrical vessel. For more details of construction and operational principles also see Coleby (1962). The Graesser contactor (QVF Glastechnik, Germany) consists of a glass cylinder with an internal rotor containing 35 compartments formed by 37 circular stainless steel boards. Each of them contains six stainless steel buckets. A 0.25-kW motor was used to drive the rotor. The rotor movement direction had been reversed to obtain more stable operational conditions (Coimbra et al., 1994). The extractor dimensions are listed in Table 1. The rotation speed varied from 6.6 rpm to 15.5 rpm. 2.2. Preparation of the aqueous two-phase system The optimization of the extraction phase system has been described in a previous paper (Giraldo-Zuniga, Coimbra, & Minim, 2002). The systems used here were composed by 18% (w/w) PEG 1500, 18% (w/w, pH 7.0) potassium phosphate (PPP) and 64% (w/w) water. The systems were prepared by adding directly 3.6 kg of PEG 1500 (Labsynth, Brazil), 1.276 kg of potassium phosphate monobasic (Vetec, Brazil), 2.324 kg of phosphate potassium dibasic (Vetec, Brazil) and 12.8 kg of distilled water, totalling 20 kg. The components were agitated for 2 h. After interrupting the agitation, the
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Table 1 Graesser extractor dimensions
3. Results and discussion
Diameter [cm] Internal Discs Bucket tubes Input and output tubes Rotor
10.0 9.40 2.54 1.30 1.30
Length [cm] Total Mixing zone Settling zone Between discs
101 92 9.0 2.54
3.1. Phase diagram for the aqueous two-phase system studied
system was left to decant for 12 h for phase separation and to achieve the equilibrium. The bottom and the top phases were separated and introduced into the column by peristaltic pumps. In all experiments, the saltrich phase was the bottom phase and the PEG-rich phase was the top. Phase densities were measured by pycnometry. Phase viscosities were determined by using an Ostwald rheometer and the interfacial tension using a spinning drop tensiometer (SITE 04, Kru¨ss, Germany). 2.3. Operational conditions Hold-up was determined at rotation speeds of 6.6, 10, 12.5 and 15.5 rpm, at phase flow rate ratios (bottom:top) of 1.0, 1.3, 2.0 and 4.0, corresponding to the ratios (bottom:top) of 80:80, 80:60, 80:40 and 80:20 ml min1 for bottom phase fixed. When the polymeric phase was maintained fixed the flow rate (bottom:top) were 1.0, 0.75, 0.5 and 0.025, corresponding to the (bottom:top) ratio of 80:80, 60:80, 40:80 and 20:80 ml min1. The upper and lower phases were pumped (peristaltic pumps, Cole Parmer) into the extractor above and below the rotor axis at opposite ends of the contactor. A digital tachometer was used, to measure the rotation speed. 2.4. Hold-up experiments The fractional hold-up of the PEG-rich phase was measured after steady state was reached, or either after approximately 120 min for each operation condition. This was performed by the simultaneous interruption of the rotation and of the inlet and outlet streams. The extractor was emptied and the total volume and the volume of both phases were measured. This method leads to an average hold-up calculated by H d ¼ V t =V
ð1Þ
where Hd is the fractional hold-up of the dispersed phase; Vt is the volume of the dispersed phase and V is the total volume of the extraction region.
In Fig. 1 is shown the phase diagram of potassium phosphate–PEG 1500 system at 25 C, indicating phase component concentrations at which separation occurs. Two-phase systems form the component concentrations above the binodal curve. This curve shows the required concentrations for the phase separation. After phase separation, the top phase becomes PEG-rich and the bottom phase becomes PPP-rich. As shown in Fig. 1, the PPP–PEG 1500 systems required lower PEG concentrations and high PPP concentrations to form the two-phase systems. This effect was also observed by Albertsson (1986) using PEG of different molecular masses. 3.2. Influences of the rotor speed and flow rate ratio on the hold-up The hold-up (H) values are the volumetric concentrations of the disperse phase in dispersion, corresponding to each of the measured data points for all operational as shown in Table 2. Fig. 2 shows the polymeric phase hold-up as a function of the rotation speed for different flow rate ratios with the flow rate of the saline phase maintained constant (80 ml min1). A very smooth variation of polymeric phase hold-up on the increase of the rotation speed was observed for four different flow rate ratio. A discreet tendency of hold-up reduction was observed at higher rotation speed. The rotation speeds affected the size and upward velocity of the drops, with high rotation speeds resulting in an intense drop breakage. Small
40
PEG (% w/w)
304
30
20 M
B
10
0 0
5
10
15
20
25
30
Potassium Phosphate (% w/w)
Fig. 1. Phase diagram for potassium phosphate–PEG 1500 system at 25 C and pH 7: (. . . . . .): binodal curve, (d): initial composition, (n): top phase, (h) bottom phase, (M): monophasic mixture, (B) biphasic region.
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Table 2 Hold-up values for the aqueous two-phases systems Flow rates
Rotation speed 1
1
Qp (ml min )
Qs (ml min )
6.6
10
12.5
15.5
80 60 40 20 80 80 80 80 60 70 40 80
80 80 80 80 80 60 40 20 60 50 80 40
0.5604 0.4553 0.4110 0.2526 0.5604 0.5635 0.5756 0.6225 0.4870 0.4788 0.4115 0.5757
0.5566 0.5440 0.4528 0.2463 0.5566 0.5644 0.6200 0.7023 0.4617 0.4300 0.4518 0.6205
0.5756 0.4781 0.4110 0.2716 0.5756 0.5844 0.6048 0.7783 0.5288 0.5180 0.4123 0.6039
0.4971 0.5034 0.3793 0.2400 0.4971 0.5067 0.6060 0.7276 0.5212 0.5250 0.3789 0.6061
0.8
1.0
0.8 Polymeric phase hold-up
Polymeric phase hold-up
0.6
0.4
0.2
Qs/Qp = 1 Qs/Qp = 1.3 Qs/Qp = 2 Qs/Qp = 4
0.6
0.4 Qs/Qp = 1.0 Qs/Qp = 0.75 Qs/Qp = 0.50 Qs/Qp = 0.25
0.2
0.0
0.0 6
8
10 12 rotation speed (rpm)
14
16
6
8
10
12
14
16
rotation speed (rpm)
Fig. 2. Dependence of polymeric phase hold-up on the rotation speed (Qs: flow rate of the saline phase [80 ml min1—fixed]); Qp: flow rate of the polymeric phase).
Fig. 3. Dependence of polymeric phase hold-up on the rotation speed (Qs: flow rate of the saline phase; Qp: flow rate of the polymeric phase [80 ml min1—fixed]).
drops tend to present higher upward velocity reducing the residence time in the extractor and consequently the hold-up, although insignificant variation in polymeric hold-up was observed. The hold-up decreased with increase of the flow rate relationship and with the decreased flow rate of the polymeric phase. This behavior is probably generated by the presence of smaller rates of polymeric phase in the equipment. Eq. (1) shows that the saline phase hold-up under the same operational conditions would be larger than that in the polymeric phase, also presenting inverse behavior when this phase is maintained fixed. Fig. 3 shows the effect of the rotation speed on the polymeric phase hold-up for different flow rate values of the saline phase at the polymeric phase flow rate of 80 mL min1. In this case, the polymeric phase holdup decreased on the increase of the saline phase volume inside the extractor. Fig. 3 shows that the hold-up was practically constant at the different rotation speeds tested here, except when the flow rates ratio was
very low. An example is the relationship of 0.25 (20/80) ml min1, for which the hold-up values between 0.62 and 0.73, showing a variation of 18%. Fig. 4 shows that the variation of the polymeric phase hold-up was maintained constant when the sum of the flow rates of the inlet streams in the equipment was kept constant. Changes in the rotation speeds did not affect the hold-up substantially. For different flow rate phases ratios, the hold-up remained in a narrow band. The rotation speed had a little influence on the holdup for the systems studied in this work. This behavior can be attributed to the fact that the Graesser contactor contains two independent flowing phases with an interface line that can be controlled in the central line of the contactor, establishing a small band of variation of the hold-up values. This behavior was also observed by Coimbra et al. (1994). Sheikh, Ingham, and Hanson (1972) described a behavior for the hold-up similar to that observed in this study. The authors analyzed the influence of a Graesser
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the continuous separation of a-lactalbumin and b-lactoglobulin from whey with ATPS, obtaining satisfactory yields at a rotor speed of 2 rpm. Recently, Jarudilokkul et al. (2000) employed a Graesser contactor to separate lysozyme from egg white using reverse micelles, also obtaining the optimal conditions for the extraction of lysozyme at a rotor speed of 5 rpm, and a phase flow relation of 60:20 ml min1, indicating low rotor speeds as good operational condition for this equipment.
Polymeric phase hold-up
0.8
0.6
0.4
0.2
60/60 70/50 80/40 40/80
3.3. Correlation
0.0 6
8
10 12 rotation speed (rpm)
14
16
Fig. 4. Dependence of polymeric phase hold-up on the rotation speed. Total flow rate constant (saline phase + polymeric phase) = 120 ml min1.
extractor (75 cm of length for 15 cm of diameter) rotation speed, using a system composed by water–n-butylamine– kerosene. An increased disperse phase hold-up with increased flow rate relations was verified, which is an opposite tendency to our results. This behavior can be expected since the hold-up is very dependent on this type of extractive system, physical properties, type of equipment and operational conditions. The physical properties of the ATPS studied are very different from the organic liquid–liquid extraction systems, mainly regarding viscosity and interfacial tension values. In the vertical columns, the rise of hold-up with the increase of the rotation speed and flow rate relation has been previously observed. For example, Coimbra et al. (1998), studying the hydrodynamic of a perforated rotating disc contactor (PRDC) using ATPS formed by PEG 6000 and dibasic potassium phosphate, observed an increase in hold-up with increased rotating disc speed and phases flow relation. Hamidi, Van Berlo, Luyben, and Wiwlen (1999) found polymeric phase hold-up values ranging from 0.1 to 0.9 and also observed an increase in hold-up with increased phases flow relation for a countercurrent sieve-plate column with ATPS composed by PEG 400 and Na2SO4. For a Karr extractor, Kader (1985) and Aravamudan and Baird (1999) verified the increase in hold-up with increased agitation speed for the systems acetone–toluene– water, kerosene oil–water and isopropyl alcohol–water. Among the different flow rate of phase tested, the interface in the extraction zone was not clearly observed at 15 rpm. The interface that should be controlled in the central line of the contactor could only be maintained in the separation zones. Wang, Ingham, and Hanson (1977) studied axial mixing and mass transfer in a Graesser contactor (15 cm diameter, 75 cm long) changing the rotor speed from 4 to 13.3 rpm, low rotor speed and low (solvent/aqueous) flow ratio being carried out to best overall mass transfer performance. Coimbra et al. (1994), also using a Graesser contactor, studied
Aiming to compare our hold-up data with those predicted by correlations developed for conventional liquid–liquid systems, we used the Richardson and Zaki (1954) correlation. This equation is applied to various types of extractors, except the Graesser contactor. The hold-up values obtained experimentally were used to determine the slip velocity (Vslip) according to Eq. (2). The slip velocity is the single most important variable controlling the mass transfer coefficients when a solute is transferred between the two phases (Misek, 1994). The direct measurement of slip velocity in the extraction columns is difficult, so it may be obtained from the flow rates of the continuous and dispersed phases and dispersed-phase hold-up. According to Slater (1985), a better slip velocity equation, first formulated by Richardson and Zaki (1954) is V slip ¼ V 0 ð1 H Þ
c
ð2Þ
V0 and exponent c are parameters calculated by the least-squares method. Within limits of 0 < H < 0.3, it was demonstrated that the slip velocity in various types of extraction equipment can be described by this empirical equation. In this work, Eq. (2) was correlated to the experimental data. Figs. 5–9 shows the experimental and correlation results for all the tested conditions.
0.001
6.6 rpm 10 rpm 12.5 rpm 15.5 rpm predicted 6.6 rpm predicted 10 rpm predicted 12.5 rpm predicted 15.5 rpm
0.0008
Vslip (m s-1)
306
0.0006
0.0004
0.0002 0.3
0.4
0.5
0.6
0.7
0.8
1—H
Fig. 5. Correlation of Eq. (2) to the experimental data for the polymeric phase (fixed).
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0.001 0.0008
0.0006
Vslip exp (m s-1)
0.0012
Vslip (m s-1)
0.0007
6.6 rpm 10 rpm 12.5 rpm 15.5 rpm predicted 6.6 rpm predicted 10 rpm predicted 12.5 rpm predicted 15.5 rpm
0.0014
307
0.0006
0.0005
0.0004 0.0002 0.0004 0 0.1
0.2
0.3
0.4
0.5
1—H 0.0004
0.0005
Fig. 6. Correlation of Eq. (2) to the experimental data for the saline phase (fixed).
0.0006
0.0007
Vslip cal (m s-1)
Fig. 8. Slip velocity: observed and correlated (Eq. (2)) values for polymeric phase—fixed (R2 = 0.77).
0.0007
Vslip exp (m s-1)
Vslip exp (m s-1)
0.000495
0.0006
0.000490
0.0005
0.000485
0.0004 0.0004
0.0005
0.0006
0.00050
0.0007
0.00052
0.00054
Vslip cal (m s-1)
Vslip cal (m s-1)
Fig. 7. Slip velocity: observed and correlated (Eq. (2)) values for saline phase—fixed (R2 = 0.95).
Fig. 9. Slip velocity: observed and correlated (Eq. (2)) values for the sum of the flow rate constant (R2 = 0.75).
According to Figs. 4 and 7, the validity of Eq. (2) to represent our experimental data has been confirmed for small hold-up values, such as the saline phase-fixed hold-up values, whose average value of hold-up was of 0.42. The mean value was obtained from an arithmetic average of measurement at each condition. The good agreement between calculated and experimental values is shown in Fig. 7, with the residual analysis for Vslip resulting in R2 higher than 0.95. For hold-up value means of 0.60 and 0.50 corresponding to polymeric phase-fixed and the sum of the
flow rate constant, respectively, the validity of the eq. (2) to represent our experimental data has not been confirmed, as can be seen in Figs. 6, 7 and 9. In Figs. 8 and 9 can be observed the residual analysis for Vslip with R2 less than 0.77 and 0.75, respectively. The difference between experimental data and correlated data is probably due to the fact that the hold-up values for our systems are not in the region of validity of Eq. (4). Also the systems used in this work are quite different from the water–organic solvent systems used to develop the correlations given in this work, such as very small
Table 3 Values of exponent c and characteristic velocity V0 Condition
Flow rate 6.6 rpm
Qs—fixed Qp—fixed Qs + Qp = 120
10 rpm 1
12.5 rpm 1
15.5 rpm 1
c
V0 (m s )
c
V0 (m s )
c
V0 (m s )
c
V0 (m s1)
0.9910 3.6224 0.0347
0.0003 0.0123 0.0005
0.9133 1.4530 0.0347
0.0003 0.0022 0.0005
0.9654 3.3617 0.0347
0.0003 0.0122 0.0005
1.0701 0.8123 0.0347
0.0003 0.0011 0.0005
308
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interfacial tension, small density and a high viscosity difference between the dispersed and continuous phases. This is uncommon in the majority of the organic liquid–liquid extraction systems. Differences in their hydrodynamic behaviors are thus likely to be expected. The exponents c and V0 resulting from the correlation of all conditions are given in Table 3. Kumar and Hartland (1994) found values of Vslip ranging from 0.0022 to 0.256 m s1 for spray columns, 0.0037 to 0.3178 m s1 for rotating discs and asymmetric rotating disc columns, 0.0025 to 0.110 m s1 for Kunhi columns, 0.0032 to 0.1464 m s1 for pulsed perforated-plate columns and 0.0084 to 0.1156 m s1 for Karr reciprocating columns. Godfrey and Slater (1991) found values for c ranging from 0.3 to 1.5 for packed columns, 0 to 4 for rotary agitated columns and 3 to 1 for sieve plate columns. These values were obtained for conventional liquid–liquid systems (water–organic solvent). Hamidi et al. (1999) found values for c ranging from 0.27 to 0.16 for countercurrent sieve plate column for aqueous two-phase systems composed by PEG 4000 and Na2SO4 and 0.31 for the V0 value. The values obtained in this work for Vslip ranging from 3.88 · 104 ml min1 to 6.93 88 · 104 ml min1 and for c values the range obtained was from 0.9133 to 3.6224. The differences in the Vslip and c values can be expected due to the different difference of type of equipments and physical properties of the extraction systems used in this work.
4. Conclusions For a constant flow rate of the saline phase, a tendency of the hold-up decrease on the increase of phase flow ratio was observed. When the polymeric phase was maintained fixed, hold-up was also decreased with the increase of the phase flow ratio. The rotation speed of 6.6 rpm and the flow rate ratio of 80/40 ml showed optimal conditions for the extractor operation regarding the best interface control. The correlation of slip velocity and hold-up using Eq. (2) is possible for the Graesser contactor using aqueous two-phase systems for small hold-up values, with the residual analysis for Vslip resulting in R2 higher than 0.95.
Acknowledgement The authors thank FAPEMIG, PADCT/CNPq (62.0167/97-1) and CAPES, for the financial support.
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