APPLICATION NOTE

imPULSE Single-Use Mixer, 2,000 L

Mixing efficiencies for the 2,000 L
imPULSE Single-Use Mixer
Introduction
The 2,000 L Thermo Scientific™ imPULSE™ Single-Use
Mixer (S.U.M.) provides innovative mixing for upstream
and downstream processes. The unique design of the
imPULSE S.U.M. produces more effective mixing at
lower working volumes than other mixers currently on themarket, and produces one-way flow mixing. The additionof the Touchscreen Console to the imPULSE S.U.M. adds
real-time sensor visualization, data logging, and the ability
to automate user-defined mixing recipes. To demonstrate
the superior mixing capabilities for various applications
and to determine optimal mixing parameters, four different
mixing studies (Table 1) were performed with the 2,000 L
imPULSE S.U.M. at various working volumes and mixing
settings. The materials used in these studies were chosen
to demonstrate the way buffers (NaCl solution), complex
media (Gibco™ AGT™ medium), and viscous liquids (cornsyrup) are mixed in the 2,000 L imPULSE S.U.M.
Materials and methods
A standard 2,000 L imPULSE S.U.M. was used with astandard 2,000 L Thermo Scientific™ imPULSE™ S.U.M.
BioProcess Container (BPC) modified with sampling andprobe ports at the bottom, middle, and top of the BPC.

The BPC was also equipped with a spray nozzle at thetop, to which a pump was connected, creating a liquid
recirculation loop. This pump was set to circulate waterfrom the bottom of the BPC through the spray nozzle at a
rate of 10 L/min during most of the mixing studies.
To test the wide range of applications of the 2,000 L
imPULSE S.U.M., mixing studies were performed atvarious working volumes. The manufacturer-recommended
maximum setting—the point where maximum agitation
occurs without excessive splashing—at various
working volumes was determined through visualinspection (Table 2).
Table 2. The manufacturer-recommended maximum setting
determined for the working volumes of 10:1, 5:1, 2:1, and full volume.

Working volume

Manufacturer-recommended
maximum setting (Hz)10:1 (200 L)

0.66

5:1 (400 L)

0.9

2:1 (1,000 L)

2

Full volume (2,000 L)

2

Table 1. Mixing studies performed on the 2,000 L imPULSE S.U.M.

Mixing type

Mixing material

Concentration

Analytics

Liquid–liquid

NaCl solution

350 g/L NaCl indeionized (DI) H₂O

Conductivity

Liquid–solid

NaCl granules

1 M (final)

Conductivity,
osmolality

Liquid–solid

AGT medium(granules)

1X (final)

Conductivity,
osmolality,
pH, glucose
concentration, visual

Viscous liquid–
liquidCorn syrup with
NaCl solution

80–88% corn syrup

Conductivity

A drawing showing the dimensions of the 2,000 L imPULSE
S.U.M. is shown in Figure 1.
384.43 cm (151.35 in)

2,000 L259.92 cm (102.33 in)
247.15 cm (97.30 in)

1,500 L

1,000 L142.64 cm (56.16 in)
303 L, middle of probe belt
287 L, max. height impeller
41 L, min. height impeller
0 L, tank bottom

90.79 cm (35.74 in)

30.87 cm (12.15 in)
20.32 cm (8.00 in)
15.34 cm (6.04 in)
12.70 cm (5.00 in)
0 cm (0 in)

Figure 1. Drawing of the 2,000 L imPULSE S.U.M.

Liquid–liquid mixing: NaCl solution
Three conductivity probes were inserted into the BPC in
the top, middle, and bottom ports. The imPULSE mixer
was filled to four separate working volumes: 10:1, 5:1, 2:1,
and full volume. At 10:1 and 5:1 working volumes, themixing settings of 0.25, 0.5, and 0.75 Hz were tested intriplicate. 0.75 Hz is above the manufacturer-recommended
maximum setting for the 10:1 volume; however, it wastested for comparative purposes. At 2:1 and full volume,the mixing settings of 0.25, 0.5, 0.75, 1, 1.5, and 2 Hz were
tested in triplicate.
The predetermined agitation was set on the 2,000 L
imPULSE S.U.M. and was allowed to mix for 2 minutes toallow mixing flow patterns to stabilize. A small volume ofconcentrated NaCl solution (350 g/L NaCl in DI H₂O) was
pipetted into the mixer as close to the surface of the water
as possible, over 3–4 seconds. Conductivity data wererecorded for 5 minutes with a data logger. This process
was repeated for each mixing setting at each volume intriplicate. An average T95 mixing time—when the measured
value reaches 95% of the final stable value—was calculated
as the average of the slowest-responding sensor among
the test replicates for each test condition.

Liquid–solid mixing: NaCl granules
Three conductivity probes were inserted into the BPC in
the bottom, middle, and top ports. The BPC was filled with
DI water to 10:1, 5:1, and full volume. For each working
volume, the manufacturer-recommended maximum setting
was used (see Table 2).
The predetermined agitation was set on the 2,000 L
imPULSE S.U.M. and was allowed to mix for 2 minutes.
Enough NaCl was quickly added to the top of the mixer
to make a 1 M NaCl solution. Samples were taken every
minute for the first 20 minutes, every 5 minutes for the next
20 minutes, every 10 minutes for the next 20 minutes, andevery minute for the last 10 minutes. Samples of 10 mL
were taken from the bottom, middle, and top ports for fullvolume, and just the bottom sampling port for 10:1 and
5:1 working volumes. Conductivity was measured online,
and osmolality was tested and recorded after samples
were collected.
Liquid–solid mixing: AGT medium
Two pH probes were inserted into the BPC in the bottom
and top sensor ports, and one conductivity probe inserted
in the middle sensor port. A camera was attached tothe top of the mixer to observe the time when foam andpowder from the medium were completely in suspension.
The BPC was filled with DI water to 90% of three working
volumes: 10:1, 5:1, and full volume. At each working
volume, the manufacturer-recommended maximum setting
was used.
The mixer was turned on to the predetermined agitation,and the manufacturer-recommended amount of AGT
medium was quickly added to the top of the mixer.
Samples were taken over a 70-minute time span using thesame method described in the previous section. Samplesof 10 mL were taken from the bottom, middle, and top
ports for full volume testing and just the bottom port for low
working volumes. The pH and conductivity were measured
online with the Touchscreen Console. The osmolality andglucose concentration of the samples were tested andrecorded after samples were collected.

Results
Liquid–liquid mixing: NaCl solution
For liquid–liquid mixing of a salt solution, T95 mixing
times were less than 4 minutes regardless of the working
volume or agitation rate, as shown in Figure 2. At 2:1 andfull volume, T95 mixing times decreased with increasing
agitation. However, at the low working volumes of 5:1 and
10:1, already rapid mixing times were not always decreased
with increased agitation.
From the data collected in this liquid–liquid study, it wasdetermined that at lower working volumes the mostaggressive agitation setting does not always correlate to
the fastest T95 mixing time. Table 3 shows the optimal
mixing setting to produce the fastest liquid–liquid
mixing times.

Table 3. The originally determined maximum agitation at all volumes
tested and the optimal agitation to produce the fastest liquid–liquid
mixing times.

Working
volumeManufacturerrecommended
maximum setting (Hz)Optimalliquid–liquidagitation (Hz)

10:1

0.66

0.5

5:1

0.9

0.5

2:1

2

2

Full volume

22

Liquid–solid mixing: NaCl granules
For liquid–solid mixing using salt granules, the conductivity
was measured for three working volumes, as shownin Figure 3. The T95 mixing times by conductivity were
2 minutes at full volume and 5:1 volume, and 8 minutes
at 10:1 volume. This mixing trend is different from theliquid–liquid mixing results observed previously, where T95
mixing times decrease with lower working volumes. It wasobserved during this mixing study that at the 10:1 working
volume a portion of the salt granules temporarily sat at thebottom of the mixer; this resulted in a longer T95 mixing
time. Similar results have been observed in the 50 L, 200 L,
and 2,000 L Thermo Scientific™ HyPerforma™ S.U.M.s
at low working volumes. Figure 4 shows the osmolality
recorded for the tested working volumes. The T95 mixing
time was reached in 2 minutes at full volume, 3 minutes at
5:1 volume, and 9 minutes at 10:1 working volume.
Liquid–solid mixing (NaCl granules): conductivity
90

Conductivity (mS/cm)

Viscous liquid–liquid mixing: corn syrup with
NaCl solution
Conductivity probes were inserted into the top, middle,
and bottom of the BPC. The BPC was filled initially with2,000 L of 88% corn syrup/12% DI water (v/v). A solutionof 350 g/L NaCl was made, and a 5 L mixture containing
12% of the salt solution and 88% corn syrup (v/v) was
prepared. The recirculation pump was not present during
this test. The agitation was set to 2 Hz, and the mixtureof salt solution and corn syrup was quickly added. The
viscosity of the 88% corn syrup solution in the imPULSE
S.U.M. was measured with a viscometer. The change
in conductivity was monitored for 15–30 minutes. This
process was repeated in triplicate at 84% corn syrup
and 80% corn syrup in the same BPC by draining apredetermined volume of the corn syrup solution and fillingit back up to full volume with DI water until the final desired
corn syrup percentage was achieved. The T95 mixing
times were calculated using the method described in theprior liquid–liquid mixing section.

80
70
60
50

T95 = 8 min

40

T95 = 2 min

30
20
10
0

0

10

20

30

40

50

60

70

80

Time (min)Full volume

5:1 volume

10:1 volume

Figure 3. Conductivity data recorded at full volume, 5:1, and 10:1.

Liquid–liquid mixing: NaCl solution

200
150
100
50
0

Liquid–solid mixing (NaCl granules): osmolality

2,500

250

0.25 Hz

0.5 Hz0.75 Hz

1 Hz

1.5 Hz

2 Hz

Agitation
Full volume

2:1 volume

5:1 volume

Osmolality (mOsm/kg)

T95 mixing time (sec)

300

2,000
1,500

T95 = 9 min
T95 = 3 min

1,000

T95 = 2 min
500
0

0

10

20

40

50

60

70

Time (min)10:1 volume

Figure 2. The average T95 mixing times (± 1 standard deviation)
calculated for four working volumes at varying mixing speeds.

30

Full volume

5:1 volume

10:1 volume

Figure 4. Osmolality recorded at full volume, 5:1, and 10:1.

80

1.2

350

1

300
250

0.8

200

0.6

150

0.4

100

0.2

50
0
0

20

30

40

50

60

Full volume
osmolality
5:1 volume
osmolality
10:1 volume
osmolality
Full volume
glucose5:1 volume
glucose0

10:1 volume
glucoseTime (min)Figure 5. Glucose and osmolality recorded at full volume, 5:1,
and 10:1.

Liquid–solid mixing (AGT medium): conductivity
Conductivity (mS/cm)

The conductivity T95 mixing times were 28 minutes at fullvolume, 2 minutes at 5:1 working volume, and 3 minutes
at 10:1 working volume (Figure 6). At low working volumes
there was a distinct upward shift in pH over time (Figure 7).
It is likely that this shift is a result of CO₂ degassing due tothe low working volume and rapid mixing. This shift is not
ideal, because cell metabolism can be altered by changes
in pH [1]. The pH shift can be corrected by the additionof acid or base. However, studies have shown that baseor acid addition can be correlated with process variability
and osmolality increase [2]. Generally, it is considereda best practice to avoid the addition of base or acid tocell culture media. During this study, the purpose wasto show how rapidly a medium could be incorporated,
and ideal conditions for cell culture medium preparation
were not followed. More ideal conditions for mixing cell
culture media are discussed in the conclusions andrecommendations section.

10

Normalizd glucose levels

Liquid–solid mixing (AGT medium): glucose and osmolality
400

Osmolality (mOsm/kg)

Liquid–solid mixing: AGT medium
For liquid–solid mixing of representative AGT medium
granules, the glucose and osmolality were measured offline
after samples were collected. The results of the glucose
concentration and osmolality are shown in Figure 5. At fullvolume, the glucose levels and osmolality had a T95 mixing
time of 30 minutes. At 5:1 and 10:1 working volumes, theglucose levels and osmolalities had T95 mixing times of
1–3 minutes.

9
8
7
6
5
4

T95 = 28 min

T95 = 3 min

3

T95 = 2 min

2
1
0
0

10

20

30

40

50

60

70

Time (min)Full volume

5:1 volume

10:1 volume

Figure 6. Conductivity data of the AGT medium recorded atfull volume, 5:1, and 10:1.
Liquid–solid mixing (AGT medium): pH
8.0
7.5
6.5
6.0

pH

The powder in this liquid–solid mixing study floated on
top of the fluid, and the settling that was observed in
the liquid–solid mixing with NaCl granules did not occur.
Therefore, the lower working volumes in this liquid–solid
mixing study had faster mixing times than full volume andthose observed in the NaCl liquid–solid mixing study.

5.5
5.0
4.5
4.0
3.5
3.0
0

10

20

30

40

50

60

70

Time (min)Full volume

5:1 volume

10:1 volume

Figure 7. pH of the AGT medium recorded at full volume, 5:1, and 10:1.

Figure 8 shows photos taken at various times during themixing study at full volume. After 46 minutes of mixing,
it was observed that the powder was completely insuspension and foaming was diminished. This correspondswith the data in Figures 5 and 6 that show osmolality,
conductivity, and glucose stabilizing in approximately thesame time frame.
Figure 9 shows photos taken at various times during themixing study at 5:1 working volume. After 2.5 minutes of
mixing, it was observed that the powder was completely
in suspension and foaming was diminished. This also
corresponds with the data in Figures 5 and 6.

Figure 8. Photos of the fluid at the top of the mixer at various time
points during the full-volume AGT medium mixing study.

Figure 10 shows photos taken at various times during themixing study at 10:1 volume. After 3 minutes of mixing,
the powder was completely in suspension and foaming
was diminished. This also corresponds with the data in
Figures 5 and 6 for the 10:1 volume.

Figure 9. Photos of the fluid at the top of the mixer at various time
points during the 5:1 volume AGT medium mixing study.

Figure 10. Photos of the fluid at the top of the mixer at various time
points during the 10:1 volume AGT medium mixing study.
1.4

Change in conductivtiy

Viscous liquid–liquid mixing: corn syrup with
NaCl solution
The measured viscosity of the corn syrup mixturescontaining 80–88% corn syrup by approximate volume
ranged from 336 cP to 881 cP, which is within therange of medium- to high-viscosity fluids seen in some
applications. The conductivity data recorded are shown in
Figure 11, with higher mixing times as viscosity increased.
The average T95 mixing times were determined to be
4 minutes for 80% corn syrup (345 cP), 8 minutes for
84% corn syrup (521 cP), and 28 minutes for 88% cornsyrup (861 cP). After the first two tests of the 88% cornsyrup testing, significant degradation of the mixing flaps
occurred, resulting in less than ideal mixing for the 88%,
84%, and 80% corn syrup.

1.2
1.0
0.8

T95 = 28 min

0.6

T95 = 8 min

0.4

T95 = 4 min

0.2
0.0

0

5

10

15

20

25

30

Time (min)
0% (1cP)

80% (345 cP)

84% (521 cP)

88% (861 cP)

Figure 11. Conductivity data recorded during viscous liquid–liquid
mixing with solutions of 0–88% corn syrup.

35

Conclusions and recommendations
It was found that the 2,000 L imPULSE S.U.M. exhibits
robust and efficient mixing for volumes ranging from 10:1 tofull volume at a wide range of mixing settings and withvarious mixing materials. It is capable of liquid–liquid mixing
at any working volume in under 4 minutes, mixing floating
solids at full volume in under an hour, and mixing highly

viscous liquids in under 30 minutes. In general, at lower
working volumes faster mixing was observed. However,mixing sinking solids was found to produce longer mixing
times at lower working volumes due to settling. Table 4shows when the solution reached a T95 mixing time basedon the analytic used.

Table 4. Average times for mixing of various materials in the 2,000 L imPULSE S.U.M.

Mixing study

Liquid–liquid

Liquid–solid

Mixing material

NaCl solution

NaCl granules

Working volume

Agitation (Hz)Analytic

T95 mixing
time (min)

10:1

0.5

Conductivity

1.0

5:1

0.5

Conductivity

0.9

2:1

2.0

Conductivity

0.4

Full volume

2.0

Conductivity

0.6

10:1

0.66

Conductivity

8.0

Osmolality

9.0

5:1

0.9

Conductivity

2.0

Full volume

10:1

Liquid–solid

AGT medium

5:1

Full volume

Viscous liquid

2.0

0.66

0.9

2.0

Osmolality

3.0

Conductivity

2.0

Osmolality

2.0

Conductivity

3.0

Osmolality

3.0

Glucose

1.0

Visual inspection

3.0

Conductivity

2.0

Osmolality

2.0

Glucose

2.0

Visual inspection

2.5

Conductivity

28.0

Osmolality

30.0

Glucose30.0

Visual inspection

46

High-viscosity liquid(700–1,000 cP)

Full volume

2.0

Conductivity

28.0

Medium-viscosity
liquid (200–400 cP)

Full volume

2.0

Conductivity

8.0

For optimal mixing, we suggest filling the mixer, setting
a desired agitation, and adding product incrementally.
For liquid–solid mixing with floating solids, it is bestpractice that a recirculation loop is added to the imPULSE
S.U.M. If a complex medium is being prepared at 5:1
working volume or lower, we recommend that the mixer
not be set to the maximum setting possible at that working
volume. Lowering the agitation rate, decreasing the overall
processing time, and paying attention to pH drift will
help ensure that the pH stays within range. Low working
volumes with sinking solids can be problematic, and longer
mixing times may be observed. For viscous liquids, thehigh-viscosity fluids can reduce the life of the agitator, andthe conditions tested in this study should be consideredworst-case. Material degradation of the mixing flaps waspresent throughout the viscous liquid–liquid mixing study.
It is recommended when mixing highly viscous fluids(700–1,000 cP) to use lower agitation for shorter periodsof time.

The 2,000 L imPULSE S.U.M. performs rapid mixing thatcan be used for a variety of applications with a wide variety
of materials ranging from liquids, floating solids, sinking
solids, and viscous fluids. The unique design of this mixer
allows it to mix effectively at low working volumes. Due tothe robust mixing, unique design, and addition of the
Touchscreen Console, the 2,000 L imPULSE S.U.M. canmeet the mixing requirements that are essential in today’scompetitive mixing industry.

References
1. Hagrot E (2011) Development of a culture system for modeling of pH effects in CHO
cells. Student thesis. KTH School of Biotechnology, Department of Process Technology,
Stockholm, Sweden.
2. Hoshan L, Jiang R, Moroney J et al. (2019) Effective bioreactor pH control using only
sparging gases. Biotechnol Prog 35(1):e2743.

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