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From Algae to a Functional Food

Project Team MembersTravis DallasEric Graves

Joaquin MartinezChristopher McNinch

Ramune Otterson-MeskyteCharu Saini

MentorsDan Rusinak, PE, Middough Inc.Dr. Jeffery Perl, PhD, PE, CHMM

Table of Contents

Executive Summary 3Discussion 4-5Conclusions and Recommendations 6Appendix 1 Design Basis 7Appendix 2 Chemistry of DHA 8-10Appendix 3 Block Flow 11-13Appendix 4 PFDs 14-38Appendix 5 Stream Information 39-53Appendix 6 Material and Energy Balance 54-55Appendix 7 Calculations 56-59Appendix 8 Annotated Equipment List 60-62Appendix 9 Economic Evaluation Factored from Equipment Costs

63-67

Appendix 10 Utilities 68-70Appendix 11 Conceptual Control Scheme 72-92Appendix 12 General Arrangement – Major Equipment Layout

92-95

Appendix 13 Distribution and End Use Review 96-97Appendix 14 Constraints Review 98-99Appendix 15 Applicable Standards 100-103Appendix 16 Project Communications 104-113Appendix 17 Information Sources and References 111-114

2

Executive Summary

AlphaAlgae has designed a plant that will produce a margarine fortified with a popular

omega-3 fatty acid, docosahexaenoic acid (DHA), from growing and cultivating a

specific algae species, Crypthecodinium Cohnii. The plant design incorporates all

aspects of functional food production, from growing the algae, to refining raw soybean

oil (SBO), to making the margarine, with each branch of the food production complying

with strict sanitary design guidelines. The final product, Golden Premium Spread, will be

the only margarine fortified with DHA available at mainstream health food markets in the

United States, allowing the product to obtain a large percentage of the market share

rapidly. 10.4 million tubs of margarine per year will be produced, with each tub having

426 grams of margarine. This means that the plant will produce about 2.3 million

kilograms of product in the first year.

This plant will be located in Cedar Rapids, IA, giving the facility easy access to a Cargill

ethanol plant. The Cargill ethanol plant will be utilized for margarine ingredients, food

for the algae, and waste disposal. AlphaAlgae’s Premium Golden Spread production

facility will cost $ 42 million to build and will cost $ 15 million per year in operating

costs. The payback period for this proposed facility is 5.7 years, and the net present

value of the project is $ 84,727,200 million.

3

Discussion

The health food market has exploded recently, from relatively niche corner stores to

mainstream grocery markets like Whole Foods. Along with the growth in the health food

market, the public’s knowledge and interest in having a healthy lifestyle has also

increased. Golden Premium Spread will fulfill the daily requirements for omega-3 fatty

acids and will be of interest to many consumers who shop regularly at health food

markets. Our ideal consumer population, people who are invested in healthy eating

habits and lifestyle, is increasing and very well informed of any new healthy eating

products.

The table spread market enjoys $ 5.2 billion in sales per year. It is a mature market that

has decreased slightly in the last decade due to concerns with the health of eating table

margarine. A new margarine product advertised as a healthy product will then be able to

obtain consumers who do not usually purchase table spread and capture consumers who

usually purchase other brands. Premium Golden Spread will enjoy $ 18.4 million per

year if it takes 1 % of the table spread market.

Healthy eating is a rapidly growing industry in the 21st century, with more consumers

becoming aware of the need for a balanced and nutritious diet. Omega-3 fatty acids are

becoming a more integral part of this healthy diet. One specifically, docosahexaenoic

acid (DHA) is linked to the healthy development of infants, and the lack of DHA is

affiliated with Alzheimer’s disease. DHA in the past has come from fish oils, but there is

potential for mercury or dioxins contamination as well as a fishy odor and taste.

4

Recently, it has been found that DHA can be extracted from algae instead of fish oils.

Specific algae strains with high concentrations of DHA can be grown in a controlled

environment and then processed, providing an uncontaminated, highly concentrated, and

completely vegetarian source of DHA. Once extracted, the DHA-rich oils can then be

utilized to create a healthy margarine. About $5.4 billion worth of margarines and table

spreads are sold every year, however a DHA-fortified margarine is not currently sold in

the United States. As word about the benefits of DHA spreads, it is apparent that the

desire for foods fortified with this omega-3 fatty acid will grow. A margarine that is

fortified in DHA will be both profitable in industry and beneficial to a public hungry for a

healthy diet.

DHA is generally regarded as safe (GRAS) by the Food and Drug Administration, so the

product will not need to go through any extra tests or vetting. Sanitation at the facility is

of the highest importance, and the production plant has been carefully designed with

sanitation kept firmly at the forefront of consideration. Each piece of equipment in the

food processing section of the facility is built with polished stainless steel, and each

section of the plant will be cleaned regularly with a clean in place (CIP) system, ensuring

that no microbes will exist in the final product. It is vital that the company enter into the

health food market as soon as possible in order to take advantage of the fractured table

spread market, in which many different companies vie for a small piece of the overall

market, to create and control the niche of healthy table spread. Our product would be the

first of its type easily available, and it is thought that our product would make a large

impression on the table spread market.

5

Conclusions and Recommendations

It is believed that this product will perform extraordinarily well in the marketplace. The

rise of healthy eating awareness has created a huge demand for healthier versions of

common foods, and a healthy table spread would interest a huge percentage of

consumers. The payback period of 5 years is then a conservative estimate. It is

recommended that production of the plant begin as soon as possible in order to ensure

that Golden Premium Spread be the first margarine in the market that can offer the

amount of DHA. The table spread market, though large, is a fractured market. This

means that there are many different companies that each control a very small percentage

of the overall sales. It is relatively easy, then, for a new product, especially one marketed

as having health benefits, to break into the market. In conclusion, a product of this type

would be very successful in the market and would attract a very large consumer base

extremely quickly. It is recommended that the plant be built.

6

Appendix 1

Design Basis

The size of all equipment and the economic evaluations are based on a 1 % market share

in the $ 5.2 billion table spread market. This equates to $ 52 million retail. The product

will sell at the market for $ 5, so selling the tubs wholesale will net $ 1.75. In order to

satisfy the 1 % market share, 18.4 million tubs of margarine will be produced every year.

However, much of the equipment is not running at full capacity, making increased

production if the market share increases easy.

7

Appendix 2

Chemistry and Biology of Omega-3 Fatty Acids

Lipids are molecules used by the body for energy storage, for cell membrane structural

building, and for signaling. Lipids include many different categories, and one kind of

lipid is the fatty acid. Fatty acids are long chains of hydrocarbons which terminate at a

carboxylic acid group. A carboxylic acid is a carbon double bonded to an oxygen

molecule and an alcohol, and it creates a hydrophilic end to the fatty acid molecule. The

other end, the hydrocarbon chain, is hydrophobic, which allows for the lipid to create cell

walls and other structures. The hydrophobic hydrocarbon chain can be saturated or

unsaturated. An unsaturated chain includes one or more double bonds while the saturated

chain has only single bonds. The term saturated refers to the hydrogens attached to the

carbons, and a saturated fatty acid then has the maximum amount of hydrogens bonded to

the carbons. For unsaturated fatty acids, the double bond or bonds occur in either cis or

trans formation, with cis being the primary form found in nature. Trans bonds in the

fatty acid form mostly as a result of processing techniques like partial hydrogenation,

though a very small amount does exist naturally as well. A cis double bond has the

adjacent hydrogen molecules on the same side of the bond and creates a kink in the

hydrocarbon chain. Trans formation, then, is just the opposite, with the adjacent

hydrogens appearing on opposite sides of the bond and maintaining the chain’s linear

shape. It has been found that trans-fatty acids, primarily because these kind of bonds in

fatty acids do not naturally occur, are unhealthy to consume, difficult for the body to

process, and lead to such things as coronary heart disease and obesity. Since the

8

discovery of the health risks of trans fats, the public has become very invested in excising

these fats from all processed food products, leading to the need for new techniques to

replace things like partial hydrogenation.

The body is unable to synthesize double bonds in the fatty acid beyond carbons 9 and 10,

so fatty acids like linoleic acid (LA) and alpha-linoleic acid (ALA) are essential. Though

omega-3 polyunsaturated fatty acids (PUFA) can be synthesized from LA and ALA, it is

a slow and energy intensive process. Therefore, it is much easier and more beneficial to

acquire long chain omega-3 fatty acids, which are used in a variety of bodily functions

and processes, through the diet. Omega-3 fatty acids are named for the double bond that

exists in the third bond from the methyl end of the carbon chain. Some examples of

omega-3 fatty acids are Eicosapentaenoic acid (EPA), Docosapentaenoic acid (DPA), and

Docosahexaenoic acid (DHA). Marine oils in particular contain a large amount of these

omega-3 fatty acids, but fish, like humans, are unable to synthesize these fatty acids

directly. Instead, the fish acquire these fatty acids from consuming algae, and humans

acquire the fatty acids from the fish.

This is the DHA molecule. The omega-3 bond at the methyl end of the carbon chain is depicted. The methyl end is denoted as the omega end of the molecule since it is considered the terminal end. Since there are multiple double bonds, DHA is considered a PUFA, and these double bonds are naturally in the cis formation. In cis formation, the hydrogens bonded to adjacent carbons in double bonds are situated on the same side of the bond. This makes the molecule bend or kink, making it easier for the body to

9

process. The figure also depicts the carboxylic acid that is on the other terminal end of the molecule.

Omega-3 fatty acids have been found to be an extremely beneficial part of the diet, with

many health benefits that come from consuming an adequate amount of these fatty acids.

Some examples of health benefits from consuming omega-3 fatty acids are heart health,

cancer prevention, and brain growth and health. DHA in particular has been singled out

as a very healthy and beneficial omega-3 fatty acid. DHA, like other omega-3 fatty acids,

can be found in various food products, but is found most concentrated in fish oils. These

fish oils are highly concentrated in DHA not because the fish can synthesize omega-3

fatty acids, but because the fish consume these fatty acids from microalgae that do

produce omega-3 fatty acids.

10

Appendix 3

Block Flow Diagram

DegummingBleaching

Removal FFADeodorization

CentrifugeCrude SBO

Citric AcidGumWater

Citric Acid

Citric AcidAcid Activated Clay

EnzymeWater

Acid ActivateClay

Interesterification

Hardfat SBO

Emulsification Pasteurization

WaterSea SaltLecithinDiacetyl

Mono/DiglyceridesB-Carotenes

Sorbic AcidErgocaliferols

Sodium Hydroxide

Citric AcidWater

Sodium HydroxideCitric AcidWater

ScrapedSurface HeatExchanger

Packaging

Refrigerant

To Wholesaler

DissolvedAir

FloatationFermentation CentrifugeCO2

LysingGlucose

Beer StillBottom

Antifoam

Water

Algae

Glucanex

SteamCoolingWater

Sterile air

Vent

Water

Air CO2

LHO-BiomassWater

NaOH

Steam

Steam

Salt

CIPSystem

Block Flow Description

11

Algae is grown in the fermentation stage, and C. Cohnii is the algae type that will be

grown in the fermentation stage. This species of algae has been chosen because of the

high concentration of DHA that exists in its lipids. After the algae is grown, the algae

bodies are separated from the liquid using dissolved air floatation. This process uses air

to push the algae to the top of the broth. After the algae is concentrated at the top of the

broth, it is separated from the bulk fluid by a scraper. This next step of the process is cell

lysing. This step is necessary because the DHA is inside the cell and the cell must be

broken in order to extract the oil. This process uses CO2 to pop open the cells using a

pressure differential. A centrifuge will then separate the oil from the water, leaving the

algae oil that will be used in the margarine process.

The margarine uses refined soybean oil (SBO) that is specially made in a process that

leaves zero trans fatty acids. Crude SBO is bought from Cargill in order to be processed

in our refining plant. The crude SBO is first degummed because the gums contained in

the SBO interfere with the rest of the process. A centrifuge then separates the SBO from

the gums. After the gums are separated, the SBO is bleached and deodorized so that the

final margarine process has a quality taste. Also in this process, free fatty acids are taken

out of the SBO so that the margarine has the correct “mouth feel.” Finally, the last block

of the SBO refining process is interesterification. This is a chemical reaction by which

the glycerides contained in the SBO are randomly reordered in the triglyceride. This

helps the SBO have the right form at the right temperature. If the SBO is too hard, the

margarine would not melt at mouth temperature, creating an unpleasant sensation when

eaten. If the SBO is too soft, then the margarine would liquefy inside the refrigerator,

12

making it very difficult to spread. The interesterification is used because the more classic

method to create margarine that melts and hardens at the right temperatures utilized

partial hydrogenation. This created trans fatty acids inside the margarine, which are

unhealthy and are avoided in this process.

After these ingredients have been prepared, they are mixed together in the margarine

production process. First the ingredients are emulsified, which is just an intense mixing,

creating a creamy mixture of water suspended within the oil. This resulting emulsion is

then pasteurized in order to ensure that no microbes are still living inside the emulsion.

This is required for a food product so that the consumer is not made ill eating the product.

In the scraped surface heat exchanger, the emulsion is crystallized, after which it is sent

to packaging. All of these systems are cleaned with a clean in place (CIP) system, which

keeps all the equipment clean and free of microbes.

Appendix 4

13

Process Flow Diagrams

Fermentation PFD

14

Main Fermenter

Steam Cooling Water

Sparged Steam

Vented Air 13

14

Sterile Air

10% Seed Fermenter

Steam Cooling Water

Sparged Steam

Vent

9

Sterile Air

1000 L Seed Fermenter

10 L Seed Fermenter

250 mL Shake Flasks

Water Water

Direct Steam Injection

MT-1

Salt

Ethanol Bottoms

Glucose

Sterile Glucanex

Sterile Anti-foam

1

2

3

4 5

7

8

11

10

12 Compressor 1

Filter 1 1 Air

Filter 2

TK-1 TK-2 TK-3

TK-4 TK-5

STEX-1

Steam 4a

P- 1 P- 2

P- 3

P- 5 P- 6

P- 7 P- 8

Filter 3 1

To DAF

15

P- 9

Harvest Tank

Rxn tank A

Rxn tank B

P-01

FermentedAlgae

FermentedAlgae Sterilized Air

Sterilize Air

Vented Air

Conc. Algae

Water

45

18

16

15

17

Dissolved Air Flotation PFD

12

Sterilized Air

Sterilized Air18

18

20

19

22 23

21

46Water

1

2

P10

P11P12

Water to Cargill treatment

Tk-6

Tk-7

DAF Tk-7

15

3

Baffle trayTowerTk- 8

CO2 Lysing PFD

Conc. Algae

Sterilized CO2

CO2 AbsorbedAlgae

LysedAlgae&CO2

Vented CO2

LysedAlgae

Throttle Valve

23

28

24

Filter 2

CO 2

Compressor 2

25 26

27

Stir tank

P13

P14

29

Lysed Algae TowerTk- 9

16

Main Centrifuge PFD

Disc Centrifuge

Wet Biomass Slurry

Algae Oil3130

Oil settling tankTK-20

P15

Algae OilCv-32

Lysed Algae29

35

36Water

17

Separation Process PFD

42

Dynamic Cross Flow Filtration

Wet Biomass

P19

40

Wet Biomass TankTk-12

41Wet Biomass

Wet Biomass

39 Water

Cv-37

P21

18

Rotary

Dryer43Dry Biomass

49Steam in

50Steam out

Conveyor Belt

Collection Hopper

Conveyor Belt

Dried Algae DistributorShipping container

37

Purge

42

Dryer Process PFD

Exhauster

Air seal dust valveFume controller

Filter

Recycle

Cv-38

Cv-39

44

45

46

47

48

Air Vapor

Water Vapor

19

Extraction PFD

Oil

Settling Tank

Tk-11

Water

33

Disc Centrifuge

Lysed Algae P16

40

30 34

38

32 Lysed Algae

Biomass

39

P18

P17

Water

45

Water

Degumming PFD

20

TK- 51Crude SBO

Storage

P-51

N2 GasTK-52

45% CitricAcid Storage

P-52

P-53 P-54

TK-534% NaOHStorage

TK-5410% EnzymeRefrigerated

Storage

N2 Gas

HLO - Gums

BatchReactor

P-55 P-56

TK-55SurgeTank

N2 GasTK-56

DegummedSBO

Storage

HX-5

CLR-1LLO

150

HX-6

Cent-5

153

131

132 133 137 142

138

136

135134

139

141140

143

144

145147146

148 149

151152

21

Refining PFD

N2 Gas

TK-56Degummed

SBOStorage

HX-8

TK-5750% Citric

AcidStorage

P-57

P-58

TK-58Acid

ActivatedClay Tank

P-59

P-60

TK-67Soft Water

Storage

P-61HX-9

P-62

VP-1

VacuumMixer

N2 Gas

TK-60SurgeTank

P-63

MX-1MX-1

VP-2

Steam

To MX-1

P-64N2 Gas

TK-61Refined

SBOStorage

HX-10

HX-11

CLR-2

CLR-3

Filter

VacuumVessel

RP-1

153

155 159154

158

162

160

161

156157

164

163

165

166

169168

188187

170

167

188187

184

185

189

192

175

173

174

172

171

180

182183

181

190191

179

178

177

176

22

Interesterification PFD

N2 Gas

TK-61Refined

SBOStorage

N2 Gas

TK-62SBO

HardfatStorage

N2 Gas

TK-63SBO

HardfatHeating

Tank

P-66

P-67

HX-14

HX-15

P-68

TK-64CH3ONaStorage

TK-66MixingTank

P-69

P-70

Filter

TK-67Soft

WaterStorage

HX-16

HLO

P-71

P-72 P-73HLO

Cent-7Cent-6

TK-6520% Citric

AcidStorage

N2 GasTK-68SBO

Storage

CLR-4

MX-3 MX-4

WaterSoftener

P-65

193

195194

196197

198 199 209 210

201200

213211

208207

228

221220 223

222219

216215

218

217

214

225

224

205204

203202

192

227226

212

N2 Gas

206

23

E

Algae OilStorage

Refined SBO Storage

LecithinStorage

Beta CaroteneStorage

DiacetylStorage

SorbicAcidStorage

Mono- diglyceridesStorage

Ergo-califerolStorage

WaterStorage

Oil

Pasteurization

CIP

Warm Water

Aqueous PhasePrep.69

TK-20

TK-21

TK-22

TK-23

CIPCIP

TK-24

TK-25

TK-26

TK-27

TK-30

TK-32

TK-31

Oil Phase Prep.Tank 1

CIP

P-30

P-31

P-32

P-33

Warm Water

Oil Oil Phase Prep.Tank 1

CIP

68

70

67

66

65

64

62 63

60 61

71Sea SaltStorage

TK-29

P-34

73

74

TK-28

75

72

72

76

CIP

79

77

75

79

77

80

Margarine PFD 1

Warm Water

78

P-35

P-35

78

24

High Pressure Pump

AmmoniaCompressor

Pin Rotor MachineScraped

Surface Heat Exchanger

Packaging

P-37

CIP

80

84

8586

CIP

87

89

90

Margarine PFD 2

83

88

Plate HX

8182

P-36

25

TK-41Acidic Solution

TK-42Alkaline Solution

TK-43Reclaim Water

TK-44FreshWater

Alkaline Storage

Acidic Storage

CIP Returning Liquid

Water

HX-41

CIP Supplying Liquid

Drain

P-41

P-42

PP

213

203201

216215 207

205

217

212

204202

214

218208206

211

210

209

Strainer

CIP PFD

Process Flow Description

26

Fermentation

The fermentation of Crypthecodinium cohnii takes place at an ideal temperature of 81°F.

The fermentation process was chosen to be a batch process over a fed-batch process or a

continuous process. The fed-batch process showed no significant benefit to a batch

process, using a glucose substrate, either in final cell density or in fermentation time.

This is likely due to the loss of an unknown growth factor in the ethanol bottoms nutrient

during sterilization. Continuous processes for algae fermentation are untested and can

potentially compromise the sterility of the fermentation. The batch fermentations are to

take place in 35,000 gallon fermentors. Four of these fermentors, operating at 60%

volume capacity, are required to achieve the amount of lipid products specified in our

economic basis.

The fermentation is to take place on a carbon substrate of glucose. Glucose was chosen as

the substrate over acetic acid and ethanol, which could have attained higher cell

concentrations. However, higher concentrations of acetic acid or ethanol are required to

conduct such a fermentation, and both substrates have corrosive qualities that would

cause damage to the fermentors over time. The time period required for acetic acid

fermentations is also longer, at 400 hours, compared to 71 hours per fermentation with

glucose substrate. Ethanol is a fire hazard, especially at the high concentrations required

to ferment C. cohnii.

27

The fermentation of C. cohnii requires a nitrogen source. The nitrogen source for this

fermentation comes from the dead yeast that collects at the bottoms of ethanol stills.

These ethanol bottoms will act as the nitrogen source and the source of any growth

factors that are not included in other feed components. The concentration of ethanol

bottoms used will be limited to 8-10% of the concentration of glucose used as it is

important that nitrogen be consumed by the end of the growth phase of our C. cohnii.

The lack of a nitrogen source will cause the algae to enter a stationary phase in which

growth no longer occurs, but the algae begin to produce excess lipids.

The growth phase of our algae fermentation was calculated using the Arrhenius equation,

, and was calculated to be 70.15 hours. The growth rate used was calculated

from data taken from de Swaaf, et al, 1998 and was found to be 0.0371 h-1. The

maximum final concentration of algae was found to be 27.7 g dw/l and was assumed to

be 27.0 for the purposes of our fermentor. The stationary phase was assumed from

literature to be optimal at 20 hours. The lipid concentration resulting from the combined

90.15 hour fermentation time was taken to be 5.54 g/l. The fermentation schedule

accounts for a 12 hour maintenance period after each batch, which can be reduced to

increase productivity.

At 60% capacity, a single fermentor will hold 79,500 liters of fluid volume. That volume

will produce 971 pounds of lipids and 4,855 pounds of algae biomass per batch. Each

fermentation will require 7.02 short tons of glucose, 0.65 short tons of ethanol bottoms,

and 2.19 tons of sea salt to achieve a salinity of 25 g/l. The glucose, ethanol bottoms, and

28

sea salt will be mixed with 60,617 liters of water, in a 27,000 liter tank at 140°F prior to

their transfer to the fermentor. Sterilization takes place in the fermentor using sparged

steam until the broth reaches 250°F and is held at that temperature for 1 hour. The

volume of sparged steam required is 15,926 pounds of steam at 293 °F and 3 barg. The

broth is then cooled with cooling water until the temperature reaches 81°F. A seed batch

of broth, fermented in a 10% seed fermentor, is then added to the main fermentor.

During the fermentation we will add 350 pounds of glucanex and 31 pounds of silicone

anti-foam per batch as needed.

Glucanex is an enzyme complex consisting of β-glucanase, chitinase, and pectinase

manufactured by Novo Nordisk and it will be used to reduce the viscosity of the

fermentation. It is available as a freeze dried powder and will be mixed into a 100g/l

solution with water. Glucanex is a polysaccharide hydrolase, which will fold the long

chain polysaccharides produced by the algae as it consumes glucose. The byproduct

polysaccharides increase viscosity, which reduces mixing efficiency and reduces aeration

rate of the broth. The glucanex solution will preferably be added at 50% of the

fermentation time to minimize the impact on growth. Thermal sterilization would

deactivate the enzymes in the glucanex solution. The solution will be filter sterilized

using 0.22 micron pore cross flow filtration and the solution will be 100g/l in water.

The anti-foam solution that will be used is an oil with the active ingredient of

polydimethylsiloxane and it is produced by Dow Corning. The oil will be retained in our

final product and its concentration must be minimized to meet FDA standards of less than

29

10 ppm. The anti-foam will be heat sterilized with a steam heat exchanger at 250°F as it

is added to the fermentor.

The fermentation broth must be aerated with filter sterilized air sparged at 157.9

Nm3/minute at 26 psi. The air will be compressed with a centrifugal compressor and will

be sterilized with a 0.22 micron filter. A dissolved oxygen saturation of 20% is desired

for the fermentation, which is 1.359 mg oxygen/ l, for optimal growth.

Degumming

The crude SBO is stored in an agitated tank with a volume of 20,000 gallons. There is a

5 psig nitrogen blanket maintained on the tank. The crude SBO is pumped from the tank

at 720gallons per batch through a heat exchanger to raise the temperature from 77 °F to

176 °F. The heating medium is Paratherm NF being pumped at 500 gph at a temperature

of 617°F and exiting at 528.3°F. 45% citric acid is pumped into the crude SBO stream at

0.07gallon per batch giving a pH of 4.06. Citric acid is stored in a 25 gallon tank. The

citric acid and crude SBO are mixed for 30 minutes prior to the temperature being

dropped to 122 °F by cooling water at a flow rate of 500 gph and an inlet temperature of

85 °F. The cooling water outlet temperature is 113.8 °F. The addition of 4% sodium

hydroxide is a controlled process in order to maintain a constant pH of 4.90 (±0.10) in the

batch reactor. 14.4 gallons of water is added to the batch reactor. The final additive to

the 1200 gallon batch reactor is 10% enzyme solution. The enzyme is Lecitase Ultra,

manufactured by Novozyme. The Lecitase Ultra is stored in a refrigerated storage tank at

30

41 °F. The mixing in the batch reactor continues for 6 hours, and then the solution is

pumped to a 1200 gallon surge tank with a 5 psig nitrogen blanket and a 50 rpm agitation

system. The surge tank is emptied over a 6 hour period with a 120 gallon per hour pump.

Out of the surge tank the solution is heated up from 122 °F to 158 °F by the Paratherm

NF. The Paratherm cools from 617 °F to 611.6 °F. After going through the heat

exchanger the solution is directed to a centrifuge. The centrifuge sends HLO to a storage

tank which will then be sent off to Cargill in order for the gums to be processed. From

the centrifuge, the crude SBO is sent to a cooler to bring the temperature down to 122°F

prior to entering a 4,000 gallon storage tank with a 5 psig nitrogen blanket and a 50 rpm

agitation system.

The degummed SBO is pumped at 120 gph through a heat exchanger to ensure the outlet

temperature is 122 °F. 50% citric acid is stored in a 35 gallon tank and added to the SBO

to achieve a pH of 3.41. The solution goes into a 100 gallon mixer where it is continually

mixed at 150 rpm for 15 minutes. From the outlet of the mixer the solution is pumped at

120 gph to a mixer where a 0.5 wt% oil of acid activated clay is added to the mixer by a

rotary feeder. Water is also added to the second mixer at a 0.05 wt% of the oil. The

solution is mixed for 15 minutes before being pumped through an economizer to raise the

temperature from 122 °F to 176 °F. The solution then enters a vacuum mixer and is

heated up to 250 °F by use of heating coils. It is continually mixed at 250 rpm for 15

minutes. The mixing is maintained at a vacuum of 10 mmHg. The solution exits the

31

mixer and is cooled by the previous economizer to 196° F. At 120 gph, the liquid flows

through a rotary disk filter to remove clay. The clay is discarded to a landfill. As it exits

the filter the liquid goes into a 1,500 gallon surge. The tank is continuously stirred at 50

rpm and utilizes a nitrogen blanket that is kept at 5 psig. The solution is pumped from

the surge tank at 120 gph to a second economizer to raise the temperature to 284° F prior

to it entering a heat exchanger where the temperature is again increased to 500 °F. The

heat exchanger heating medium is Paratherm NF, which cools from 617°F to 568.4°F.

The crude SBO enters the vacuum vessel where it is steam sparged on 4 trays at a

vacuum of 10 mmHg. The oil is maintained on each tray for 15 minutes before it falls

down to the next tray where it is sparged with additional steam. The amount of steam

from top to bottom is 0.1, 0.7, 1.5, 1.5 wt%, respective to the amount of oil. The steam,

FFA, citric acid, and water exit from the top of the vacuum vessel and are sent to Cargill

for processing. The refined oil leaves the vacuum vessel via a 120 gph pump through the

second economizer which lowers its temperature from 482 °F to 415.4 °F. At the outlet

of the economizer the oil goes through the heating coils of the vacuum mixer where its

temperature is decreased to 356 °F. The oil must be cooled down to 122 °F by cooling

water at 85° F entering the cooler and exiting at 108.4 °F. The refined oil enters the

4,000 gallon storage tank with a 5 psig nitrogen blanket and 50 rpm mixing.

Interesterification

32

The refined SBO is pumped from the storage to tank at 350 gph through a heat exchanger

to raise the temperature from 122 °F to 230 °F. The heating medium is Paratherm NF

which has a resulting temperature change of 617 °F to 542 °F. The refined SBO then

enters the interesterification mixing tank.

The hardfat SBO is stored in a 10,000 gallon tank with a nitrogen blanket at 5 psig and

continually stirred at 50 rpm. The hardfat SBO is pumped at 175 gph to a 1,000 gallon

processing tank where the hardfat SBO is heated from 77 °F to 122 °F while being mixed

at 50 rpm with a 5 psig nitrogen blanket on the tank. The tank is heated with Paratherm

NF entering at 500 gph at a temperature of 617 °F to maintain the temperature of the

hardfat SBO. The SBO is then pumped at 100 gph through a heat exchanger to raise the

temperature to 230 °F. The hardfat SBO then enters the interesterification mixing tank.

The hardfat and refined SBO are mixed and maintained at 230° F at 37.5 mmHg for 40

minutes. Cooling water is used to cool the tank to 194 °F, while still maintaining

vacuum. 99.8% sodium methoxide is added to the tank at 0.5 wt% of the oil in the

interesterification tank. Mixing is continued at 500 rpm for 30 minutes. Cooling water

cools the SBO down to 176 °F. The process is inactivated by 2 wt% addition of 20%

citric acid to the tank. Stirring is continued for an additional 15 minutes. When the

process is complete, the SBO is pumped at 500 gph through a rotary disk filter to remove

the sodium methoxide, which is sent to the landfill.

33

Water washing is done with soft water from a 7,500 gallon storage tank. The soft water

is used for two washing procedures. Each wash with soft water is done with 5 wt% oil in

a 500 gallon mixer, stirring at 150rpm. The mixing lasts for 10 minutes in each mixer.

At the outlet of each mixer is a centrifuge to handle the 500 gph oil and water mixture.

The waste from the centrifuge will need to be neutralized and disposed of. The washed

SBO leaves the second centrifuge to enter a 500 gallon atmospheric pan dryer. The

temperature is then increased to 230 °F for 30 minutes. The SBO is then pumped at 500

gph from the dryer to cooler to bring the temperature to 122 °F, using cooling water as

the medium utilized for the cooling. The SBO then enters a 5000 gallon storage tank

with a 5 psig nitrogen blanket and is mixed at 50 rpm.

Margarine Process

The margarine process is fairly straightforward, with no major chemical reactions or

exotic pieces of equipment. The ingredients are placed into two categories: the oil phase

and the water phase. The oil phase consists of the oils and the oil-soluble ingredients.

These are 669 kg / hr of the refined soybean oil gathered from the soybean refining

process, 22.9511 kg / hr of the DHA-rich algae oil taken from the harvested algae, 1.8

kg / hr of lecithin, 4.3 kg / hr of the mono and diglycerides, .00007 kg / hr of

ergocaliferol, .02 kg / hr of the beta-carotene, .002 kg / hr of the diacetyl, and .9 kg / hr

sorbic acid. These ingredients are prepared in an 1500 L oil phase preparation tank.

There are two of these tanks in order for the process to fun semi-continuously because

two tanks can be prepared at the same time. The water phase phase consists of the

34

potable, clean water and the sea salt. The water runs into the water preparation tanks at

154 kg / hr and the salt is added to the preparation at 12 kg / hr. These are prepared

together in a water phase preparation tank. There is only one of these tanks because the

ingredients are quickly and easily mixed together. After one batch is sent through,

another batch is quickly prepared. The water tank for this process is small, at 1000 L

because it is connected to the city water supply and is refilled as the water is used.

After these initial preparations, the oil phase is sent to one of the two 2000 L

emulsification tanks where it is heated with a warm water jacket over the tank. As it is

being mixed, the water phase is then slowly added, creating a water in oil emulsion. This

emulsion is heated to around 100 °F to allow for better mixing, and the emulsion tank

agitators stir the mixture with about 8 hp. After the emulsion is well-mixed, it is sent

forward to a plate heat exchanger which heats the emulsion to 110 °F. Pasteurization is

needed for two major reasons. First, it kills any remaining microbes which may still be

living inside the emulsion. Obviously, for sanitary reasons, these microbes must be

killed. Secondly, the pasteurization stage heats the emulsion to a consistent temperature,

which may not have occurred in the emulsification tanks. After pasteurization, the

emulsion is transferred to a high pressure, plunger type pump or homogenizer. This

pump pressurizes the emulsion to 75 bar, causing a slight temperature increase of 10 °F.

This high pressure keeps the emulsion from separating further down the line. After this

pressurization, the emulsion travels through a long section of insulated pipe. This part of

the process is used for pasteurization, keeping the emulsion at around 120 °F for ten

35

minutes to ensure that any microbes are completely killed. The length of this tube is 330

ft and the pipe will be wrapped around the entire plant as is common practice.

The next stage of the margarine process is considered the heart of the production line.

Here, using a scraped heat exchanger, the emulsion is crystallized to 50 °F using an

ammonia based refrigeration system. An external ammonia compressor is utilized in this

refrigeration system, with an ammonia circulation of 18 tons. The crystallized margarine

is scraped from the walls of the SSHE by mechanical means. After crystallizing, the

margarine is then kneaded in a pin rotor machine which ensures that the final margarine

project has the correct viscosity. Then the margarine is sent through a tub filling and

packaging machine, ready to be sent out the door.

CIP Process

Cleaning and sanitizing equipment is critical in the food processing industry. The

cleaning in place (CIP) system is an integral part of all modern margarine production

facilities, preventing bacteria growth and assuring sanitary equipment conditions. The

CIP system involves the recirculation of the cleaning fluids through assembled food

processing equipment in a continuous circuit with rinse and detergent solutions at the

proper concentrations, temperatures, pressures and times. For a traditional margarine

products the normal equipment cleaning interval is once a week. However, the cleaning

interval is shorter for the products with a high water content, a low fat content, or a high

protein content.

36

The CIP tanks can vary in size depending on the design and the necessary duty of the

system, but they should hold enough liquid to completely fill the largest cleaning circuit

plus a 25% reserve. The usual lay out for CIP set is to have three tanks, one that contains

fresh water, one that contains detergent, and one that contains reclaimed water. A fourth

tank with secondary detergents is found in some systems. The four CIP tank set is usually

used for margarine facilities. The tanks are usually round with a conical bottom, and

made with stainless steel.

CIP pumps fall into roughly three groups: delivery pumps, mixing pumps and return

pumps. Delivery pumps are usually centrifugal, offering high flow at relatively low

pressures and are designed to work best with a flooded suction. These are used to deliver

the cleaning solution to the portion of the plant that is being cleaned. Mixing pumps are

usually of the same design as delivery pumps, but have smaller capacities. These are used

to ensure adequate mixing of the detergents. Return pumps are used to return the cleaning

solution from the portions of the plant that are being cleaned back to the CIP tanks. They

should be designated to pump well even without a flooded suction; this allows them to

cope with the large quantities of air that become entrained in the cleaning solutions

during CIP.

Generally the most effective CIP process incorporates five steps:

1. An initial rinse;

Rinse with water at ambient temperature to remove any residues. 10 to 15 minutes

are usually sufficient for this part of the cycle.

37

2. Rinse with alkaline detergent;

Rinse with an alkaline detergent, typically a 2-2.5% solution of Caustic Soda

(NaOH) at about 70˚C for a time period of 20-30 minutes. This phase should

remove any organic matter.

3. Intermediate rinse with water;

Rinse with water at ambient temperature for a time period of 5 to 10 minutes. This

step should remove any residual detergents

4. Rinse with acidic detergent;

Rinse it with an acidic detergent, typically a 2.5% or lower concentration of Nitric

Acid (HNO₃) solution at ambient temperature for a time period of 10-15 minutes.

This phase should remove mineral salts, proteins, lime and other deposits.

5. Final rinse with clean water.

Rinse with clean water at ambient temperature to remove all traces of the cleaning

detergents. 10 to 15 minutes are usually sufficient for this part of the cycle.

During the CIP cycle it is important to keep the detergent concentrations constant; a

significant increase in concentrations can damage the system equipment, while a

significant decrease in concentration can negatively affect the detergents cleaning

efficiency.

38

Appendix 5

Stream Data

Stream No

Stream Name

Stream DataFlow rate, kg/batchFlow rate, kg/hrTemperature, °CPressure, bargPhaseSp grBulk density, kg/m3

Water, kgDry solids, kgSolvent, kgpHAir, Nm3/min

1 2 3 4 4a 5

Glucose Sea SaltEthanol Bottoms

Water to Water Heater

Steam to Water Heater

Water to Mix Tank #1

6,371.2

1,989.1

592.1

56,827.4

3,790.0

60,617.3

17 17 17 17 145 60

1 1 2 3 2Liquid Solid Liquid Liquid Gas Liquid

1.54 1.00 0.002 1.00

7 8 9 10 11 12

39

Medium to Fermenter

Steam Sparged Seed Batch

Sterile Anti-foam

Sterile Glucanex Sterile Air

69,569.7

7,224.3

8,277.2 14

1,590

14,168

7,222

7,224.3

0.20

202.01

60 145 121 27 -

- 3 2.0 2.0 0.8 Liquid Gas Liquid Liquid Liquid Gas 1.20

1.20 -

- - 60,617

7,224.3

7,305.7

1,431 -

592

215.6

159 -

8,360

755.9 -

7.0 7.0 - - 156.32

13 14 15 16 17 18

Vented AirInlet from

Harvest Tank Rxn tank A Rxn tank BSterile Air In Sterile Air In

14,168

86,663

43,332

43,332

202.01

14,444

7,222

7,222 1,000 1,000

27 27 0.5 7 7 Gas Liquid Liquid Liquid Gas Gas

- 1.20

1.60

-

- 76,583

38,292

38,292

- 3,095

1,548

1,548

- 6,985

3,492

3,492

-

156.32 13 13

19 20 21 22 23 24

40

inlet DAF

Waste Water

Conc Algae

Vented air

Inlet from daf tk Sterlized CO2 In

86,663 63,295

23,368 23,368

14,444

30,162 10,882 1,000

27 27

27

27 27

3 10.3422 Liquid Liquid Gas Liquid Gas 1 2

1,600 1,600

76,583 57,437

19,146 19,146

3,095 619

2,476 - 2,476

6,985 5,239

1,746 - 1,746

26

25 26 27 28 30 31Absorbed

CO2 Algae

Throttled Algae

Vented CO2

Lysed Algae Ctfg. Inlet lipids

23,368 23,368 466

6,581 27 3 1.01353 0 Liquid Liquid gas Liquid Liquid

18,787 18,787

19,146 19,146 -

624 2319 2,476 2,476 466

2,319 2,319 1,746 1,746 -

32 33 34 35 36 37

41

biomass

Biomass + Water In Biomass Water Out Dry Biomass Evaporated Water

22,903

22,902.52

6,168.82

16,733.71 2,339.66 3,829.16

31,360 Liquid Liquid Liquid Solid Liquid 19,146

19,146

3,829

15,317

2,011 2,011

1,990.42 20.11

1,746 1,746

349 1,397

60 61 62 63 64 65

Alg. Oil In 1 Alg. Oil In 2 SBO In 1 SBO In 2 Lecithin In

B-Carotene In

22.9511 22.9511 668.831 668.831 1.73047 0.01716870 70 70 70 70 70

Liquid Liquid Semi-Solid Semi-Solid Liquid Solid

0.9 900 900 900 900 1078.2 941

- - - - - -- - - - - -- - - - - -- - - - - -- - - - - -

42

66 67 68 69Diaceyl In Sorbic Acid In Mono- Di Glycerides In Ergocaliferol

0.002163 0.865236 4.32618 7.52E-05

70 70 70 70 Liquid Liquid Solid

990 1200 - - - -- - - -- - - -- - - -- - - -

70 71 72Potable Water in 1 Potable Water In 2 Oil Phase prep. 1

153.803 153.803 698.72370 70 70

liquid Liquid Liquid

1000 1000 933153.803 153.803 -

- - -- - -- - -- - -

73 74 75 76

43

Sea Salt In Aqueous Phase 1 Aqueous Phase 2 Oil Phase 2

12.3042 166.1072 166.1072 698.72370 70 70 70

Solid Liquid Liquid Semi-Solid

920 980 980 933- 153.803 153.803 -- - - -- - - -- - - -- - - -

77 78 79Heating Water In Heating Water In 2 Heating Water Out

22712.5 22712.5 22712.5120 120 119

Liquid Liquid Liquid

1000 1000 100022712.5 22712.5 22712.5

- - - -

80 81 82 83 84Emulsion Out Plate Hot Plate Hot Water PHX Out HPP Out

44

Water In Out

864.8305922 22712.5 22712.5 864.8305922 864.8305922100 120 119 110 120

75Liquid Liquid Liquid Liquid

933 1000 1000 933 933153.803 22712.5 22712.5 153.803 153.803

- - -- - -

5.3 5.3 5.3- - -

85 86 87 88Ref. In Ref. Out SSHE Out SSHE Out 2 - - 864.8305922 864.8305922

-20 50 50 Liquid Vapor Semi-Solid Semi-Solid

20 - 933 933- 153.803 153.803- - -- - -- - -- - -

89 90Pin Rotor Out To Wholesaler

45

864.8305922 864.830592250 50

Semi-Solid Semi-Solid

933 933153.803 153.8

- -- -- -- -

131 132 133 134 135

Nitrogen Blanket

Crude SBO Inlet HX-5

Crude SBO Outlet HX-5

Heating Medium Inlet

HX-5

Heating Medium Outlet

HX-5 720.00 720.00 500.00 500.00 77 176 617 528.3

0.5 psi Gas Liquid Liquid Liquid Liquid

0.92 0.89 7.00 7.00

136 137 138 139 140From 45% Citric Acid

Tank

Inlet Batch Reactor

From 4% NaOH Tank

From Enzyme Tank

Cooling Water Inlet

0.07 720.00 0.11 0.50 500.00

77 176 77 77 85

Liquid Liquid Liquid Liquid Liquid1.24 0.92 1.00 1.00

1.75 4.06 12.60 7.00

141 142 143 144 145Cooling Water

OutletBatch Reactor

OutletNitrogen Blanket

Surge Tank Outlet

Centrifuge Inlet 5

720.00 500.00 120.00 120.00113.8 122 122 158

0.5 psi

46

Liquid Liquid Gas Liquid Liquid 0.92 0.92 0.92 4.90 4.90 4.90

146 147 148 149 150Heating

Medium Inlet HX-6


HX-6HLO - Gums

LLO - Degummed

SBO

Inlet Degummed

SBO Storage

500.00 500.00 3.63 120.00 120.00617 611.6 158 158 122

Liquid Liquid Liquid Liquid Liquid

0.92 4.90 7.00 7.00

151 152 153Cooling Water

Inlet CLR-1Cooling Water Outlet CLR-1

Nitrogen Blanket

500.00 500.00

85 88 0.5 psi

Liquid Liquid Gas

153 154 155 156 157

Nitrogen Blanket

Degummed SBO Storage Tank Outlet

Outlet HX-8


HX-8 Paratherm


HX-8 Paratherm

120.00 120.00 500.00 500.00 122(77 S/U) 122 617 610.4


47

0.92 0.92 7.00 7.00

158 160 161 162 163From 50% Citric Acid

Tank

Acid Activated Clay Inlet MX-2

Water Inlet MX-2 Outlet MX-2 Inlet Vacuum

Mixer

0.07 4.61 lb/hr 0.11 120.00 120.0077 77 122 176

Liquid Solid Liquid Liquid Liquid1.24

1.72 3.41 3.41

164 165 166 167 168

Vacuum Pump 1

Outlet Vacuum Mixer Inlet Filter Waste Outlet

FilterInlet Surge

Tank

120.00 120.00 0.35 120.00 249.8 195.8 195.8 195.8

10.00 Gas Liquid Liquid Solid Liquid

3.41 3.41 3.41 3.41

169 170 171 172 173

Nitrogen Blanket

Outlet Surge Tank Inlet HX-11 Outlet HX-11


HX-11 120.00 120.00 120.00 500.00 195.8 284 500 617


48

3.41 3.41 3.41

174 175 176 177 178Heating

Medium Outlet HX-11

Vacuum Pump 2

Tray 1 Sparging Steam



500.00 0.13 0.93 1.99568.4 224.6 224.6 224.6

6.00 Liquid Gas Gas Gas Gas

179 180 181 182 183Tray 4

Sparging Steam

Waste Outlet Vacuum Vessel

Outlet CLR-2 Cooling Water Inlet CLR-2

Cooling Water Outlet CLR-2

1.99 6.46 6.46 500.00 500.00

224.6 374 140 85 94.5

Gas Gas Liquid Liquid Liquid 1.72 1.72

184 185 187 188 189

Recycle to MX-1

Outlet Vacuum Vessel Outlet HX-10 Outlet Vacuum

Mixer Heater

Inlet Refined SBO Storage

Tank 120.00 120.00 120.00 120.00 482 415.4 356 122


7.00 7.00 7.00 7.00

49

190 191 192

Cooling Water Inlet CLR-3

Cooling Water Outlet CLR-3

Nitrogen Blanket

500.00 500.00

85 108.40 0.5 psi

Liquid Liquid Gas

193 194 195 196 197

Nitrogen Blanket

SBO Hardfat to Heating Tank

Nitrogen Blanket

Heating Medium Inlet Heating Tank

Heating Medium Outlet Heating Tank

175.00 500.00 500.00 77 617 551.7

0.5 psi 0.5 psi Gas Liquid Gas Liquid Liquid

7.00

198 199 200 201 202

Inlet HX-14 Outlet HX-14Heating

Medium Inlet HX-14


HX-14Inlet HX-15

100.00 100.00 350.00 500.00 500.00

122 230 617 595 122


7.00 7.00 7.00

203 204 205 206 207

50

Outlet HX-15Heating

Medium Inlet HX-15


HX-15

Nitrogen Blanket

Cooling Medium Inlet Mixing Tank

350.00 500.00 500.00 500.00

230 617 542.3 85 0.5 psi

Liquid Liquid Liquid Gas Liquid

7.00

208 209 210 211 212Cooling

Medium Outlet Mixing Tank

Outlet CH3ONa Storage Tank

Outlet 20% Citric Acid

Storage Tank Inlet Filter Outlet Filter

1.00 10.00 500.00 500.00500.00

97.6 77 77 176 176

Liquid Solid Liquid Liquid Liquid 1.93 4.50 4.50

213 214 215 216 217

Waste Outlet Filter

Outlet Soft Water Storage

TankInlet MX-3 Inlet MX-4


HX-161.63 100.00 50.00 50.00

500.00176 77 190 190 617

Solid Liquid Liquid Liquid Liquid

4.50 7.00 7.00 7.00

218 219 220 221 222

51


HX-16

Inlet Centrifuge 6

HLO Centrifuge 6

Soaps

LLO Centrifuge 6

Inlet Centrifuge 7

500.00 32.79 500.00 500.00500.00 573.9 178.5 178.5 178.5 180.5


4.50 4.50 7.00 7.00

223 224 225 226 227HLO

Centrifuge 7 Soaps

LLO Centrifuge 7

Inlet SBO Storage Tank

Cooling Medium Inlet

HX-13

Cooling Medium Outlet

HX-1332.36 500.00 500.00

500.00 500.00180.5 180.5 122 85 123.5


7.00

228Nitrogen

Blanket SBO Storage Tank

0.5 psiGas

52

Appendix 6

Material and Energy Balance

Refining Mass Balance on Yearly Basis

Centrifuge 1Inlet HLO LLO

Crude SBO 2856361.3 kg Crude SBO 17401.3 kg Crude SBO 2838960.0 kg

53

FFA 43503.3 kg Phospholipids 323.3 kg Phospholipids 29.0 kgPhospholipids 352.4 kg Citric Acid 380.5 kg FFA 43503.3 kgCitric Acid 380.5 kg Water 12305.0 kgWater 12305.0 kg NaOH 486.0 kgNaOH 486.0 kg Enzyme 60.0kgEnzyme 60.0 kg

2913448.5 – 30956.2 – 2882492.3 = 0Filter

Inlet Waste OutletCrude SBO 2838960.0 kg Crude SBO 8516.9 kg Crude SBO 2830443.2 kgPhospholipids 29.0 kg Phospholipids 20.4 kg Phospholipids 8.7 kgFFA 43503.3 kg Clay 14412.3 kg FFA 43503.3 kgCitric Acid 2331.8 kg Citric Acid 2331.8 kgClay 14412.3 kg Water 4762.7 kgWater 4762.7 kg

2903999.3 – 22949.6 – 2881049.7 = 0Sparging

Inlet Waste OutletCrude SBO 2830443.2 kg Crude SBO 28304.4 kg Crude SBO 2787986.5 kgPhospholipids 8.7 kg Phospholipids 5.8 kg Phospholipids 2.9 kgFFA 43503.3 kg FFA 42633.3 kg FFA 870.0 kgCitric Acid 2331.8 kg Citric Acid 2331.8 kg Mono/

Diglycerides14152.3 kg

Water 4762.7 kg Water 4762.7 kgSteam 109533.3 kg Steam 109533.3 kg

2990582.0 – 187571.3 – 2803011.7 = 0Interesterification

Inlet OutletCrude SBO 2787986.5 kg SBO 3488739.4 kgPhospholipids 2.9 kg Phospholipids 2.9 kgFFA 870.0 kg FFA 870.0 kgMono/Diglycerides

14152.2 kg Mono/Diglycerides

14152.2 kg

SBO Hardfat 700752.9 kg Citric Acid 18688.7 kgCitric Acid 18688.7 kg CH3ONa 6863.4 kgCH3ONa 6863.4 kg Soft Water 60359.0 kgSoft Water 60359.0 kg

3589675.6 - 3589675.6 = 0Filter

Inlet Waste OutletSBO 3488739.4 kg SBO 13955.0 kg SBO 3474784.5 kgPhospholipids 2.9 kg CH3ONa 5998.7 kg Phospholipids 2.9 kgFFA 870.0 kg FFA 870.0 kgMono/Diglycerides

14152.2 kg Mono/Diglycerides

14152.2 kg

Citric Acid 18688.7 kg Citric Acid 18688.7 kgCH3ONa 6863.4 kg CH3ONa 864.7 kg

54

Soft Water 60359.0 kg Soft Water 60359.0 kg3589675.7 – 19953.7 – 3569722.0 = 0

Centrifuge 2Inlet HLO LLO

SBO 3474784.5 kg SBO 20848.7 kg SBO 3453935.8 kgPhospholipids 2.9 kg FFA 435.0 kg Phospholipids 2.9 kgFFA 870.0 kg Citric Acid 18688.7 kg FFA 435.0 kgMono/Diglycerides

14152.2 kg CH3ONa 432.4 kg Mono/Diglycerides

14152.2 kg

Citric Acid 18688.7 kg Soft Water 199413.0 kg CH3ONa 432.4 kgCH3ONa 864.7 kg Soft Water 34685.3 kgSoft Water 234098.2 kg

3743461.4 – 239817.8 – 3503643.6 = 0Centrifuge 3

Inlet HLO LLOSBO 3453935.8 kg SBO 20723.6 kg SBO 3433212.1 kgPhospholipids 2.9 kg FFA 435.0 kg Phospholipids 2.9 kgFFA 435.0 kg CH3ONa 432.4 kg Mono/

Diglycerides14152.2 kg

Mono/Diglycerides

14152.2 kg Soft Water 173287.4 kg Soft Water 34473.7 kg

CH3ONa 432.4 kgSoft Water 208111.6 kg

3677069.9 – 195228.4 – 3481840.9 = 0

(3447367.2 kg) * 99.3% = 3424720 kg

Appendix 7

Calculations

For the fermentation process, an algae Monad growth rate was assumed.

55

Fermentation Time Requirement (hours/batch)= Lag Phase + Growth Phase + Stationary Phase + Maintenance= 0 + [ln(27.7 g/L)-ln(2 g/L)] / 0.03747 hr-1 + 20 + 12= 102.14 hours

Algae Production (tons/batch)= Final Concentration (g/L) x Broth Volume (L) / (1000 g/kg) x

(2.2046lb/kg) / (2000lb/ton)= 27.7 x 79,500 / 1000 * 2.2045 / 2000= 2.427 tons/batch

Annual Amount of Algae Produced (tons)= Oil needed (tons) / 0.2 (g lipids / g dw cells)= 133 / 0.2= 665.0 tons

Annual Amount of Batches Required= 665.0 tons / (2.427 tons/batch)= 274.0 batches

Plant Operating Time (hours/year)= Time per batch (hours) * Batches per year / Number of fermentors = 102.14 hours * 274.0 / 4= 6,996.6 ~ 7,000 hours/year

Steam Sparged into Fermentor to Sterilize, 60 to 121°C (kg steam)

56

(Steam at 145°C and 3 barg)mbroth (kg)*cp(kJ/kg*K)*ΔT (K) = msteam(kg)*Hvap (kJ/kg) + msteam (kg)*cp

(kJ/kg*K)*ΔT (K)17,300,000 kJ = msteam *[2,163.47 (kJ/kg) + 0.5*4.187*(144-

120)+0.5*4.187*(144-60)] msteam = 7,240 kg steam

Steam Used to Heat Mix Water from 17 to 60°C (kg steam)(Steam at 145°C and 3 barg)

mbroth (kg)*cp(kJ/kg*K)*ΔT (K) = msteam(kg)*Hvap (kJ/kg) + msteam (kg)*cp (kJ/kg*K)*ΔT (K)

9,873,628 kJ = msteam *[2,163.47 (kJ/kg) + 0.5*4.187*(144-17)+0.5*4.187*(144-60)]

msteam = 3,719.7 kg steamHeat Evolved from Cell Growth (kcal)

= 0.12 * Oxygen Uptake Rate (mmol/(h)) * Fermentation Time (h)= 0.12 * 295.35 mmol/h * 70.14 h= 2,485.88 kcal

Co 2 absorption and LysingMass Balance:ΣMass = Mass in – Mass out = 0 0 = 86,663 (kg/batch of algae) + 1000 (kg/hr of air) – 63,295 (kg/batch waste water) -23,368 (kg/batch of conc. algae) - 1000 (kg/hr of air) Energy Balance:Q = MCp(∆T) + Qin Q = throttling process is adiabatic, so Qin is the steam jacket to tankQ = 12,700.6 kg/hr of steamCentrifuge #1Mass Balance:ΣMass = Mass in – Mass out = 00 = 23,368 (kg/batch of conc. Algae) - 466 (kg/batch of lipids) - 22, 902 (kg/batch of lysed algae)Energy Balance:Q = MCp(∆T) + QinQ = assumption of no temperature change , Qin HP supplied to centrifugeQ = 150 HP x 2545 = 381, 845 BTU/ Hr

Centrifuge #2Mass Balance:ΣMass = Mass in – Mass out = 00 = 22,902.52 (kg/batch of lysed algae) – 6,168.82 (kg/batch of algae) – 16,733.71 9 (kg/batch of waste water)Energy Balance:Q = MCp(∆T) + QinQ = assumption of no temperature change , Qin HP supplied to centrifugeQ = 100HP x 2545 = 244,500 BTU/ Hr

57

Centrifuge #3Mass Balance:ΣMass = Mass in – Mass out = 0Energy Balance:Q = MCp(∆T) + QinQ = assumption of no temperature change , Qin HP supplied to centrifugeQ = HP x 2545 = 381, 845 BTU/ Hr

Vacuum VesselSparging Steam flow designTray1- 715.59/6/3785.412*0.092/1000*0.001 = 0.42 gal/hrTray2- 715.59/6/3785.412*0.092/1000*0.007 = 2.91 gal/hrTray3- 715.59/6/3785.412*0.092/1000*0.015 = 6.23 gal/hrTray4- 715.59/6/3785.412*0.092/1000*0.015 = 6.23 gal/hrHX-11 Oil into the Vacuum VesselQoil = mcp(∆T)Qoil = 921.33*0.55*(500-284)Qoil = 109454 BTU/hrQPoil = mcp(∆T)T = 617- (109454/2900/.79)T = 569.2°F∆Tlm = [(T1 – t2) – (T2 – t1)]/ln[(T1 – t2)/ (T2 – t1)]∆Tlm = [(617-500) – (569.2-284)]/ln[(617-500)/(569.2-284)]∆Tlm = 188.8°FQ = UA ∆Tlm A = 109454/60/188.8A = 9.68 ft2 Annual Energy Cost For 1 Fermentor Motor Power (kWh) * Cost of energy ($0.10/kWh) * # of BatchesAnnual Energy Cost = 400 hp * 0.745 kW/hp * 90.2 hours * $0.10/kWh * 272 batchesAnnual Energy Cost = $ 731,125.12

Interesterifaction Rate ConstantAssumed that the rate constant depended on a Arrhenius relationship, assumed activation energy is similar to digestion, breaking triglycerides in the bodyk=A*e^(Ea/RT)T= 353 KEa = 9 kcal/mol = 37,681.2 J/molR = 8.312 J/mol KA = 1.948 * 10^9 1/mink = 5.171 * 10^3 1/min

Heat needed for temperature change of emulsion

58

Q=w Cp ΔTAssume clean heat transfer area and an emulsion specific heat of .5(1906.2) * (.5) * (100-70)= 28,599.3 BTU / hr

Heat needed for Plate Heat ExchangerQ = w Cp ΔT= (1906.62) (.5) (110-100)= 9533.1 BTU/hr

Heat Removed by SSHEQ = w Cp ΔT= (1906.62) (.5) (50-120)= -66,731.7 BTU/ hr

Length of Pasteurization Piping needed Volumetric Flow rate/(Area of pipe) = velocity of fluid.0155 m3/min/(π * .022252) = 9.966 m/min9.966 m/min * 10 min = 99.66 m needed for pasteurization time99.66 m (3.28 ft / 1 m) = 326.885 ft

Appendix 8

Annotated Equipment List:

Equipment Size (gal)TK-01 11,000TK-02 40,000TK-03 1,200TK-04 300TK-05 300

59

MT-1 22,00010L Seed Fermenter 2.651000L Seed Fermenter 26510% Seed Fermenter 3,500Main Fermenter 35,000CO2 Lysing Tank 12,000DAF Tank 24,000

Component Sizing Component Sizing

TK-51 20000 galTK-52 25 galTK-53 400 galTK-54 15 galBatch Reactor 1500 galTK-55 1500 galTK-56 4000 galTK-57 35 galTK-58 250 galTK-59 600 galTK-60 100 galTK-61 4000 galTK-62 10000 galTK-63 1000 galTK-64 250 galTK-65 1200 galTK-66 400 galTK-67 7500 galTK-68 5000 gal

MX-1 100 galMX-2 100 galMX-3 500 galMX-4 500 gal

Component Sizing Component Sizing

HX-5 202490 BTU/hrHX-6 12604 BTU/hrCLR-1 12604 BTU/hrHX-8 15340 BTU/hrHX-9 19901 BTU/hrHX-10 36568 BTU/hrHX-11 109454 BTU/hr

60

CLR-2 39542 BTU/hrCLR-3 97016 BTU/hrHX-14 49572 BTU/hrHX-15 159621 BTU/hrHX-16 92228 BTU/hr

Component SizeP-30 0.25 GPMP-31 3.5 GPMP-32 1 GPMP-33 1 GPMP-34 3.5 GPMP-35 110 GPMP-36 4.25 GPMEmulsion Tank 1 2000 LEmulsion Tank 2 2000 LHigh Pressure Pump 2475 L / hrPlate Heat Exchanger 6 ft^2Scraped Surface Heat Exchanger 2150 L / hrPin Rotor Machine 2000 L/hrTub Filling Machine 2740 L/hrAmmonia Compressor 18 ton

Component SizeTK-20 1000 LTK-21 5000 LTK-22 20 LTK-23 2 LTK-24 2 LTK-25 5 LTK-26 10 LTK-27 2 LTK-28 500 LTK-29 1000 LTK-30 500 LTK-31 1500 LTK-32 1500 L

61

Appendix 9

Economic Evaluation Factored from Equipment Costs

All data was obtained through the use of Aspen Economic Evaluator. For this evaluation, the design basis of 1 % of the $ 5.2 billion dollar table spread market was assumed.

Total Project Capital Cost 37,328,232.77 USD

Operating Costs 18,962,032.85 USD/Year Raw Materials 6,031,280.00 USD/Year Utilities 3,532,157.83 USD/Year Maintenance 2,030,000.00 USD/Year

62

Operating Labor Costs 2,828,000.00 USD/Year Operating Charges 707,000.00 USD/Year Plant Overhead 2,429,000.00 USD/Year G and A Costs 1,404,594.96 USD/Year

Total Product Sales 25,963,238.43 USD/Year

Payback Period 5.7 Years

Tax Rate is 40% Straight Line Depreciation over 10 years 2,986,258.62 USD/Year

YearCumulative Cashflow 1%(1+.05)^15

NPV

IRR

1 2 3 4 5 6

-6,743,784.40-

37,526,542.44-

36,965,246.61-

26,892,098.71-

15,922,960.38-

4,000,701.501.05 1.10 1.16 1.22 1.28 1.34

8594606.905 8896573.13984,727,200.17

1.255 1.575 1.977 2.481 3.113 3.9073523334.564 3051379.281

25.5% 37,830.75

7 8 9 10 11 12

8,935,054.09 22,948,103.27 38,105,847.45 54,479,485.96 72,144,219.55 91,179,464.491.41 1.48 1.55 1.63 1.71 1.80

9193199.989 9484583.267 9770817.048 10051993.69 10328203.88 10599536.65

4.903 6.154 7.723 9.693 12.164 15.2662638066.389 2277103.861 1962641.785 1689304.503 1452198.915 1246906.129

13 14 15

111,669,077.84 133,701,594.40 157,370,476.071.89 1.98 2.08

10866079.43 11127918.04 11385136.78

63

19.159 24.044 30.1761069461.925 916330.0812 784371.5829

Cumulative Cash Flow

-$50,000,000

$0

$50,000,000

$100,000,000

$150,000,000

$200,000,000

0 2 4 6 8 10 12 14 16Year

Cum

ulat

ive

Rev

enue

(USD

)

Another evaluation was done in which the market share of Golden Premium Spread would increase to 2 % in 7 years:

Cumulative Cash Flow 1% (2% in year 7)(1+.05)^15

NPV

IRR

-6,743,784.40-

37,526,542.44-

36,965,246.61-

26,892,098.71-

15,922,960.38-

29,650,701.501.05 1.10 1.16 1.22 1.28 1.34

8594606.905 -10243851.78158,394,244.01

64

1.345 1.81 2.43 3.27 4.40 5.922491148.688 2012941.306

34.5% -3,699.31

-3,779,190.32 24,246,908.04 54,562,396.40 87,309,673.42 122,639,140.60 160,709,630.48

1.41 1.48 1.55 1.63 1.71 1.8018386399.98 18969166.53 19541634.1 20103987.38 20656407.75 21199073.29

7.97 10.72 14.41 19.39 26.08 35.083247429.924 2615327.379 2103165.333 1688999.289 1354680.215 1085259.492

201,688,857.18 245,753,890.30 293,091,653.641.89 1.98 2.08

21732158.85 22255836.08 22770273.55

47.19 63.47 85.37868468.9858 694272.6917 554484.1595

Cumulative Cash Flow

-$100,000,000

-$50,000,000

$0

$50,000,000

$100,000,000

$150,000,000

$200,000,000

$250,000,000

$300,000,000

$350,000,000

0 2 4 6 8 10 12 14 16Year

Cum

ulat

ive

Rev

enue

(USD

)

65

Another situation was examined in which Golden Premium Spread would enjoy a 2 % market share at the beginning of its life. The payback period for this situation is about 5 years:

YearCumulative Cashflow 1%(1+.05)^15

NPV

IRR

1 2 3 4 5 6

-6,743,784.40-

43,226,542.44-

42,103,950.78-

21,957,654.98 -19,378.32 23,825,139.441.05 1.10 1.16 1.22 1.28 1.34

17189213.81 17793146.28198,627,853.09

1.428 2.039 2.911 4.157 5.936 8.476-9445737.657 -36810605.43 385592.3691 4846230.113 3695843.23 2813207.6

25.5% -3,665.08

7 8 9 10 11 12

49,696,650.62 77,722,748.98 108,038,237.34 140,785,514.36 176,114,981.54 214,185,471.421.41 1.48 1.55 1.63 1.71 1.80

18386399.98 18969166.53 19541634.1 20103987.38 20656407.75 21199073.29

12.103 17.282 24.676 35.235 50.312 71.8412137653.163 1621736.506 1228526.21 929388.4216 702200.8618 529925.6511

13 14 15

255,164,698.12 299,229,731.24 346,567,494.581.89 1.98 2.08

21732158.85 22255836.08 22770273.55

102.582 146.477 209.154399477.6092 300832.6647 226329.326

66

Cumulative Cash FlowBasis: 2% Market Share from Start

-$100,000,000

-$50,000,000

$0

$50,000,000

$100,000,000

$150,000,000

$200,000,000

$250,000,000

$300,000,000

$350,000,000

$400,000,000

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15Year

Cum

ulati

ve R

even

ue (U

SD)

Appendix 10

Utilities

Chemical and Margarine Ingredient Costs:

Chemical Unit Price Amt. Required Yearly Price TotalCrude SBO $867.40/ton 3,196.9423 tons $2,773,027.75SBO Hardfat $0.46/lb 1,528,431.473 lb $703,078.4850% Citric Acid $8.51/gal 9,118.579 gal $77,599.11Acid Activated Clay $209/ton 15.887 tons $3,320.3899% Sodium Methoxide

$14/kg 6,863.348 kg $96,086.87

Nitrogen $7.7825/m³ 1.3114 m³ $102.064% Sodium $4.39/L 484.737 L $2,128.00

67

HydroxideEnzyme $576/L 60.053 L $34,590.53Sea Salt (Fermentation)

$600/ton 672.44 tons $403,464.00

Ethanol bottoms $140/ton 199.46 tons $27,924.40CO2 Free 1,096,000 kg -Lecithin $0.81/lb 19,534.7 lb $15,823.10Sea Salt (for Margarine Production)

$600/ton 739.449 tons $443669.40

Ergocaliferol $18234/kg 0.36628 lb $3,029.43Mono/Di Glycerides $0.1/kg 48,836.8 lb $10,780.75Beta-Carotene $1250/kg 193.8 lb $109,883.00Diacetyl $2.5/kg 24.42 lb $27.69Sorbic Acid $8250/ton 9,767.36 $40,290.40Dextrose $0.3085/lb 2141 tons $1,320,997.00

Fermentation Facility:

Equipment Energy Requirements:

Fermentation Mixer x 4: 300 kWSteam Boiler: 597 kWDecanter Centrifuge x 2: 106 kWDisk Centrifuge x 2: 15 kW

Refining Process:

Heat Loads and Sink:

Heat Exchanger 5Q= 202,490 BTU/ hr

Heat Exchanger 6Q = 12,604 BTU/ hr

Cooler 1Q = 12,604 BTU / hr

Heat Exchanger 8Q = 15,340 BTU / hr

68



Heat Exchanger 11Q = 109454 BTU / hr

Cooler 2Q = 39542 BTU / hr

Cooler 3Q = 97016 BTU / hr




Cooler 4Q = 161,694 BTU / hr

Batch Reactor Cooling CoilsQ = 119,232 BTU / hr

Vacuum Mixer Heating CoilsQ = 29,317 BTU / hr

TK-63 Heating CoilsQ = 139,588 BTU / hr

TK-66 Cooling CoilsQ = 52,261 BTU / hr

Margarine Plant:

Equipment Electrical Requirements:

Emulsion Mixer: 8 kW

69

Emulsion Mixer 8 kWHigh Pressure Pump (Homogenizer): 15 kWSSHE: 22 kWPin Rotor Machine: 28 kWAmmonia Compressor: 55 kWTub Filling and Packing: 2.75 kW

Heat Loads and Sinks:

Temperature of Margarine rises from 70 ºF to 100 °F in emulsion tanks:Q = 29,000 BTU/hr

Temperature of Margarine rises from 100 °F to 110 °F in plate heat exchanger:Q = 9,600 BTU/hr

Temperature of Margarine decreases from 120 °F to 50 °F in scraped surface heat exchanger:Q = 58,000 BTU/hr

Appendix 11

Conceptual Control Scheme

70

Main Fermenter

Cooling Water

SpargedSteam

Vented Air 13

14

Sterile Air

10% Seed Fermenter

Cooling Water

SpargedSteam

Vent

9

Sterile Air

1000 L Seed Fermenter

10 L Seed Fermenter

250 mLShake Flasks

Water Water

Direct Steam Injection

MT-1

Salt

Ethanol Bottoms

Glucose

Sterile Glucanex

Sterile Anti-foam

1

2

3

4 5

7

8

11

10

12Compressor 1

Filter 11Air

Filter 2

TK-1 TK-2 TK-3

TK-4 TK-5

STEX-1

Steam 4a

P- 1

P- 2

P- 3

P- 5 P- 6

P- 7 P- 8

Filter 3

To DAF

15

P- 9

Harvest Tank

PIC

QIC

QIC FIC

FIC

TIC

TIC

TIC

TICCompressor 1Air

PIC

Filter 21

PIC

PIC

QIC

P- 10

P- 11

QIC

QIC

QIC

QIC

FIC

Fermentor Controls

71

Main Fermenter

Cooling WaterSparged

Steam

Vented Air 13

Sterile Air

Sterile Glucanex Sterile

Anti-foam

7

8

11

Filter 1

Filter 2

P- 8

Filter 3

TIC

14

Compressor 1Air

PIC

PIC

FIC

QIC

QIC

9

P- 3

TIC

10QIC

Visc

Foam

QIC

P- 6

Turbidity

QIC

QIC

Fermentor Controls

72

Rxn tank A

Rxn tank B

P-01

FermentedAlgae

FermentedAlgae

Sterilized AirSterilize Air

Vented Air

Conc. Algae

Water

45

18

16

15

17

Dissolved Air Flotation Controls

12

Sterilized Air

Sterilized Air

20

19

22 23

21

46Water

1

2

P10

P11

P12

Water to Cargill treatment

Tk-6

Tk-7

DAF Tk-7

PIC

Cv-21

Cv-23

PIC

FIC

Cv-20

FIC

18 18

Cv-22

FIC

FICFIC

Cv-24

Cv-25

FIC

Cv-26

73

3

Baffle trayTowerTk- 8

CO2 Lysing Controls

Conc. Algae

Sterilized CO2

CO2 AbsorbedAlgae

LysedAlgae&CO2

Vented CO2

LysedAlgae

ThrottleValve

23

24

Filter 4

CO 2

Compressor 2

25 26

27

Stir tank

P13

P1429

Lysed Algae TowerTk- 9

Cv-30

FIC

28

FIC

Cv-27

Cv-29

Cv-28FIC

Cv-30

4748

Steam in & out

TIC

Cv-31

PIC

74

Main Centrifuge Controls

Disc Centrifuge

Wet Biomass Slurry

Algae Oil3130

Oil settling tankTK-20

P15

Algae OilFICCv-32

Lysed Algae29

35

36Water

75

Extraction Controls

Oil

Settling Tank

Tk-11

Water

33

Disc Centrifuge

Lysed Algae

P16

40

30

34

38

32 Lysed Algae

Biomass

39

P18

P17

Water

45Water

FICCv-33

FICCv-34

Cv-35

FIC

76

Separation and Drying Processes Controls

Rotary

Dryer

42

Dynamic Cross Flow Filtration

Biomass P19

40

43

Biomass /water tank

Tk-12

Dry Biomass

P20

37 water vapor

41

Biomass

Biomass

FICCv-36

49Steam in

TICCv-38

50 Steam out

Conveyor Belt

Collection Hopper

Conveyor Belt

Shipping container

Table Spread Distributor

Dried Algae Distributor

39Water

Cv-37

P21

FIC

Degumming Controls

77

TK- 51Crude SBO

Storage

P-51

N2 GasTK-52

45% CitricAcid Storage

P-52

P-53P-54

TK-534% NaOHStorage

TK-5410% EnzymeRefrigerated

Storage

N2 Gas

HLO - Gums

BatchReactor

P-55

P-56

TK-55SurgeTank

N2 Gas

TK-56Degummed

SBOStorage

HX-5

CLR-1LLO

HX-6

Cent-5

TIC

TIC

FIC

PIC

PIC

PIC

TIC

pHCQIC

QIC LIC

TCFIC

LIC

LIC

Refining Controls

78

N2 Gas

TK-56Degummed

SBOStorage

HX-8

TK-5750% Citric

AcidStorage

P-57

P-58

TK-58Acid

ActivatedClay Tank

P-59

P-60

TK-59Water

Storage

P-61

HX-9P-62

VP-1

VacuumMixer

N2 Gas

TK-60SurgeTank

P-63

MX-1MX-1

VP-2

Steam

To MX-101

P-64

N2 Gas

TK-61Refined

SBOStorage

HX-10

HX-11

CLR-2

CLR-3

Filter

VacuumVessel

RP-1

TIC

TICTIC

FIC

FIC

PIC

PIC

PIC

PIC

PIC

FIC

To VacuumMixer

LIC

FIC

FIC

LIC

LIC

Interesterification Controls

79

N2 Gas

TK-61Refined

SBOStorage

N2 Gas

TK-62SBO

HardfatStorage

N2 Gas

TK-63SBO

HardfatHeating

Tank

P-66

P-67

HX-14

HX-15

P-68

TK-64CH3ONaStorage

TK-66MixingTank

P-69

P-70

Filter

TK-67Soft

WaterStorage

HX-16

HLO

P-71

P-72 P-73HLO

Cent-7Cent-6

TK-6520% Citric

AcidStorage

N2 GasTK-68SBO

Storage

CLR-4

MX-3 MX-4

WaterSoftener

P-65

TIC

TIC

TIC

TIC

PIC

PIC

PIC

PIC

FIC

TIC

FIC

QIC

FIC

FIC

LICQIC

TC

FIC

LICTC

TC

N2 Gas

PIC

FIC

QICLIC

80

E

Algae OilStorage

Refined SBO Storage

LecithinStorage

Beta CaroteneStorage

DiacetylStorage

SorbicAcidStorage

Mono- diglyceridesStorage

Ergo-califerolStorage

WaterStorage

Oil

Pasteurization

CIP

Warm Water

Aqueous PhasePrep.

TK-20

TK-21

TK-22

TK-23

CIPCIP

TK-24

TK-25

TK-26

TK-27

TK-30

TK-31

TK-32

Oil Phase Prep.Tank 1

CIP

P-30

P-31

P-32

P-33

Warm Water

Oil Oil Phase Prep.Tank 1

CIP

Sea SaltStorage

TK-29

P-34

TK-28

CIP

Margarine Controls 1

Warm Water

QIC

QIC

QIC

QIC

QIC

QIC

QIC

QIC

QIC

QIC

QIC

QIC

P-35

P-35TIC

TIC

81

High Pressure Pump

AmmoniaCompressor

Pin Rotor MachineScraped

Surface Heat Exchanger

Tub Filling and Packaging

P-37

CIPCIP

Margarine Controls 2

Plate HX

P-36

TIC

TICVIC

PICPIC

82

TK-41Acidic Solution

TK-42Alkaline Solution

TK-43Reclaim Water

TK-44FreshWater

Alkaline Storage

Acidic Storage

CIP Returning Liquid

Water

HX-41

CIP Supplying Liquid

Drain

P-41

P-42

PP

LIC

LIC

LIC

LIC

LIC

LIC

LIC

LIC

FIC

TICFIC Strainer

CS

CIP Controls

Control Descriptions:

83

Fermentation

P-1, 2P1 will be centrifugal pump on a quantity control switch allowing it to pump 5309.3 gal of glucose to MT-1. P2 will also a centrifugal be on a quantity control switch allowing the pump to transfer 156.4 gallons of ethanol bottoms to MT-1.

P-10P10 will be a rotary feeder an will transfer 4,385.2lb of crystalline sea salt to MT-1.

P-11P11 will pump culinary (soft) water into the mix tank and will be controlled by the quantity control switch measuring the transfer of heated water in stream 5 added to MT-1.

Valve 4aValve 4a will control the amount of steam added to the water stream in stream 4 by a temperature sensor and quantity control switch. Temperature of the water in stream 5 will be maintained at 140 °F by the addition of steam. Stream 5 will add 60617.3 liters of water to MT-1.

P-3Pump 3 will be a centrifugal pump and will be controlled by a quantity control switch to transfer 57,974.8 liters of medium to the Main Fermentor from MT-1. The switch will allow for a 2 hour mixing time in the Main Fermentor before transferring to an empty Fermenter.

Valve 8During sterilization, steam will be sparged in stream 8 and will be controlled by a temperature sensor and added in an amount, 15,926.8 lb, which will make the temperature of the broth 250 °F. The temperature will be maintained at 250°F for 1 hour after which the cooling water will be run at full flow rate.

P-7Pump 7 is a centrifugal pump and it will be controlled by a quantity control switch to transfer the seed batch volume to the main fermentor. The volume transferred will be 2100 gallons at approximately 80.6°F.

Compressor 1, Valve 12Compressor 1 is a centrifugal compressor and will be controlled by the pressure in stream 12 before the valve at a pressure of 3 barg. The valve will be controlled by a flow rate control, which will allow for a transfer of 158 normal cubic meters per minute of air.

P-5

84

Pump 5 will be controlled by a rotational viscometer transmitter monitoring a slip stream on the side of the fermentor and will transfer a volume of 420 gallons of glucanex solution to be filtered and transferred to the main fermentor batch when viscosity reaches 20cP.

P-6Pump 6 will be controlled by a rubber sheathed electrode monitoring the presence of foam in the fermentor. A maximum quantity of 3.6 gallons of silicone based anti-foam oil will be transferred to be heat sterilized and then to the Main Fermenter.

STEX-1The amount of steam transferred to the heat exchanger used to sterilize stream 10 will be controlled by a temperature sensor and a valve controlling the flow rate of steam at 293°F and 3 barg.

CW-1Cooling water flow into the cooling jacket on the main fermentor will be controlled by a valve and temperature sensor. The temperature inside the fermentor will be maintained at 80.6°F.

Valve 13Valve 13 will control stream 13, which is the vented air stream in the fermentor. The valve will be controlled by a pressure sensor in the vessel and will not allow the pressure to exceed 3 barg while steam sparging. Operating pressure will be maintained below 1.79 barg during fermentation and air sparging.

P-8Pump 8 will transfer the final volume of the fermentation broth to the harvest tank and will be controlled by a quantity control switch. The amount of liquid broth transferred will be 22,893.9 gallons when the fermentation has been completed. A turbidity sensor will monitor cell density and will determine when a batch is completed and when to transfer the broth.

Extraction and Lysing

Stream 15 (FIC)Stream 15 is divided into 2 equal streams, 16 and 17, that go into reaction tanks A and B and the feed inlet control valve (Cv-20) is a mass flow rate measurement equalizing valve that ensure that stream 16 and 17 are of equal valve and direct flow to one of the streams if they are not equal.

Reaction tanks A and BThe fermentation broth will be pumped out of each of these and controlled by mass flow meters (Cv-23 & 22) that allow the pumps to operate at specified gpm flow rate and the

85

valves will open or close adjusting for a constant specified gpm. The air inlet flow rate will be controlled by an inlet pressure valve (Cv-21) and the pressure is regulated at 500 Kg per hour, the valve will open or closed if the 500 Kg per hour is not maintained.

DAF tank 7The Dissolved Air Flotation tank has a regulating valve (Cv-24) for the in air that is maintained at 1000 Kg/hour and the valve will adjust to maintain this flow rate. The waste water being pumped out the tank will be maintained at specified gallons per minute by the regulating valve (Cv-26) and this valve will open or close to adjust for the correct gpm.

CO 2 Absorption TowerThe absorption tower is maintained at 150 PSI and a pressure relief valve (Cv-30) if the pressure is higher than 150 PSI the valve will open and relieve the excess pressure. The CO 2 is pumped into the tower at 150 PSI and the control valve (Cv-29) will open and or close to maintain the appropriate pressure. The tower also has a recirculation pump that operates at set gpm and valve (Cv-27) adjusts to ensure the correct gpm is applied for recirculation.

CO 2 Throttling and lysing Tower tank 9The control valve (Cv-28) maintains the 150 psi and will adjust according to maintain the pressure going into the throttling valve. The lysing tower is maintained at a temperature of 80ºC by steam and the valve (Cv-31) and this valve is a temperature control valve that opens and closes to keep the specific temperature of 80ºC. The lysed algae is pumped from the tower and controlled by mass flow meters (Cv-30) that allow the pump to operate at specified gpm and the valve will open or close adjusting for a constant specified gpm.

Storage tank 10The algae oil is pumped from this storage tank and controlled by mass flow meters (Cv-32) that allow the pump to operate at specified gpm and the valve will open or close adjusting for a constant specified gpm.

Stream 30The lysed aglae is pumped from pumped from the first centrifuge in a second centrifuge and is controlled by a mass flow meter (Cv-33) that allow the pump to operate at specified gpm and the valve will open or close adjusting for a constant specified gpm.

Settling tank 11The waste water being pumped out the tank will be maintained at specified gallons per minute by the regulating valve (Cv-35) and this valve will open or close to adjust for the correct gpm. The oil is be pumped from this tank also and will be maintained at specified gallons per minute by the regulating valve (Cv-34) and this valve will open or close to adjust for the correct gpm.

Biomass Storage tank 12

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The Biomass and water being pumped out of this tank will be feed into a third centrifuge and will be maintained at specified gallons per minute by the regulating valve (Cv-36) and this valve will open or close to adjust for the correct gpm into the centrifuge. The water leaving the centrifuge will be pumped into a series of pipes that collects all the waste water and vavle (Cv-37) will open or close the valve to ensure the proper gpm is maintained.

Rotary DryerThe rotary dyer uses steam to dry the biomass and the steam will be regulated by valve (Cv-38) which will regulate the steam inlet by opening and or closing to maintain the correct drying temperature.

Refining

TK-51,55,56,60,61,62,63,66,68 (PIC)All SBO storage tanks will have a nitrogen blanket of 0.5psig place on them. The controllers will be dead band controller to allow the pressure to get to the required amount, and then the valve will go shut.

P-51,52 (FIC)The flow into the batch reactor will be controlled to allow only 720 gallons of crude SBO to enter the batch reactor per batch. This quantity will be sensed by the controller and match the amount of 45% citric acid flowing into the batch reactor.

HX-5,6,8,11,14,15,16, Heating Coils of TK-63 and Vacuum Mixer, Cooling Coils of TK-66 and Batch Reactor (TIC)The SBO oil will be temperature monitored on the outlet of the heat exchangers in order to control the position of a valve positioned on the outlet of the heating/cooling medium to control flow. The heating/cooling coils will have temperature monitoring of the associated tanks. The temperature sensor will adjust a valve on the return line of heating/cooling medium.

Batch Reactor (pHC)The pH of the batch reactor will be monitored in order to maintain it at 4.8-5.0. A small pump will circulate the liquid from the batch reactor through a line where the pH will be monitored. The 4% NaOH will be added accordingly to maintain the proper pH.

Batch Reactor, P-54, P-55 (LIC, TC, FIC, TC)The level in the batch reactor will be monitored to control when the impellers can begin mixing. Once mixing begins, there will be a 30 minute time delay until the addition of the enzyme. The enzyme will be monitored for flow in order to secure the pump when the proper amount has been added. The mixing will continue for 6 hours, and then the

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mixer will be secured. Once the mixer is secured P-55 will pump the contents of the batch reactor to the surge tank.

P-56 (LIC)In order to create a continuous process, there will be level control from the surge tank starting the flow of P-56. The pump will shut off when level goes below a desired level.

CLR-1,2,3 (FIC)There will be continuous flow of cooling medium through the cooler. The hot liquid flow will be monitored and will adjust the outlet flow of cooling water in order to give an adequate cooling amount.

VP-1,2 (PIC)Pressure sensors will monitor the vacuum pressure in the tank. The vacuum pump will run continuously and have a recycle valve on it to maintain the proper vacuum pressure. The valve will open more in order to cause less of an increase in vacuum.

P-57,58 (LIC,FIC)The level in the tank will control the flow of P-57. The flow through the downstream line will be monitored and will match the proper flow of 50% citric acid through P-58.

RP-1 and P-60 (FIC)The flow will be monitored to determine the amounts of acid activated clay will be added to MX-1.

P-63 (LIC)The flow of oil from the surge tank will be controlled by the level present in the tank, which will control P-63.

Steam to Vacuum Vessel (FIC)The flow of oil into the vacuum vessel will be monitored by flow control. This will control the position of each of the four tray steam sparging inlet valves. The flow of steam will be matched against the flow of oil. The flow of steam will be monitored as well to better match the amount of flow of steam going into the vacuum vessel.

P-64 (LIC)The flow of oil from the vacuum vessel will be controlled by a level control switch which will control P-64.

Hardfat SBO Storage, P-65 (LIC, QIC)The flow of liquid from the tank is not a continuous process. The flow will be controlled by a level indicator to indicate that P-65 can pump to the processing tank. There will be a quantity controller in order to control how much is sent to the processing tank.

Refined SBO and Hardfat SBO (QIC)

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The quantities of the SBOs will be monitored to maintain the 20 to 80 ratio of hardfat to refined SBO going into the interesterification tank by control valves. P-66 and P-67 will be on the quantity control switch causing the flow of liquid into the tank. When the proper amount has been sent, the pumps will secure themselves.

TK-66, P-68, P-69, P-70 (LIC, QIC, TC, FIC)The level in the mixing tank will be monitored and once the proper level is reached the impellers will begin mixing. At the proper level the sodium methoxide will be introduced to the system in order to initiate mixing. The quantity of sodium methoxide will be monitored and when the 1.7 gallons of it has been added the pump will be secured. When the P-68 is secured and time control will be set for 30 minutes at which time the 20 citric acid will be pumped into the mixing tank to secure the interesterification process. The flow rate of citric acid will be monitored and P-69 will be secured when 17.21 gallons have been added to the tank. The tank will stir for 15 additional minutes at which time the impeller will be secured, and P-70 will begin running to pump out the tank. The amount of flow out of P-70 will be monitored at the outlet of the filter in order to control the pump.

P-71 and Soft Water Flow (FIC)P-71 will turn on the sensing of flow in the line going to the washing system. The amount of soft water going into each mixer will be controlled by sensing lines monitoring the SBO flow. The amount of soft water will correspond with 5 wt% of SBO.

Margarine Plant

P-30 and P-31

These pumps are controlled by a QIC control. The controllers measure the amount of SBO and algae oil and then shut off the pumps when the correct amount of each has been transferred.

TK-22, 23, 24, 25, 26, 27

The amounts of ingredients added to the margarine are controlled by QIC controls that open and close valves added to the line. These ingredients are gravity controlled. When the right amount of ingredients have been transferred, the controller closes the valves.

P-32

This pump is controlled by the water flow. Once enough water has entered the water preparation tank for one batch, then the pump is turned off.

TK-29

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The flow of the salt as it leaves the tank is controlled by this controller. Once the right amount of salt for one batch has left the tank, then the valve is closed.

P-33 and P-34

Each of these pumps are controlled by the flow out of the preparation tanks. Once the tanks are emptied, then the pumps are shut off.

Hot water inflow valve

This valve controls the amount of hot water that flows into the jackets of the emulsion tanks. It is controlled by the temperature of the emulsion inside the tanks. The valve can be opened in different degrees, such as totally closed, 50 % open, 75 % open, et cetera. If the temperature of the emulsion inside the tanks is too low, then the valve is opened to allow for more heat to transfer to the fluid. If the emulsion is too warm, the valve is closed to compensate.

PHX Hot Water inflow Valve

The plate heat exchanger’s hot water inflow is controlled by the margarine outflow temperature. If the emulsion is leaving the plate heat exchanger too cool, then the hot water inflow is increased. If the emulsion is too warm, the valve is partially closed to lower the amount of heating.

HPP

This plunger pump is controlled by the pressure of the outflowing emulsion. The pressure of this fluid needs to be 75 bar. If the pressure isn’t high enough, then the valve increases the amount of pressurization that occurs within the pump.

SSHE

The control for this piece of equipment comes from the temperature of the crystallized margarine just as it leaves the heat exchanger. The temperature of the fluid at this part of the flow should be 50 °F. The valve controls the ammonia refrigerant. If more cooling is needed, the valve is opened more fully, allowing more ammonia to flow into the SSHE and increasing the amount of cooling.

Pin Rotor Viscosity

A separate line that splits off the main line, using a very small recirculation pump, allows for this controller to check the viscosity of the margarine as it leaves the pin rotor machine. If this value is not ideal, the controller can increase or decrease the pin rotor machine’s kneading, raising or lowering the margarine’s viscosity until it is ideal.

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Appendix 12

General Arrangement – Major Equipment Layout

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Overall Plant Layout:

Otis Rd

Cargill

A

B

C

D

E

F

A. Storage/BoilersB. FermentationC. Offices/Labs/Miscellaneous SpaceD. Water Cooling TowerE. SBO RefiningF. Margarine Processing

Wind DirectionGate

General Plant LayoutAlpha Algae Plant1928 SE Otis RdCedar Rapids, IA 52401

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S2

Seed 1

S1

S4

S3

Seed 2

Seed 3

Seed 4

Mix Tank

Fermenter 1

Fermenter 2

Fermenter 3

Fermenter 4

Harvest Tank

DAF

RXN 1

RXN 2

Absorption

Lysed Tank

Lipid Storage

Centrifuge 1 Centrifuge 2

C3 C4 Dryer

Glucose Storage

Ethanol Bottoms

Glucanex Antifoam

Salt Boiler

Cooling Tower

Electrical Room

85 ft

35 ft

115 ft

Fermentation Layout

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SSHE

AmmoniaCompressor

Filling,Packing

Margarine Plant First Floor Layout

= 4 feet

CIP system

Emulsion Tanks

Ingredient PreparationTanks

HPP

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Margarine Plant Mezzanine Layout

Electrical Room

Ingredient Storage Tank Area

= 4 feet

Appendix 13

Distribution and End-use Issues Review

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Once fully processed and packaged, Golden Premium Spread will be distributed

across the United States by a trucks owned by a wholesale company. This wholesaler

will be fully experienced in and authorized to handle and distribute food products safely

and professionally, and the these trucks will utilize refrigeration to keep the product fresh

and safe for all consumers to enjoy. Despite the professionalism of the distribution

service, problems could still arise. It has to be assumed that not every truck will reach its

destination without problem, but we assume that all of the trucks will be able to distribute

the product without issue or loss.

The product has been processed carefully in order to ensure the health of the

consumer, however contamination could possibly arise at some time in the future. If any

of our valued customers becomes sick as a result of our product, then production will

immediately cease and a recall will be issued. However, the chances for such a recall are

very slim due to the careful processing and strict sanitation standards that will be strongly

enforced in the production facility. Each margarine tub will be marked for identification

and accountability in case of any contamination instances, making the problem batch or

batches easy to find and recall. Such an issue will not occur in such a carefully designed

facility except for gross negligence from a worker in the facility, so each worker will be

trained in sanitation and cleanliness.

Some consumers will be concerned about complications that may occur as a result

of consuming too much of the product. Some studies have claimed that consuming too

much DHA could have adverse side effects like increased bleeding, increased risk of

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heart failure for people with congestive heart failure, or even hemorrhagic strokes.

However all of these possible side effects only occur with extremely large doses of

omega-3 fatty acids. In Food and Drug Administration has stated that intake of DHA and

other omega-3 fatty acids is generally regarded as safe (GRAS) in doses of up to 3000 mg

per day. In order for a consumer to reach that level, he or she would have to consume

almost an entire tub of Golden Premium Spread. Therefore, a problem in which a

customer overdoses on DHA from eating Golden Premium Spread should not occur.

Omega-3 are considered GRAS which means that the margarine product will be safe.

Appendix 14

Constraints Review

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Despite all precautions and careful design, it cannot be assumed that each

fermentation batch will be successful. Industry standard is to assume that approximately

1/1000 batches will go wrong in some way, whether through contamination or some other

flaw. Another constraint that arises in the fermentation period is the growth rate of the

algae. Our fermentation system has been designed to utilize the maximum, exponential,

growth of the algae, however, a limit is imposed on the system once the algae reach a

certain concentration. This concentration is avoided during the step-up procedure, with

the growing algae placed into larger and larger containers in order to lower the

concentration and keep the algae from approaching the stationary phase of growth, but it

is inevitable that at some point, the algae will stop growing. Also, the algae can only

make a certain amount of DHA concentrated lipids during the starvation portion of the

fermentation. There is no known way to increase the DHA content in these lipids, so

only 40 % of the lipids are DHA.

Sanitation is a constraint to this process as well. Every vessel needs to be

regularly cleaned in order to keep the final product free of any pathogenic microbes. This

means that the entire system must be continually cleaned, and it cannot run continuously.

There are enough vessels in order for cycling to occur, however. This means that while

one vessel is being used, another can be rigorously cleaned with the CIP system. A

consumer becoming sick after ingesting the product would have catastrophic results on

the market share of the product. Care must be taken in the cleaning of the system and the

entire plant, which lowers the amount of actual production that can be done.

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Another major constraint on this design is market demand. It is important to

make just enough product to meet the market demand, but not so much that some of the

product goes unsold. In order to ensure that the product just meets the market demand,

the first year of production will just meet the 1 % market share level. If too much is

made, then money will be wasted in the production while unsold tubs of Golden Premium

Spread pile up or have to be discarded. In this way, the market is the most major of the

constraints on this process.

Appendix 15

Applicable Standards

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The final margarine product must comply with FDA regulations about vitamin

content in margarines. All margarine products must replicate the nutritional content of

butter, so all margarines are fortified with Vitamin A and many are fortified with Vitamin

D as well. Golden Premium Spread is fortified with both Vitamin A and D which are

included in the list of ingredients. From the nutritional label figure, it can be seen that

our product will supply 10 % of the recommended daily dosage of Vitamin A in every

serving. It will also include 10 % of the recommended daily dosage of Vitamin D with

every serving.

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Other than the vitamin requirements that the final product must have, the most

important applicable standard to which AlphaAlgae’s Golden Premium Spread facility

must conform are sanitation standards. Obviously, sanitation and immaculate cleanliness

are of great importance to a food plant. Without these standards, consumers can become

sick, which a public health issue. Both for the safety of the valued customer, and for the

company’s profit, the Golden Premium Spread plant has been carefully designed from the

bottom up to conform to these applicable sanitation standards. Most of these standards

come from the IMACE Code of Hygiene which was written in January 1999. These

codes inform the design engineer how to design a plant which will manufacture fat

spreads using good manufacturing practice. Some of these standards are as follows:

Equipment Design• Cleanable to microbial level• Made of compatible materials• Accessible for inspection, maintenance and cleaning• No liquid collection and no niches• Equipment must perform as designed• Hygienic compatibility with other plant systems• Validated cleaning procedures

Facility Design• Physical separation of distinct hygienic zones• Material flow and personnel movement control • Prevent water accumulation inside building materials• Temperature and humidity control• Air flow and air quality control• Building envelope sanitary conditions• Interior spatial design that enables cleaning• Sanitation integrated into facility design

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The IMACE Code of Hygiene also describes the need for fat products to conform

to standards regarding the taste and consistency of the final product, as well as its mouth

feel. Golden Premium Spread should be a solid, but spreadable, at refrigerator

temperatures, softer but still a solid at room temperatures, and should melt at mouth

temperature. This feeling of having the table spread melt in one’s mouth, literally, is a

standard to which AlphaAlgae’s Golden Premium Spread adheres. Using industry

standards for the margarine processing and ingredients keeps the process clean and free

of contamination as well as providing a superior product that will not only attract new

customers, but will keep customers. The overall goal is to keep customers coming back

to our table spread every time the customer goes to the health food grocery store.

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Appendix 16

Projects Communication File

Middough Visit March 22- Group presented to John Micheli-Filters should have magnets-CIP is a daily process, but with evidence of good cleanliness can make it once a week, we will use 1/week-Will operate 24 hrs, with only a 2-3 workers to monitor fermenters-Want flow into dryer to have lowest possible water content-Vacuum pumps-steam boosters with 3-4 stages-SBO storage tanks have nitrogen blanket of 0.5 psig is sufficient-Use a homogenizer as the HPP-Pump out of the votator is for back pressure regulator-CIP filter should be an inline strainer-CIP alkaline and acid tanks are probably industrial 55 gal poly bottles with installed flow meters-Must have code, date for each box and tub-Have workers packing boxes with tubs-Pastuerization is a piping run of 30 minutes at the constant temperature

Meeting Notes - 3/11/2009 - by CharuPresentation Feedback:

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Strong Points: Well organized PowerPoint, maintained standard time, Handouts ( Mass balance), Q&A

Improvements: Units consistency, Give a summary before and after the presentation and mention future works

List of things we need to work on:1. Have drawing standards, Assign numbers for streams used in each process and

use same format for spreadsheets.2. Work on Instrumentation of PFD, using Icarus.3. Work on Process description: Including Equipment list and details, Chemistry

Section, Products section4. Break down the cost estimate, have estimate cost for the building, using Icarus.5. We should avoid using centrifuges just to keep our capital cost down.6. Land cost for our project is similar to cost of produce the amount of glucose we

need.7. Dan will be sending the contact information to find more about the dryers.8. For the CO2 lysing we only need 2 feet diameter column, as a continuous process

Meeting Notes - 3/4/2010 - by Travis

The following topics were discussed during a conference call: Enzymes can be stored in a kegerator at the required temperature, just label it on

the PFD and designate the temperature on the mass balance sheet Heat exchanger should assume a overall heat transfer coefficient for all heat

exchangers and go from there Cooling water inlet at 85 F and out at a maximum of 120 F Determine the duty required for heaters, boilers, cooler, ammonia system. Allow

Aspen to determine sizing and pricing required Discussed putting things into Aspen to determine the pricing Discussed the washing process of SBO For the presentation, have handouts for the mass balance and any other specifics

associated For the presentation order, have overall block flow, pfds, and whatever else Will be sending chemical list pricing to Dan by Friday Will send presentation via email to Dan 24 hours prior to Tuesday

Meeting Notes - 2/26/2010 - by Ramune

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The following topics were discussed during the phone conference: The C02 for our fermentation will come from ethanol refinery The prices of commodity chemicals can be found in the library sources Look at the different extractors GEA separators Disk centrifuge Agitation and heat transfer from biochemical engineering books Need to make overall Block Flow Diagram including hexane Need mass balance kg/batch Every 24h equipment must be cleaned - five step process CIP Fermentation vessel is sterilized Energy balance on glucose going in Respiration rate The design of the dissolved air unit can be found in engineering book Use ASPEN to generate prices of equipment Peristaltic pump for SOB process

Meeting Notes -2/23/2010 -By Ramune

Our mentor Dan came to meet with us at CEB. The following topics were discussed: Communication between group members must be improved - Dan suggested

working together and exploring all ideas Color code was used on our block flow diagram; red - add certain things, green -

take the stuff out Always have the back-up slides for the equipment or process that was not chosen

by us, but it is alternative to our process or equipment Do not take prices from the catalog (reagent grade for laboratory purposes) Chemical properties of sodium methoxide Bottoms that are used for the fermentation is not yeast In order to find the price of certain equipment or component we can look at the

alternative and estimate the price Vitamin D comes from shrimp For the incoming air use the absolute filter (about 400hp motor) Need to have drop tanks and storing capacity for our raw materials (1.5 times the

size of the truck) Our process is combination of batch and continuous Use enzyme to brake slime after CO2 lysing Use hexane after centrifuge Dry the biomass in direct dryer Cascaded CSTRs are like PFR and can be replaced by batch reactor

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Refrigeration unit will come with margarine equipment

Meeting Notes – 2/18/2010 -By Charu

1. Presentation was missing the overall mass balance2. BFD needs to improved ( It should serve as a road map of the process) and needs

specific streams name3. PFD needs to be developed from BFD4. We need to use consistent units ( lbs/hr is preferred)5. Cost Estimate on main equipments6. Utilities and waste products ( all inputs and outputs) should be in diagram7. For recovery process use the minimum antifoam8. analyze gas absorption, mix system, dissolve oxygen, power number and agitator9. How and why are we starving algae? By limiting amount of Nitrogen and to let

algae produce oil.10. Make a page on Wikispaces for all the assumptions made in the process.11. Hexane Extraction method was discussed taking account for carotene present in

the oil12. Eric will be talking to John for the IPE program13. Talked about, how nickel rainy works.14. Hardening Ratio for the Margin is 20:8015. We need refrigerated room for margin16. Look into Pesticide control, Clean up system, Sanitary design and EPA system

Phone conference 2/11/2010 - By Ramune

All group members were participating in the phone conference. The following topics were discussed during our conference call with mentor Dan:

Making margarine in our facility and shipping it off We looking to take 1% of butter/margarine market and base the rest of the process

on this number Use available recipe for margarine Show how margarine plant opperates Need to make contact with Cargill The fermentation vessel size is standard Scale-up gradually from lab to production vessel We will be using glucose for fermentation (buying from Cargill, need to figure

out the pricing) Lysing with CO2 and remaining biomass will be treated with hexane We will base our marketing on the trends of “Whole Foods” Need to make detailed block flow diagram Calculations in excel spreadsheets

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Phone Call 3 Notes - by Charu

1. Each group member talked about whatever they found through their research.2. We decided to run the batch process as compare to continuous process3. Economy Calculations – Travis needs to look for how the market for spread butter will grow? Then do the rough calculations on net present value. We also need to take in consideration that 5-10% of our process will be spoiled. Yield will be affected.4. We will have to buy 0 trans fatty acid of certain melting point to mix with our product (lipid) to make the spread. That can be purchased from Cargill or AVM , etc.5. Do we need to refine the crude oil? No, because it doesn’t contain any harmful material when compared with FDA standards. There is no strong smell in the lipid, so little favoring should work for deodorization6. Bio-reactor- How the heat will be removed from? Cooling jacket? Coils inside or out the reactor? Because increasing the surface is not the only way.7. Yeast Extract- How yeast will be used in the process?8. Extraction Processes where discussed. Volume will be reduced in order to operate the lysing process. We want to produce the lipids in liquid form after all the processes.9. Two Different separation methods were mentioned single-stage separation and Countercurrent separation.

Phone Call 2 Notes - by Dan

Team Alpha: Re On Stream factor, Yields, Pricing.

You have determined how much you want to sell. Now you have determine how many batches you need to produce.

The only commercial fermentation process is ethanol. It has many biological pressures to keep it from being contaminated. Yet only one technology provider, Volgebusch, guarantees their process. Many process that started off as continuous have switched back to batch. If a batch goes bad you have the other fermentors to continue manufacturing.

No plant operates 100% of the time.The best chemical plants operate 85% 0f the time or 7500 hr/year. Federal holidays, plant utilities shut downs, no feed stock, unschedule break downs, waiting for analytic results, batches gone bad, off spec material, acts of god etc.

Besides the actual operating time you have to allow time for discharge, cleaning, charging, sterilizing, induction time etc.

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No reactions are 100% complete nor are recoveries 100% complete. There are losses at every step. Including wash out losses when cleaning equipment. Even when shipping finished product to customers. They all effect the overall yield from raw materials to sold goods. As mentioned above some batches go bad and have to be wasted. Not only loss of reactor time but also loss of raw materials.

The selling price in the is not the price that you sell it for as it leaves the plant. Retailers typically buy goods at 50% of what they sell it for. The wholesaler that you sell it to also has to make money to cover his warehousing and distribution cost. So your selling price may only be 35% of what the supermarket sells it for. Confirm the number for table spreads.

A good deal of chemical engineering is business.

Phone Call 1 notes - By Charu• Decided as a group to go with functional food• Discussed about the economy and market of the product• Everyone discussed about what they are working on.• Tried to come up with site location ( FL, CA ?)• Specified the type of Bioreactor we will be using ( Clear tubes)• Discussed if product is in or outside the cell.• Got the reference for oilgae• Referred to look more into:Membrane Filtration, Reusing the water, Gross separation, concentrating the solution, Acrylic Chemicals, Cross flow Membrane

First Meeting Thursday Jan 21 Notes - By Charu• Start wikispace and set communication matrix of group members• Whenever emailing Dan, make sure to email at both email ids written on the business card• Importance of Fatty acids DHA , Omega 3 acid• Main Source of Fish Oil Algae• Selling Target $3-4/gallon• Open pond vs. closed pond Depends on water sources, yield, carbon dioxide utilization, sulfur dioxide extraction if used from power plant, contamination• Nutraceutical we can make functional food, use the nutrients available along with color of alage.• Yellow color Pro Vitamin A, using double bond absorption makes it saturated and color goes away.• DHA Properties , 6 Double Bonds

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• Other fatty acids zero trans fatty acid• Resources Bailey’s Books• Goal Make a flow diagram and discuss the challenges and uncertainity• How Lipids are transported? High molecular weight compounds aren’t absorbed by the body• Sanitation Big issue in Designing Nutraceutical products• Product Making Butter, supplements or just crystallize the fatty acids and sell them to food industries• Bioreactors requires enrich source of CO2• Target, Market, Money 2 acres?• Process of growing algae Use sunlight or feed glucose in dark?• Recylcing very important• Economy • Make the Block diagram of the project• Harvesting microfilteration , centrifugation• Hardening & interesterfication process• Bulk or fractional extraction?• To lower the price of butter spread, we can use soybeans to go trans fatty acids• Can DHA and carotenes be removed?

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Appendix 17

Information Sources and References

Advanced Dairy Chemistry. Volume 2: Lipids. Ed. by P.F. Fox and P.L.H. McSweeney. New York: Springer Science + Business Media, Inc., 2006

Amato, T. et al. “An integrated approach to dissolved air flotation.” Water Science & Technolgy Ed. Peter Wilderer. Germany: Technical University of Munich, 2001. 19-26. Print.

Bailey, James E. (James Edwin), 1944-. Biochemical Engineering Fundamentals / James E. Bailey, David F. Ollis. Ed. David F. Ollis joint author. New York : McGraw-Hill, 1977. Web.

Arterburn, Linda M., et al. “Bioequivalence of Docosahexaenoic Acid from Different Algal Oils in Capsules and in a DHA-Fortified Food.” Lipids (2007) 42: 1011-1024

Baker, R.W. Encyclopedia of Separations Science. Academic Press, 2000. Web 15 Feb. 2010.

Beal, R.E., Sohns, V.E., “Treatment of Soybean Oil Soapstock to Reduce Pollution.” Journal of the American Oil Chemistrys' Society (1972) 49, No.8: 447-450.

Converti, Attilio, et al. "Estimation of Viscosity of Highly Viscous Fermentation Media Containing One Or More Solutes." Biochemical engineering journal 4.1 (1999): 81-5. Web.

da Silva, Teresa Lopes, et al. "Effect of n-Dodecane on Crypthecodinium Cohnii Fermentations and DHA Production." Journal of industrial microbiology & biotechnology 33.6 (2006): 060601. /fndfed/zgw.ebscohost.com:210/aph/. Web.

da Silva, Teresa Lopes, et al. "The use of Multi-Parameter Flow Cytometry to Study the Impact of n-Dodecane Additions to Marine Dinoflagellate Microalga Crypthecodinium Cohnii Batch Fermentations and DHA Production." Journal of industrial microbiology & biotechnology 35.8 (2008): 080801. /fndfed/zgw.ebscohost.com:210/aph/. Web.

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