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Streptomycin Production via Fermentation – Process Modeling and Techno-

Economic Assessment (TEA) using SuperPro Designer.

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 Production of Streptomycin
Process Modeling and Evaluation with

SuperPro Designer®
by

Amir Mustafa, Rafael da Gama, Nikiforos Misailidis and Demetri Petrides

January 2022

This is the ReadMe file of a SuperPro Designer example that deals with process modeling, cost analysis
and optimization of Streptomycin produced by Streptomyces griseus. The flowsheet of the process is
appended to the bottom of this document. You may test-drive the model by downloading the functional
evaluation edition of SuperPro Designer from the downloads page of our website (www.intelligen.com).
The files of this example can be found in the Examples \ Pharmaceuticals \ Streptomycin folder. The
default installation path of the SuperPro Designer Examples folder follows below.

C:\ Users \ Public \ Public Documents \ Intelligen \ SuperPro Designer \ v12 \ Process Library \ Examples

If you have any questions about this example and SuperPro Designer in general, please send an email
message to dpetrides@intelligen.com

INTELLIGEN, INC.
Simulation, Design, and Scheduling Tools
For the Process Manufacturing Industries
www.intelligen.com
Introduction

Penicillin's success in fighting bacterial infections motivated many scientists and researchers to look for
additional antibiotics. One such endeavor in the fall of 1943 by a scientist named Albert Schatz under the
supervision of Dr. Selman A Waksman of Rutgers University led to the discovery and isolation of
streptomycin. Waksman and his students have also been credited with discovering numerous other
antibiotics such as actinomycin, neomycin, clavacin, etc.

Waksman received the Nobel Prize in Physiology and Medicine in 1952 for the discovery of streptomycin.
Although Albert Schatz was the legal co-discoverer of streptomycin, he was not rewarded with the Nobel
Prize, but received a share of the drug's royalties [5].

Figure 1. Selman A Waksman, showing some of the antibiotics discovered in his laboratory [left] (Copyright,
Rutgers University). and Albert Schatz [right], February 1989.

Streptomycin produced by Streptomyces griseus (S. griseus) is a broad-spectrum antibiotic that is highly
effective against both Gram-negative and Gram-positive organisms. Streptomycin has been found to be
very useful in treating infections caused by Gram-positive bacteria such as Mycobacterium tuberculosis,
which are particularly resistant to penicillin. It is also useful in combating plant diseases caused by
bacteria because it acts systemically in plants.

According to the World Health Organization (WHO), streptomycin is the safest drug used to treat
tuberculosis. Streptomycin has been added to the WHO's list of essential medicines for public healthcare.

Streptomycin was the first antibiotic made from bacteria of the Streptomyces genus (actinomycetes).
Krinsky was the first scientist to isolate S. griseus from Russian soil during World War I. S. griseus is
common in the soil and is a Gram-negative bacterium. It produces gray mycelium during sporulation and

1
generates gray-yellow pigment when it grows in the form of a colony [1-3]. A Petri dish culture of S.
griseus is shown in Figure 2.

Figure 2. Antibiotic-producing strain of Streptomyces griseus.

Streptomycin is mostly produced as acidic salts of hydrochloric acid or sulfuric acid, as these salts are
easier to precipitate with ketones such as acetone. Some of the physical and chemical properties of
streptomycin sulfate are listed in Table 1 [3,6].

Table 1. Physical and chemical properties of Streptomycin.

Physical and Chemical Properties Streptomycin

Physical state, form, and Solid, hygroscopic white to off-white powder

appearance

Solubility Soluble in water (50mg/ml).

Odor Faint amine-like odor

Taste Slightly bitter

Chemical name D-Streptamine, O-2-deoxy-2-(methylamino)-α-L-

glucopyranosyl-(1→2)-O-5-deoxy-3-C-formyl-α-L-
lyxofuranosyl-(1→4)-N,N1-bis(aminoiminomethyl)-,sulfate (2:3)

Molecular formula (C21H39N7O12)2-3H2SO4

Molecular weight 1457.41

The chemical structural formula, 2D structure, and 3D structure of streptomycin sulfate are shown in

2
Figure 3 and Figure 4, respectively [7,8].

Figure 3. Chemical structure of streptomycin sulfate.

Figure 4. 2D and 3D structure of streptomycin sulfate.

3
Streptomycin Market Size
The worldwide streptomycin market is projected to reach $600 million by 2025 and grow at a compound
annual growth rate (CAGR) of 0.7% in the forecast period between 2020 and 2025 [9-11].

Some of the top key producers of streptomycin include:

• Astellas Pharma
• GlaxoSmithKline
• Dr. Reddy’s Laboratories
• Johnson and Johnson
• Merck (MSD)
• Gilead Sciences
• Lonza
• Eli Lilly
• AstraZeneca
• Takeda Pharmaceutical
• Thermo Fisher Scientific

Figure 5 illustrates the streptomycin market growth by region and clearly shows that the Asian and
Australian markets are experiencing the highest market growth while Africa and the Eastern European
region are experiencing very little or no market growth [11].

Figure 5. Global Streptomycin Market Growth by Region, 2019-2025.

Streptomycin is useful in treating tuberculosis, Mycobacterium avium complex, plague, rat bite fever, and
other infections. Antibiotics can also prevent many infections in certain cases, such as before a surgery.
They act quickly and some work within hours. Due to the increased investment in research and
development with new product developments, the streptomycin market is experiencing high growth. The

4
increasing use of antibiotics in cell culture laboratories offers opportunities for market development.
Because they suppress infections, antibiotics are often used in cell culture media. Streptomycin is one of
the most common antibiotics for cell culture. Many cell culture laboratories often add antibiotics and
antifungal agents to cell culture medium during primary cell culture preparation. The streptomycin market
is expected to grow considerably as cell culture research is rising remarkably [10].

Large Scale Production of Streptomycin

Although streptomycin may be produced using wild strains of S. griseus, the production yield can be
increased by mutation of the microorganism. The production yield can also be increased by optimizing the
culture media.

Spores of S. griseus maintained as soil stocks or lyophilized in a carrier such as sterile skimmed milk are
employed as stock culture. The inoculum of S. griseus is added into the sporulation medium at 28 ⁰C.

Figure 7. Growth of Streptomyces griseus.

The spores of S. griseus sporulate and build up the mycelial network in the flask or seed fermentor.
Endospores are formed under low nutrient availability.

A list of typical ingredients for fermentation media follows.

1. Carbon source: Monosaccharides like glucose are the best carbon source for microorganism
growth and streptomycin production. Oligosaccharides and polysaccharides such as maltose and
starch can also be used as the carbon source, but they give lower yield.

2. Nitrogen source: Soy meal, meat extract, ammonium salts, etc. serve as common nitrogen
sources.

3. Mineral source: small amounts of magnesium, calcium, potassium, sodium along with sulphates,
phosphates and chlorides are required for growth and product formation.

5
 4. Growth stimulating source: L-naphthalene, acetic acid, and phenylacetic acid function as growth-
stimulating factors in the production of streptomycin.

In addition to the sources listed above, compounds such as proline, oils, fatty acids, and antioxidants are
also used to speed up streptomycin production. Precursors are not required for the production of
streptomycin [3].

The fermentation process is aerobic, takes 5 to 7 days and goes through 3 phases as shown in Figure 6
[14].

Phase I
Rapid
growth and
formation of abundant
S. griseus cell mass occurs
during this phase.
Glucose is utilized slowly and low
production of streptomycin is witnessed.

Phase II
Streptomycin production takes place at a rapid
rate without increase in the mycelial growth.
Glucose and oxygen are required in large
quantities during this phase.

Phase III
Cells undergo lysis, requirement of oxygen decreases and the
contents of the medium including glucose get exhausted. Finally,
streptomycin production ceases.

Figure 6. Three phases of streptomycin fermentation.

After the completion of fermentation, the broth is acidified with sulfuric acid to produce streptomycin
sulfate. The mycelial biomass is removed by filtration and the solution is neutralized with sodium
hydroxide. The solution is loaded onto an ion exchange column that retains the product. The column is
then washed with water and the product is eluted with hydrochloric acid. The product solution is
concentrated under vacuum at about 60°C using an evaporator. The product is precipitated with methanol
and acetone. A basket centrifuge is used to recover the precipitate and the cake is washed with acetone
to remove impurities. The washed precipitate is freeze-dried. The purified product is recovered as
streptomycin sulfate [1].

6
Process Description

A conceptual process for producing streptomycin sulfate was modeled and economically evaluated in
SuperPro Designer to estimate the expected raw material requirements, process equipment capacities,
utility requirements, capital investment and production costs. The model was based on a fed-batch
upstream fermentation and a semi-continuous downstream purification train. The development of the
model was based on data available in the technical and patent literature supported by our engineering
judgment and experience with related processes.

For reporting and analysis purposes, the process flowsheet has been divided into three sections:

• Fermentation (black icons)

• Product Recovery (blue icons)
• Solvent Recycling (dark green icons)

Flowsheet sections in SuperPro are simply sets of related unit procedures (processing steps). For
information on how to specify flowsheet sections and edit their properties, please use the Help tool (Help
Index Section) or refer to Chapter 8.1 of the SuperPro manual provided in PDF format with the SuperPro
installation. The contents of each process section are described in greater detail next. The flowsheet of the
process is appended to the bottom of this document.

Fermentation

The preparation of the fermentation media solution is managed by a continuous Mixture Prep unit procedure
(P-1 / MX-101). Table 2 displays the composition of the media solution.

Table 2. Fermentation media composition.

Component Name Composition % (w//w)

Glucose 20

Soy Hydrolysate 2

Sodium Chloride - NaCl 0.2

Ammonium Sulfate – (NH4)2SO4 5

Water 72.8

To ensure that the prepared media is free of pathogens the prepared media solution is sterilized using a
continuous sterilizer (P-2 / PZ-101). The sterilized media is stored in a tank (P-3 / V-101). For the sake of

7
simplicity, a continuous storage tank (V-103) was used to represent the storage of the sterilized media with
an average liquid residence time of 24 hours, which is equal to the cycle time of the overall process (see
the Process Scheduling section for details on the cycle time of the overall process).

A seed train composed of three growth steps is used: the first step is carried out in batch mode in three 6L
shake flasks operating in staggered mode (P-5 / SFR-101); the second takes place in seven 800L
fermentors operating in staggered mode (P-6 / SFR-102); and the third step is carried out in seven 8000L
fermentors operating in staggered model (P-7 / SFR-103). The role of multiple equipment units operating
in staggered mode is explained later in the Process Scheduling section.

The bacterial growth for inoculum preparation is modeled by the following stoichiometric equation. The yield
of the reaction is assumed to be 70%.

200 Glucose → 15 S.griseus DCM + 185 H O 2 (1)

S. griseus grows for two days in shake flasks and for five days in each seed fermentor. The media solution
is distributed to the fermentors using a Flow Distribution procedure (P-4 / FDIS-102). The media demand
for each fermentation step is controlled by a Pull-In operation. The role of the Flow Distribution procedure
(P-4 / FDIS-102) is to sum up the media demands and back-propagate the total demand to the media
preparation procedure (P-1 / MX-101). All the seed and production fermentors are also supplied with
sterilized and compressed air using a continuous air filtration procedure (P-9 / AF-101), gas compression
procedure (P-10 / G-101) and a flow distribution procedure (P-11 / FDIS -101). The average aeration rate
is 0.2 VVM.

The bacterial growth in all seed fermentors is modeled by the following stoichiometric equation. The yield
of the reaction is assumed to be 97%.

100 Glucose + 90 O2 + 25 (NH4)2SO4 + 10 Soy Hydrolysate →

(2)

25 S.griseus DCM + 115 CO2 + 10 Streptomycin + 75 H2O

The production of streptomycin occurs in seven fermentors operating in staggered mode (P-8 / FR-101).
Mycelium growth and product formation in the production fermentors is represented by the following mass
stoichiometry, assuming a conversion of 98%. A heat of reaction of -3700 kcal / kg of consumed O2 at 28
⁰C was assumed.

100 Glucose + 90 O2 + 25 (NH4)2SO4 + 10 Soy Hydrolysate →

(3)

15 S.griseus DCM + 115 CO2 + 20 Streptomycin + 75 H2O

8
The fermentation in the production fermentors is carried out for six days. Most of the media solution (50,000
kg out of a total of 60,000 kg) is supplied in fed-batch mode. This is specified through the Fed Batch tab of
the fermentation operation dialog window (see Figure 7).

Figure 7. Fed-Batch tab of the fermentation operation dialog window.

Product Recovery

Upon completion of the fermentation step, the broth is sent to a reactor (P-13 / R-101) for acidification with
a 20% sulfuric acid solution, which leads to the formation of streptomycin sulfate. A six-hour reaction time
was assumed with a conversion of 98% represented by the following mass stoichiometry.

2.0 Streptomycin + 3.0 H2SO4 → 1.0 Streptomycin.3H SO 2 4 (4)

After the completion of acidification, diatomaceous earth (Celite) is added to the reactor to a final
concentration of 2% w/w. The role of Celite is to aid the subsequent filtration and removal of the mycelium
by a rotary vacuum filter (P-14 / RVF-101) [12]. The filter cake is washed with water to minimize product

9
loss. The filtrate stream (S-135) containing excess sulfuric acid is neutralized with sodium hydroxide in a
neutralizer (P-15 / V-102). The neutralization reaction is modeled by the following stoichiometric equation
with a conversion of 100% and a heat of reaction of -55.2 kJ/mol of NaOH at 25 ⁰C.

1.0 H2SO4 + 2.0 NaOH → 1.0 Na SO 2 4 + 2.0 H2O (5)

The neutralized solution (stream S-137)) which contains streptomycin sulfate in dissolved form along with
impurities is then stored in a flat bottom tank (P-16 / V-103) which feeds the ion exchange chromatography
column (P-17 / C-101). It was assumed that 96% of the streptomycin is retained by the ion exchange resin.
The column is washed with 1.2 bed volumes (BV) of USP water and eluted with 1.2 BV of 0.5M HCl solution.
The product is recovered in 0.5 BV of eluant. Finally, the resin is regenerated with 2 BV of 1M NaOH
solution.

It is worth mentioning that several unit procedures in the product recovery section including the ion
exchange column are scheduled to cycle independently of the main recipe and operate back-to-back.
Independent cycling of procedures can be identified with a clock icon ( ) on the lower left side of that unit
procedure. The concept of Independently cycling procedures will be discussed in greater detail in Appendix
I at the end of this document.

The product stream of the ion exchange column (S-142) contains a significant amount of hydrochloric acid
because the eluant is a 0.5M HCl solution. The solution is neutralized with NaOH in a continuous neutralizer
(P-18 / V-105). This neutralization reaction is modeled by the following stoichiometric equation with a
conversion of 100% and a heat of reaction -59.9 kJ/mol of NaOH at 25 ⁰C.

1.0 HCl + 1.0 NaOH → 1.0 NaCl + 1.0 H O 2 (5)

The neutralized solution (stream S-144) is stored in a continuous flat bottom tank (P-19 / V-104) with a
residence time of 24 h after which it is preheated to 55 ⁰C (P-20 / HX-104). A single-stage evaporator (P-
21 / EV-101) operating under vacuum at 60 ⁰C removes approximately 80% of the water reaching a final
streptomycin sulfate concentration of 28% w/w. Operation under vacuum is necessary in the evaporator
because streptomycin sulfate is sensitive to high temperatures [13,14]. The water vapor from the evaporator
is condensed in a cooling procedure (P-22 / HX-103) and is recycled back to the ion exchange column (P-
17 / C-101) through a centrifugal pump (P-23 / PM-101) and a custom mixer (P-24 / MX-103) that manages
the supply of the makeup water.

It is worth mentioning that virtual energy integration was specified between the pre-heater (P-20 / HX-104)
and the condenser (P-22 / HX-103) of the evaporator, which satisfies 100% of the heating requirement of
the preheater and 5.14% of the cooling requirement of the condenser. The concept of Virtual Energy
Recovery will be discussed in greater detail in Appendix II at the end of this document.

10
After evaporation, the concentrated streptomycin sulfate solution is mixed with methanol (P-25 / R-102) and
then precipitated by adding acetone to a final concentration of 20% w/w at 15 ⁰C. The crystallization reaction
is modeled by the below stoichiometric equation with a conversion of 99%.

Streptomycin Sulfate → Streptomycin-Cr (6)

Two reactor vessels are assigned to P-25 operating in staggered mode with a cycle time of 12 hours (cycling
independently of the overall process cycle time which is 24 hours). Detailed information on the subject is
provided in Appendix I.

A flat bottom tank (P-26 / V-106) functions as a buffer tank between the crystallizer and the basket centrifuge
(P-27 / BCFBD-101) which is used to recover the product crystals. The cake containing the crystals is
washed with acetone to remove impurities. The volume of the washing acetone is equal to three times the
volume of the cake. The LOD of the cake is 25%. Two basket centrifuges are assigned to P-27 operating
in staggered mode with a cycle time of 2 hours (cycling independently of the overall process cycle time
which is 24 hours). As mentioned above, detailed information on the subject is provided in Appendix I.

A solids bin (P-38 / SB-101) receives the washed crystals and supplies a freeze dryer (P-39 / FDR-101)
which outputs 2,846 kg/batch of dried streptomycin sulfate crystals as the final product.

Solvent Recovery

To minimize the need for fresh methanol and acetone in the process, the filtrate of the centrifuge (stream
S-157) which contains most of the solvents is stored in a flat bottom tank (P-28 / V-107) which feeds a train
of three continuous distillation columns (P-29 / C-102, P-30 / C-103, P-31 / C-104) that separate the
solvents. Most of the water (~ 60%) and all non-volatile materials end up in the bottom stream of the first
column (stream S-159) while the distillate stream (S-160) contains almost all the acetone, methanol, and
the remaining amount of water. The distillate of the first column is then fed to the second column (P-30 / C-
103) which removes the remaining water in the bottom stream. The distillate of the second column, which
contains only acetone and methanol, is fed to the third column (P-31 / C-104) which generates a distillate
containing 97% acetone and a bottom stream containing 62% methanol, the remaining being acetone and
a small amount of water. The bottom stream of the third column (stream S-163) is cooled to 40 ⁰C (in P-33
/ HX-102) and stored in a buffer tank (P-34 / V-109) which supplies methanol to the crystallizer. Makeup
methanol is added directly to the crystallizer. The distillate of the third column (stream S-164) is also cooled
to 40 ⁰C (in P-32 / HX-101) and stored in a buffer tank (P-35 / V-108) which supplies acetone to the
crystallizer and the basket centrifuge. A flow adjusting procedure (P-36 / FAD-101) manages the supply of
the acetone makeup.

11
Process Scheduling and Debottlenecking

The total batch time for this process is approximately 505 h (21 days). This is the time from the start of a
particular batch (i.e., media preparation, fermentation in shake flasks, etc.) to the end of that batch (drying
of streptomycin sulfate crystals). However, the process cycle time is only 24 h since most of the individual
procedures in this process are much shorter than the overall batch time, and several (staggered) equipment
items are used for those that are longer.

To visualize the process schedule, click Charts Equipment Occupancy Multiple Batches. This will
generate the Equipment Occupancy Chart (EOC) shown in Figure 8. The chart displays 21 consecutive
batches of streptomycin sulfate production; each color indicates a different batch. The two process sections
– Fermentation & Product Recovery section – are also indicated on the chart. The Fermentation section
shows that seven fermentors are employed in each seed fermentation step (SFR-102 and SFR-103)
operating in Stagger Mode (alternating from batch to batch). The same is observed for the main
fermentation step (FR-101). The allocation of multiple equipment units to a single procedure that operate
in staggered mode reduces the effective cycle time of that procedure and enables the overall process to
operate with a shorter cycle time. Allocation of staggered equipment is specified by right-clicking on the
procedure icon, selecting “Equipment Data” and focusing on the Staggered Mode variables on the lower
left corner of the dialog window. Detailed information on Staggered Equipment and Independently Cycling
procedures can be found in the “Miscellaneous Modeling Tips” section of the Industrial Enzymes example
ReadMe file in the Examples \ Bio-Materials \ IndEnzymes folder of SuperPro Designer. Information on
equipment operating in Stagger Mode also can be found in the Online Help Facility of SuperPro Designer
and Chapter 6 of the SuperPro Designer user manual.

12
 Product Recovery

Figure 8. Equipment Occupancy Chart (EOC) for the streptomycin sulfate manufacturing process. The solvent recovery equipment units that operate
in continuous mode do not appear on the EOC.

13
Figure 9. Operations Gantt Chart for the streptomycin sulfate manufacturing process

14
The operations Gantt diagram (in the style of MS Project) offers another way to visualize the execution of
a batch process. This chart shows detailed scheduling information for one or multiple batches. The Gantt
chart for a single batch is generated by selecting Charts Gantt Charts Operations GC. Figure 9
shows part of the the Gantt chart for the current example, which illustrates the scheduling of operations in
the seed fermentation and production fermentation procedures. The dark blue and cyan colored bars
represent the durations of procedures and operations, respectively.

The Gantt chart enables users to visualize the execution of a batch process in detail. It also makes batch
recipe editing easier. Double clicking on one of its bars opens the dialog of the corresponding entity (e.g.,
operation, procedure, recipe, etc.). The simulation calculations can then be redone, and the chart can be
updated by clicking on the refresh button of the chart ( )

In addition, SuperPro can export its scheduling data to MS Project by selecting File Export to MS
Project XML File. SuperPro can also export its recipe data to SchedulePro by selecting File Export to
SchedulePro’s Recipe DB. SchedulePro is a resource management, production planning and scheduling
tool available from Intelligen. Please consult the Help facility for information on these two exporting options
(HelpIndexExporting).

Material Balances

Table 3 shows overall process data such as batch size, annual production rate and number of batches per
year. This table was extracted from the RTF version of the Materials & Streams report, which can be
generated by selecting Reports Materials & Streams in the main menu bar of SuperPro. The format of
the report can be set using the dialog that appears when you select Reports Options in the main menu
bar. The total number of batches per year is calculated as 309, resulting in an annual production rate of
879,605 kg/yr.

Table 4, which was also taken from the Materials & Streams Report, displays the raw material requirements
in kg/yr, kg/batch, and kg/kg of MP (“MP” stands for main product, the purified streptomycin sulfate in this
case). It clearly shows that USP water and the solutions of HCl and NaOH used in the Product Recovery
section of the process are the dominant raw materials in terms of quantity. It is noteworthy that the
consumption of fermentation media is relatively low compared to the downstream solutions.

15
Table 3. Overall process data

OVERALL PROCESS DATA

Annual Operating Time 330 days

Unit Production Ref. Rate 879,605 kg MP/yr
Batch Size 2,846 kg MP
Recipe Batch Time 21.03 days
Recipe Cycle Time 1.00 day
Number of Batches per Year 309.00
MP = Flow of Component 'Plasmid' in Stream 'S-245'

Table 4. Material requirements

BULK MATERIALS (Entire Process)

Material kg/yr kg/batch kg/kg MP

(NH4)2SO4(aq) 1,029,760 3,332.56 1.17
Acetone 290,460 940.00 0.33
Air 30,655,005 99,207.13 34.85
Celite 404,957 1,310.54 0.46
CIP-Acid 738,492 2,389.94 0.84
CIP-Caustic 1,478,431 4,784.57 1.68
Glucose 4,119,289 13,331.03 4.68
H2SO4(aq) 196,081 634.56 0.22
HCl (0.5M) 27,465,094 88,883.80 31.22
Methanol 128,754 416.68 0.15
NaCl 41,193 133.31 0.05
NaOH (1 M) 37,816,625 122,383.90 42.99
NaOH (20% w/w) 937,944 3,035.42 1.07
Soy Hydrolysate 411,915 1,333.06 0.47
USP Water 22,000,463 71,198.91 25.01
Water 20,160,383 65,243.96 22.92
TOTAL 147,874,846 478,559.37 168.12

16
Cost Analysis

SuperPro Designer performs thorough cost analysis calculations, estimating the capital expenditure
(CAPEX) as well as operating costs (OPEX) of a project. It generates the following three pertinent reports
(via the Reports menu): the Economic Evaluation Report (EER), the Cash Flow Analysis Report (CFR),
and the Itemized Cost Report (ICR). Table 5 displays the Executive Summary of the Economic Evaluation
Report.

The total estimated capital investment for such a facility is approximately $78 million. This includes the
equipment purchase and installation costs; other costs related to plant construction; startup and validation
costs; and the working capital required for this project. Plant construction costs such as the cost of buildings
and piping are estimated through multipliers in SuperPro Designer, and these have been modified in this
example to more accurately represent the capital costs associated with an antibiotic manufacturing facility.
For more information on this topic, please refer to the Industrial Enzymes example located in the
“…Examples \ Bio-Materials \ IndEnzymes” folder or consult the Help facility (under the topic “Capital
Investment Dialog: DFC Tab”).

Table 5 also shows the annual operating cost (AOC) and the unit production cost, which are $43 million
and $49/kg of streptomycin sulfate, respectively. Assuming a sales price of $75/kg of streptomycin sulfate,
a gross margin of 34.5% and a return on investment (ROI) of 30.7% can be expected for this project.

Table 5. Executive summary for the streptomycin manufacturing process

EXECUTIVE SUMMARY (2021 prices)

Total Capital Investment 77,703,000$

Capital Investment Charged to This Project 77,703,000$
Operating Cost 43,212,000$/yr
Savings (due to Heat Recovery) 7,980$/yr
Revenues 65,970,000$/yr
Batch Size 2,846.62kg MP
Cost Basis Annual Rate 879,605kg MP/yr
Unit Production Cost 49.13$/kg MP
Net Unit Production Cost 49.12$/kg MP
Unit Production Revenue 75.00$/kg MP
Gross Margin 34.51%
Return On Investment 30.74%
Payback Time 3.25years
IRR (After Taxes) 21.73%
NPV (at 7.0% Interest) 88,943,000$
MP = Total Flow of Stream 'Streptomycin'

17
Figure 10 displays a breakdown of the operating costs. This chart was copied from the Economic Evaluation
Report. Charts can be included in the reports by selecting Reports Options and checking the “Include
Charts” box on the lower right corner of the dialog.

Facility-dependent, raw materials, and labor-dependent costs are the largest components of the operating
costs, accounting for 32%, 31% and 22% of the AOC, respectively.

Figure 10. Annual operating cost breakdown for the streptomycin manufacturing process

Table 6 provides a breakdown of the operating costs per section. It was taken from the Itemized Cost Report
(ICR). The Product Recovery section is the most expensive, accounting for 51% of the AOC. It is closely
followed by the Fermentation Section, which is responsible for 45% of the AOC. The Solvent Recycling
Section accounts for only 4% of the overall operating cost.

Table 6. Breakdown of the annual operating cost by cost categories and process sections

4. BREAKDOWN PER COST ITEM AND SECTION (1000$/year)

Consum- Lab/ Waste

Section Materials Facility Labor Utilities TOTAL %
ables QC/QA Trt/Dsp
Fermentati 3,366 7,594 5,025 0 754 2,760 38 19,537 45.21
on
Product
9,664 5,795 3,848 1,360 577 207 646 22,099 51.14
Recovery
Solvent
214 250 668 0 100 284 13 1,528 3.54
Recycling
Auxiliary
0 48 0 0 0 0 0 48 0.11
Equipment
TOTAL 13,244 13,687 9,541 1,360 1,431 3,251 697 43,212 100.00

18
It is worth mentioning here that the cost of several equipment items such as fermentors, ion exchange
columns, crystallization/sulfation reactors and freeze dryers are based on user defined cost models. A
detailed discussion about the concept of user defined cost models can be found in the Appendix III at the
end of this document.

Summary
Our goal with this example was to present a simple streptomycin production model in SuperPro Designer
that is easy to understand and follow. As indicated in the above analysis, a plant with capacity of around
879 metric tons of streptomycin sulfate per year requires a CAPEX of around $78 million and annual
operating expenditures (including depreciation) of around $43 million, resulting in a unit production cost of
$49/kg. The facility-dependent expenses, raw materials, and labor are the dominant items of the
manufacturing cost.

Appendix I – Independently Cycling Procedures in Batch Processes

If a batch (cyclical) procedure operates autonomously with its own rhythm (cycle time) that is not linked to
the cycle time of the overall process, it is modeled in SuperPro by selecting the Cycles Independently of
Main Recipe checkbox in the procedure data dialog (see Figure 11) which is brought up by right-clicking
on the procedure icon and selecting Procedure Data. In this model, the ion exchange columns (P-17 / C-
101), the crystallization reactors (P-25 / R-102) and the basket centrifuges (P-27 / BCFBD-101) operate in
that mode. More specifically, the ion exchange procedure (P-17 / C-101) utilizes three columns operating

in staggered mode (staggered mode is indicated by the Staggered Equipment icon, ) with an
independent cycle time of 2 hours, which is equal to the duration of the LOAD operation. Each column
goes through the following set of operations: Loading for 2 hours, Wash for 42 minutes, Elution for 52.5
minutes, and Regeneration for 70 minutes. As soon as the Load operation of the first column is
completed, the inlet flow is directed to the second column while the first undergoes Wash, Elution and
Regeneration. When the Loading operation of the second column is completed (after 2 hours), the inlet
flow is directed to the third column. By the time the Loading of the third column is completed, the first
column is available to be reloaded. Figure 12 displays the operation of the three columns as a function of
time. The Cycle Duration of each column (the sum of the Load, Wash, Elution, and Regeneration
operation times) is 4.74 h. The availability of three 3 column operating in staggered mode with a cycle
time of 2 h enables the chromatography columns to handle the continuous inlet flow and therefore
operate continuously from a macroscopic point of view. The 2 h cycle time specified for this procedure
(Figure 11) represents the time between consecutive chromatography cycles (batches), which is different
from the overall recipe cycle time of 24 h determined by the production fermentors. This way each
fermentation batch is processed by 12 ion exchange cycles (batches). It is possible for the user to specify
that ratio instead of the independent cycle time of an autonomous procedure (see Figure 11).

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Similarly, the two crystallization reactors (P-25 / R-102) operate in staggered mode and independently
cycle every 12 h, which is equal to the duration of the TRANSFER-IN operation that receives the material
from the evaporator. Consequently, each fermentation batch is processed by 2 crystallization cycles
(batches). Finally, the two basket centrifuges (P-27 / BCFBD-101) operate in staggered mode and
independently every 2 h. Consequently, each fermentation batch is processed by 12 basket centrifugation
cycles (batches).

NOTES

1) A clock icon ( ) is visible on the lower left side of each unit procedure that has been set to cycle
independently.

2) Independently cycling procedures do not appear on the Gantt chart. However, their equipment
occupancy is displayed on the EOC. Also, operations of regular procedures in the flowsheet cannot
be scheduled relative to operations of independently cycling procedures.

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Figure 11. The Independent Cycling specifications for the Ion Exchange Columns (P-13/C-101)

Load Wash Elution Regeneration

Appendix II – Virtual Energy Recovery

Figure 12. The EOC of the Ion Exchange Columns (P-13/C-101) that cycle independently of the main
process. The (i/c) symbol next to the equipment names denotes the independent cycling.

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Appendix II – Virtual Energy Recovery

This model has many operations that either consume or generate heat. To improve the efficiency and
economics of this process, heat can be exchanged between hot streams (heat sources) that require
cooling and cold streams (heat sinks) that require heating. This reduces the overall usage of the utility
and associated utility costs. There are two ways to carry out heat exchange in SuperPro Designer:

1) Add heat exchange procedures directly to the flowsheet by selecting Unit Procedures \ Heat
Exchange \ Heat Exchanging from the main menu, connecting the relevant input and output
streams, and defining operational data for the procedure.

2) Define the heat recovery network by right-clicking on the flowsheet in an open area and selecting
Energy Recovery.

The first method above is preferred when only a few heat exchangers are used or if it is desired to
explicitly define the operating and/or equipment parameters for the exchangers. An example of the use of
this heat exchange setup can be found in the Ethanol example located in the Examples \ Bio-Fuels
folder. That example includes heat exchangers that are used to pre-heat cool streams while
simultaneously cooling hot streams (e.g., procedures P-7 and P-20 in the Ethanol flowsheet).

The second method above is preferred when many heat exchangers would be necessary. This method
allows the user to define the heat recovery network in a faster and easier manner. In addition, there are
fewer streamlines and unit procedure icons on the flowsheet, and the entire heat recovery network can be
easily viewed and modified in a single dialog. This method was applied to this example, and it is
described in greater detail below.

To access the Energy Recovery window, right-click on an empty area of the flowsheet and select Energy
Recovery from the menu that appears. The following dialog window will open:

Figure 13. Energy Recovery dialog window.

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This window shows all the operations in the model that require cooling, their required cooling loads and
their inlet and outlet temperatures. For example, the COOL-1 operation in procedure P-22 requires
14,495,000 kcal/batch (or 604,000 kcal/h) of cooling to achieve an outlet temperature of 50 oC from an
inlet of 60 oC.

This window makes it possible to match operations that require cooling with operations that require
heating, thus reducing the need for heating and cooling utilities. The operations that require heating can
be viewed by clicking the Show Recipients of Energy Recovered >> button at the bottom left of the
dialog in Figure 13. This will expand the Energy Recovery dialog, as shown in Figure 14:

For instance, the COOL-1 operation in P-22 requires 14,495,000 kcal/batch for cooling the relevant
stream from 60 to 50 oC, provided by cooling water. Meanwhile, the HEAT-1 operation in P-20 requires
744,896 kcal/batch to heat up the stream from 31.9 to 55 oC using steam. This operation can be
adequately satisfied by the cooling load and the temperature difference of the COOL-1 operation in P-22.

Figure 14. The expanded Energy Recovery dialog.

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To set up a heat exchange match between two operations, it is first necessary to click the checkbox in the
Recovered column in the upper table of Figure 14 for the appropriate Heat Source. Then the ellipsis

button ( ) in the View/Edit column in the Energy Recovery dialog is enabled. For instance, clicking
the ellipsis button for the P-22 / COOL-1 operation brings up the following dialog:

Figure 15. The Energy Matches dialog for the P-22 / COOL-1 operation.

Figure 15 shows the available operations that could potentially act as heat recipients for the P-22 /
COOL-1 operation. To select the heat recipient(s) to match with the COOL-1 operation in P-22, simply
click the checkbox in the Match column for the desired recipient operation, as shown in Figure 15.

For each operation that is matched, the “Match %” is calculated and an indication of the energy load that
has been matched so far, as well as the remaining load that can still be recovered with additional
matches. In this example 5.14% of the cooling load required by the COOL-1 operation of P-22 is provided
by operations HEAT-1 of P-20 (see Figure 14). Accordingly, the entire heating load of the HEAT-1
operation of P-20 is provided by the COOL-1 operation of P-22. This match results in an annual utility
savings of $7787. The dollar value of savings is reported underneath the UTILITIES COST table of the
Economic Evaluation Report which can be generated by selecting “Reports \ Economic Evaluation …”
from the application’s main toolbar.

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Note that excess heat from operations that require cooling can also be matched to heating agents. For
example, the heat from the P-22 \ COOL-1 operation could be used to generate Hot Water which in turn
can supply heat to other operations. To specify this, first click the “Match with spent heating agents”
option on the right side of the Energy Matches dialog shown in Figure 15. This will bring up the dialog
shown in Figure 16. Then select the appropriate row of the table (in this case, only the Hot Water option is
available) and click the Select button. This will cause the heat from the P-22 \ COOL-1 operation to be
used to produce the Hot Water utility. Note that heat from multiple hear sources can be used to generate
the same utility.

If there is a demand for the recipient utility agent in the process, then, matches with utility agents result in
savings. If the available heat exceeds the demand for the recipient agent, the excess agent generated
can be sold. The selling price is specified in the Properties Dialog of the agent (it corresponds to the
Agent Credit Price). Selling of the excess agent is considered when the “Use Excess Heat as Credit”
check box for that agent is checked (lower-right corner of Figure 16).

Figure 16. Matching a cooling operation with a heating agent.

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Appendix III – User Defined Equipment Cost Models

SuperPro Designer facilitates estimation of capital and operating costs and performs a thorough project
economic evaluation, profitability analysis and cash flow analysis of simple or integrated manufacturing
processes. The purchase cost of equipment (PC) is an important parameter that affects the direct fixed
capital investment of a project and, indirectly, its operating costs. SuperPro is equipped with correlations
to estimate the purchase cost of a great number of equipment types. However, the built-in correlations for
most types of vessels, membrane filters, chromatography columns and various other types of equipment
are more suitable for facilities manufacturing high-value biopharmaceutical than for commodity antibiotics.

For commodity bioproducts we recommend users to enter their own equipment cost data for better
accuracy. A good source of equipment cost data for such processes is available in the literature. [15]

Figure 17 displays the Purchase Cost tab of the equipment data dialog for an equipment item (fermentor
FR-101 in this case). This tab can be accessed by right clicking the corresponding procedure’s icon,
selecting Equipment Data, and then switching to the Purchase Cost tab. As you can see, the cost
estimation options include a user-defined purchase cost, estimation through the built-in cost model, and a
user-defined cost model.

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Figure 17. Equipment Purchase Cost Tab.

A new custom cost model can be specified by selecting the User-Defined Cost Model option and clicking
on the Parameters button. This brings up the dialog shown in Figure 18, which allows the user to specify
cost correlations in a power-law format (i.e., the dark red equation at the top of the window). The parameters
of the equation are as follows:

• C0 is the base cost,

• Q0 is the base capacity, and
• Q is the actual capacity, and
• a is the exponent of the power law
The capacity range can be broken down into any number of intervals.

27
Figure 18. User-Defined Equipment Cost Correlation Dialog.

For this example, the cost model parameters shown in Table 7 were used for various types of equipment.
Equipment not listed in this table, such as basket centrifuge, storage tanks & distillation columns employed
SuperPro’s built-in cost models.

Table 7. User-defined cost model parameters for the streptomycin example.

Equipment Capacity range (m3) Base Capacity Base Cost ($) Exponent
(m3)
Low End High End (Qo) (Co) (a)
Seed Fermentors 0 50 1 $60,000 0.6
Main Fermentors 0 25 5 $40,000 0.6
25 350 25 $250,000 0.7
Reactors 0.3 3 3 $75,000 0.4
3 140 3 $75,000 0.53
Ion Exchange Columns 0 20 5 $500,000 0.6
Freeze Dryer (kg) 0 1000 900 $1500,000 0.7

The user-defined cost models for each type of equipment can be stored in the user database, so that users
can quickly re-use the same data in multiple models by sharing a common User Database. Figure 18

28
displays the relevant buttons( ) for access the User DB data. To retrieve a cost model from the
user database, check the box Link to Cost Model. The latter option requires the user to define the cost
model in the user databank before it can be retrieved. This option is grayed out if no compatible equipment
cost model exists in the database. An advantage of defining the cost model in the database is that the data
can be retrieved by multiple equipment items of that type, which may exist in multiple SuperPro files
developed by various SuperPro users. As a result, using the user cost model database can ensure
consistency in equipment cost calculations across many projects. For more information on the user
databanks of SuperPro, please consult the corresponding sections of the SuperPro manual and the help
facility, as well as the SynPharmDB ReadMe file of the SynPharm example in the Examples \
Pharmaceuticals folder.

SuperPro is also equipped with databases for storing cost and other data for equipment of specific sizes
that can be purchased from various vendors. That data becomes accessible to a piece of equipment in a
file by matching it with suitable equipment in the DB through the Allocation tab of the Equipment Data
dialog of an equipment item. For further information on this subject, once again consult the corresponding
sections of the SuperPro manual and the Help facility, as well as the SynPharmDB ReadMe file of the
SynPharm example.

The purchase costs of all equipment of a Section are summed up and multiplied by installation and other
factors to come up with the Direct Fixed Capital. Like the default cost models for individual equipment items,
the default multipliers for estimating the fixed capital investment are appropriate to fine chemical and
pharmaceutical types of facilities. It is therefore important to modify these multipliers for other types of
processes, to avoid greatly overestimating the fixed capital investment. The default multipliers and the
multipliers used in this example are shown in Table 8:

Table 8. Fixed Capital Investment Multipliers.

Factor Multipliers Used in this Default Multipliers

Example File
Unlisted Equip. Purchase Cost 0.10 0.20
Piping 0.35 0.35
Instrumentation 0.30 0.40
Insulation 0.03 0.03
Electrical Facilities 0.10 0.10
Buildings 0.40 0.45
Yard Improvement 0.15 0.15
Auxiliary Facilities 0.40 0.40

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To edit the built-in multipliers for a specific section of the process, click the Section Capital Cost Adjustments

button ( ) on the Sections toolbar. The capital cost factors associated with that section can then be
modified (see Figure 19).

Figure 19. The Capital Cost Factors associated with the Fermentation section.

Note that the capital cost multipliers should be verified (and modified, if necessary) for each section of the

30
flowsheet. Also note that it is possible to re-use the same capital cost multipliers for many different projects
by associating them with a specific “site” and then allocating the process to that site. This is explained in
detail in the “Using Databanks to Facilitate Cost Analysis” section of the “SynPharmDB” document within
the SynPharm subfolder of the Examples \ Pharmaceuticals folder of your installation.

References

[1] Biosynthesis of Antibiotics, edited by J.F. SNELL Vol I. (Academic Press, 1966).

[2] Streptomycin: Structure, Biosynthesis, Process and Uses of Streptomycin | Biotechnology

https://www.biotechnologynotes.com/antibiotics/streptomycin/streptomycin-structure-biosynthesis-
process-and-uses-of-streptomycin-biotechnology/13848

[3] Production of Streptomycin

https://biologyreader.com/production-of-streptomycin.html

[4] Streptomycin: Structure, Biosynthesis, Process and Uses of Streptomycin | Biotechnology

https://www.biotechnologynotes.com/antibiotics/streptomycin/streptomycin-structure-biosynthesis-
process-and-uses-of-streptomycin-biotechnology/13848

[5] Streptomycin: Discovery and Resultant Controversy by Milton WainwrightSource: History and
Philosophy of the Life Sciences, Vol. 13, No. 1 (1991), pp. 97-124Published by: Stazione Zoologica
Anton Dohrn - Napoli

[6] Streptomycin sulfate

https://www.enzolifesciences.com/ALX-380-277/streptomycin-.-sulfate/

[7] Streptomycin Sulfate

https://www.drugs.com/pro/streptomycin-sulfate.html

[8] Compound Summary Streptomycin sulfate

https://pubchem.ncbi.nlm.nih.gov/compound/Streptomycin-sulfate

[9] Penicillin and Streptomycin Market Size 2021 Industry Share, COVID-19 Impact, Development
Strategy, Global Trend, Geographical Statistics, Growth Status, Manufacturing Cost Structure and
Future Investments Analysis Report 2027

https://www.marketwatch.com/press-release/penicillin-and-streptomycin-market-size-2021-
industry-share-covid-19-impact-development-strategy-global-trend-geographical-statistics-
growth-status-manufacturing-cost-structure-and-future-investments-analysis-report-2027-
2021-11-24

[10] Penicillin and Streptomycin Market - Forecast (2021 - 2026)

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 https://www.industryarc.com/Report/15050/penicillin-streptomycin-market.html

[11] Penicillin and Streptomycin Market By Product, By Manufacturing Process, By Route of

Administration, By End-User- Global Industry Share, Growth, Competitive Analysis and Forecast,
2019-2025

https://www.omrglobal.com/industry-reports/penicillin-and-streptomycin-market

[12] US Patent 2,701,795 ; STREPTOMYCIN EXTRACTION

Arne N. Wick and Milton J. Vander Brook, KalamaZ00, Mich., assignors to The Upjohn Company,
Kalamazoo,Mich., a corporation of Michigan No Drawing. Application December 12, 1945,
Serial No. 634,622 15 Claims. (CI. 260-210)

[13] The Effect of Heat on Streptomycin and Para-Aminosalicylic Acid in Tuberculosis Culture Medium,
MARGARET C. DRUMMOND, M.S., GEORGE T. LEWIS, Ph.D., MARTIN M. CUMMINGS, M.D.,
F.C.C.P. https://journal.chestnet.org/article/S0096-0217(15)31839-2/pdf

[14] Production of Antibiotics, In book: Fundamentals, Industrial and Medical Biotechnology.Chapter: 8;

Publisher: Universal Academic Services, Beijing.Editors: Ogbonna, Ubi, and Enweani

[15] DOE/NETL-2002/1169 (2002). Process Equipment Cost Estimation. Available at

http://www.osti.gov/bridge/purl.cover.jsp?purl=/797810-Hmz80B/native/

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