Bio-processing overview (Upstream and downstream process)

Summary

This video offers an insightful overview of bioprocessing, a technique that harnesses biological resources like living cells to produce biomedically relevant products such as enzymes and recombinant proteins. It explains the process taking place in bioreactors, involving various cell types, and differentiates between upstream and downstream processing. Upstream focuses on cell growth and product generation, while downstream encompasses recovery, purification, and quality control of the synthesized products. The video also touches on industrial-scale challenges and the equipment used, such as centrifuges and chromatography systems, highlighting the transition from lab-scale to bulk production in the bioprocessing industry.

Takeaways

• 🧪 Bioprocessing is a technique that utilizes biological resources such as living cells to produce enzymes, metabolites, and other products with biomedical relevance.
• 🌱 The process takes place in a bioreactor, which provides a controlled environment for the biological cells to function, and can involve bacteria, mammalian cells, or plant cells.
• 🛠️ Bioprocessing is an industrial-scale process, differentiated from lab-scale by the scale of the reaction, not the type of reaction.
• 🔬 Products generated through bioprocessing can include secreted metabolites, enzymes for research or therapeutic drugs, and recombinant proteins for biomedical research.
• 🏭 The industrial production of biological products involves scaling up from a small culture volume to a large bioreactor, where the product is generated in bulk.
• 🌱 The bioprocessing industry starts with a seed stock of bacteria, which is then transfected with a plasmid and scaled up to produce the desired product.
• 🔄 The process involves both upstream and downstream processing; upstream refers to cell growth and product generation, while downstream involves recovery, purification, and packaging of the product.
• 🌡️ Bioreactors are equipped with various components including a sterile tank, stirring mechanism, temperature control, and aeration to ensure optimal conditions for cell growth.
• 🔬 Centrifuges and filtration units are used in the downstream process to separate cells from the product and to purify the product based on its location (intracellular or extracellular).
• 🔄 Cell disruption systems are used for intracellular products to release the product from within the cells, followed by purification steps such as chromatography.
• 🏷️ Quality control is a critical step in bioprocessing to ensure the functionality and safety of the product before it is released for use.

Q & A

What is bioprocessing?

Bioprocessing is a technique that utilizes biological resources, such as living cells, to produce products like enzymes and metabolites with biomedical relevance in a controlled bioreactor environment.

What are the types of biological cells that can be used in bioprocessing?

The biological cells used in bioprocessing can be bacteria, mammalian cells, or plant cells.

What are some examples of products generated using bioprocessing?

Examples of products generated using bioprocessing include secreted metabolites, enzymes for biomedical research, therapeutic drugs, and recombinant proteins.

What is the purpose of a bioreactor in bioprocessing?

A bioreactor is used to carry out the bioprocess in a controlled environment, allowing for the growth of biological cells and the production of the desired products.

How does the scale of bioprocessing differ from a small-scale lab experiment to an industrial process?

In a lab, a small vessel is used for a small scale work, whereas in an industrial process, a much larger bioreactor is used to produce the product in bulk, catering to a larger demand.

What is the role of a seed stock in bioprocessing?

A seed stock contains bacteria that are transfected with the desired plasmid. It serves as the starting point for gradually scaling up the culture volume in bioprocessing.

What are the components of a bioreactor?

A bioreactor consists of a sterile tank for the broth, a stirring paddle attached to a motor, a coolant system to maintain temperature, an aeration unit for oxygen supply, and electronic control systems.

What is the difference between upstream and downstream processing in bioprocessing?

Upstream processing involves the generation of cells in large amounts and the production of the product, while downstream processing includes the recovery, purification, polishing, quality control, and packaging of the product.

How is an extracellular product purified and recovered in bioprocessing?

An extracellular product is purified and recovered by connecting the bioreactor tank to a centrifuge and then to a filtration unit, which separates the cells from the metabolites and purifies the product of interest using column chromatographic techniques.

How is an intracellular product such as an enzyme recovered in bioprocessing?

An intracellular product is recovered by first disrupting the cells using a cell disruption system, which mechanically shears the cells to release the product. The product is then separated from the debris using centrifugation and further purified through downstream processing steps.

What is the importance of quality control in bioprocessing?

Quality control is crucial in bioprocessing to ensure that the final product is functional, meets the required specifications, and is safe for release into the market.

Outlines

00:00

🌱 Introduction to Bioprocessing

This paragraph introduces the concept of bioprocessing, a technique that utilizes biological resources such as living cells to produce biomedically relevant products like enzymes and metabolites. It explains that bioprocessing occurs in a controlled environment known as a bioreactor and involves biological cells, which can be bacteria, mammalian, or plant cells. The paragraph also distinguishes between different types of products generated through bioprocessing, such as secreted metabolites, enzymes, and recombinant proteins. The speaker uses an analogy of cooking to explain the scale difference between laboratory and industrial bioprocessing and sets the stage for a deeper dive into the industrial aspect of bioprocessing, including the production of enzymes in bulk by companies.

05:01

🔬 Bioprocessing Industry Overview

The second paragraph delves into the specifics of the bioprocessing industry, starting with the seed stock containing bacteria that are transfected with a plasmid to express the desired product. It outlines the process of scaling up the culture volume and the use of a bioreactor for product harvesting. The paragraph describes the downstream processing, which includes cell harvesting, centrifugation, separation, purification, and quality control, emphasizing the importance of each step in ensuring the product's functionality and market readiness. The speaker also explains the components of a bioreactor, including the tank, stirring mechanism, aeration unit, and control systems, and discusses the industrial-scale centrifuges used in the process. This paragraph provides a comprehensive overview of the bioprocessing steps, from upstream processing in the bioreactor to downstream processing for product recovery and purification.

10:02

🛠️ Purification and Recovery of Bioproducts

The third paragraph focuses on the purification and recovery of bioproducts, distinguishing between extracellular and intracellular products. For extracellular products, the process involves centrifugation to separate cells from metabolites, followed by filtration and chromatographic techniques to purify the product of interest. In contrast, intracellular products require cell disruption systems to release the product, such as enzymes, into solution. The paragraph explains the use of industrial-grade pistons for mechanical shearing and the subsequent steps of purification, including dewatering, concentration, and chromatographic methods like HPLC. The importance of polishing steps to achieve high purity and quality control to ensure product functionality and preservation is highlighted. The paragraph concludes with the packaging and distribution of the final bioproducts, providing a complete picture of the bioprocessing journey from raw materials to finished goods.

Keywords

💡Bioprocessing

Bioprocessing is a technique that utilizes biological resources, such as living cells, to produce products like enzymes and metabolites that have biomedical relevance. It is central to the video's theme as it is the overarching process being discussed. The script explains that bioprocessing takes place in a bioreactor within a controlled environment, highlighting its importance in producing substances for biomedical research and therapeutics.

💡Bioreactor

A bioreactor is a controlled environment where bioprocessing occurs. It is a key component in the script's explanation of how biological cells are used to generate products. The bioreactor is described as having various components like a tank for the broth, a stirring mechanism, a cooling system, and an aeration unit to ensure the right conditions for cell growth and product generation.

💡Biological Cell

Biological cells, such as bacteria, mammalian cells, or plant cells, are the key players in bioprocessing. They are used as the machinery to produce the desired products. The script mentions that the type of biological cell used can vary depending on the product being generated, which underscores the versatility of bioprocessing.

💡Metabolites

Metabolites are one of the types of products that can be generated through bioprocessing. They are chemical compounds resulting from metabolism and can have significant biomedical relevance. The script uses metabolites as an example of a product that could be secreted outside the cell during the bioprocessing.

💡Enzymes

Enzymes are proteins that act as catalysts for biochemical reactions and are also a product of bioprocessing. They are highlighted in the script as being relevant for biomedical research and potentially important as therapeutic drugs. The video discusses how enzymes can be produced at an industrial scale for use in research and medicine.

💡Recombinant Proteins

Recombinant proteins are proteins that are produced by an organism that has been genetically engineered to express a gene from another organism. The script mentions recombinant proteins as one of the products generated using bioprocessing, used for biomedical research, indicating their significance in scientific and medical applications.

💡Upstream Processing

Upstream processing refers to the initial steps in bioprocessing where cells are generated in large quantities. It includes inoculum development, media development, and optimizing growth kinetics. The script explains that this phase occurs in the bioreactor and is crucial for the efficient production of the desired substances.

💡Downstream Processing

Downstream processing involves the recovery and purification of the biosynthetic product generated during upstream processing. The script describes this as a series of steps including centrifugation, filtration, chromatography, and quality control, which are necessary to purify and prepare the product for use.

💡Cell Disruption

Cell disruption is a process used to break open cells, particularly for the extraction of intracellular products like enzymes. The script describes industrial-grade pistons that mechanically shear cells to release the product into solution, which is then further processed and purified.

💡Quality Control

Quality control is a critical step in bioprocessing that ensures the product is functional and suitable for release into the market. The script emphasizes the importance of this step to verify the purity and efficacy of the product before it is packaged and distributed.

💡Industrial Scale

The term 'industrial scale' refers to the large-scale production of bioprocessing products. The script contrasts lab-scale production with industrial scale, highlighting the need for companies to produce and purify liters of enzymes and other products to meet global demand, which is a significant aspect of the video's discussion on bioprocessing.

Highlights

Bioprocessing is a technique that uses biological resources like living cells and their machinery to produce biomedically relevant products.

Bioreactors are used in bioprocessing to provide a controlled environment for biological cells to produce products.

The key player in bioprocessing is the biological cell, which can be bacteria, mammalian, or plant cells.

Bioprocessing can generate secreted metabolites, enzymes, and recombinant proteins for biomedical research and therapeutic drugs.

An analogy is made between cooking for a family versus a community to explain the scale differences in bioprocessing.

In labs, enzymes are produced through cloning and transfection into bacteria, which is then scaled up for industrial production.

Companies produce enzymes and proteins in bulk using bioprocessing techniques for research and therapeutic use.

The bioprocessing industry starts with a seed stock of bacteria that is transfected with a plasmid to express the desired product.

Culture volume is gradually scaled up from small to large in bioreactors for the production of large quantities of the product.

Products are harvested from the bacteria through centrifugation and chromatographic separation methods.

Quality control is crucial to ensure the functionality and safety of the bioprocessed products before market release.

Upstream processing involves the generation of cells in large amounts in bioreactors, while downstream processing includes recovery, purification, and packaging.

Bioreactors consist of a sterile tank, stirring paddle, DC motor, coolant system, aeration unit, and electronic control units.

Industrial-scale centrifuges are used to separate cells and metabolites in the bioprocessing of extracellular products.

Intracellular products require cell disruption systems to release the product, followed by purification steps.

Concentration and dewatering steps are essential for efficiently recovering products from large volumes of culture broth.

High-throughput column chromatographic methods such as HPLC are used for the initial purification of the product.

Polishing steps ensure the product is 98 to 100 percent pure, and quality control is vital for product development.

The video provides an overview of bioprocessing, with subsequent videos diving into more detailed aspects of the process.

Transcripts

hello everyone

in this video we would learn about

bioprocessing and it's just an overview

of that

so what is bioprocessing

bioprocessing is a technique

by which we can use biological resources

such as

living cells and using their machineries

we can use products such as enzymes

metabolites etc which have important

biomedical relevance now

definitely the bioprocess would take

place

in a bioreactor in a controlled

environment right

and the key player of this bioprocessing

is the biological cell sometimes it

could be a bacteria sometime it could be

a mammalian cell

or sometime it could be a plant cell as

well so bioprocess

involves bacteria mammalian cell or even

cell free systems

now let's try to understand what type of

products

could be generated using this bio

processing method

the product could be secreted metabolite

which is secreted outside

the product could be an enzyme which is

relevant for biomedical research or

maybe this enzyme is

important therapeutic drug now

it could be also a recombinant protein

used for biomedical research so all of

these type of products can be generated

using a bio processor now the

bioprocessor can

produce these products in a controlled

environment

but before we understand bioprocessing

we need to understand some very basics

imagine you're cooking for your family

so you need a very small vessel right

but

when you cook for the whole village or

the whole community you need a bigger

cooking pot so the environment where

these

cooking would happen is different but

the cooking is same

the reactions are same but only the

scale is different

similarly bioprocessing is also an

industrial process or it's a macro level

reaction okay so let's try to take a

specific example

so let's say you work with this

particular enzyme

and you want this enzyme to be produced

so what you are going to do

in lab if you need these enzyme for a

small scale work

you would definitely try to generate

this enzyme

you would use the cloning workflow where

you

clone your gene of interest using the

cloning methods

and ultimately you would transfect that

recombinant plasmid

into a bacteria this plasmid would

express

its product inside the bacteria and you

would grow the bacteria as the bacteria

grows the product also grows inside the

bacteria later on

using column chromatography and other

chromatographic tools you would

purify that protein and finally

that protein is with you and it

would be enough for your work but

imagine

once you need a some kind of product

such as an antibody

such as a restriction enzyme or such as

recombinant proteins

for your own research each time

you are not going to make it from the

scratch right

you are going to look for companies who

deliver

all of these enzymes right and

the question is how does these companies

produce these things in bulk the company

would deliver these products to your

doorstep and you would start

using it but the question is how does

the bulk preparation takes place

inside the industry right so we are

going to talk about that

industrial aspect of biological

processing so obviously

the overall formula is same you need to

have a recombinant plasmid you need to

have

bacteria which would be generating the

product but

the reaction container or the reaction

conditions

are different and the scale is different

so you can clearly understand in a

factory there is a amp up

in the culture volume from which you are

purifying

in lab you might need one microliter of

this particular enzyme

but worldwide people need one microliter

so

the factory has to generate liters and

liters of that

enzyme and that's is not an easy process

so let's begin from the scratch what is

happening inside of bioprocessing

industry

so they have a seed stock so this seed

stock would contain

bacteria you would transfect the

bacteria with the plasmid

that you want to express inside this

bacteria

now you would gradually scale up your

culture volume from a small culture

volume to a big one and ultimately you

would take your culture

into a fermentation reactor or a

bioreactor

we'll come to the details of the

bioreactor in a moment

but in the bioreactor your product would

be harvested that means your bacteria

would grow in number as the bacteria is

growing in number your product is also

generated

now after that your product need to be

recovered from the bacteria right

so you have to harvest the cells and you

have to do centrifugation

followed by some kind of separation by

chromatographic method

then there would be several rounds of

purification steps polishing step

and ultimately it would pass through a

quality control

you have to understand whether the

product which is generated is actually

functional or not whether it is okay for

it to be released in the market so

quality control step is very important

and ultimately it would be packaged and

delivered

to its proper location so the step

where the cells are generated in a

massive amount

is known as the upstream processing

so this upstream processes happens in

the

bioreactor whereas the recovery process

purification process

all of these comprise a downstream

processing which

includes processing purification

polishing quality control and packaging

so first of all you can take a frozen

seed stock

then you can put it in put that culture

in a

relatively bigger vessel

then this culture would be revived

after that you have to amp up the volume

so you have to gradually amp up the

volume

and once the volume reach quite a lot

then you have to put it

in a bioreactor tank so let's talk about

the bioreactor so there are lots of lots

of component in this bioreactor tank

so we are going to talk about it one by

one so let's look at the bits and pieces

okay

so first there is the tank where the

broth would stay right

and this tank is actually sterile it

ensures the microbe that you need

only is able to grow it does not ensure

growth of a random microbe obviously

there is a stirring pedal attached to a

dc motor so it would allow this pedal to

rotate

and its speed can be controlled so in

this rotation process

heat might be generated so overall in

order to maintain the temperature

of the bioreactor there would be coolant

systems as well

so there are coolant systems running

around these

broth tank after that there would be a

aeration unit

which bubbles oxygen

through this media now some

microbes might need oxygen some might

not need oxygen so this component is

variable

so this ensures the dissolved oxygen

level

in the bioreactor is in a controllable

amount

after that there would be electronic

display units and there would be control

boards and control panels there would be

inlet

and outlet chamber as well so all that

comprises bioreactor

after that the product that is generated

in the bioreactor would pass through

centrifuge and this centrifuge does not

look like the centrifuge

which is in your lab bench so these

centrifuge

are industrial scale centrifuge and they

are very different looking right

overall this is a bioreactor and this is

how the pedals are spinning

so what we learned so far is the

upstream process

the upstream process refers to

uh the massive amplification of the

microbes of the cell

and that would be generating the

substances that is our

that is of our interest in a massive

amount but it still remain

still remain in an unpurified stage

right so upstream process can be induced

upstream process can include um inoculum

development

media development improvement of the

innoculum by genetic engineering process

so all the molecular biology process at

the initial step

and ultimately optimizing the growth

kinetics

so you have to come to a growth i mean

growth condition

which was which is faster cost effective

and very efficient so all of these

things would be

important when you're making a product

in an industry skill

now coming to the downstream process so

downstream processing refers to

recovery and purification of the

biosynthetic product

which is generated in the upstream

process now it need to be purified

so the products that are generated let's

see how it is purified

now before that let us take a simple

example of two kind of scenario

one type the product could be

extracellular some kind of metabolite

which is excreted out of these bacteria

now the product would be also

intracellular for example this is the

enzyme inside the bacteria

so let's see how each type of products

can be purified and recovered

so coming to the extracellular product

purification

so in this case the bioreactor tank

would be connected to a centrifuge and

this centrifuge the output of the

centrifuge will be connected to a

filtration unit so the bio react in the

bioreactor tank there would be bacteria

and the bacteria would have several

metabolites some of these metabolites

would be of our interest and some are

useless for us so dur in the centrifuge

the first pass separation takes place

so you settle down all the big heavy

cells in the bottom

and all the metabolites including useful

and non useful metabolites are in the

solution

now when the solution is passed through

the filtration unit

you can purify your product of interest

based on

column chromatographic techniques or

many other techniques so this is how a

extracellular product can be

or extracellular metabolite can be

filtrate

now coming to a intracellular product

such as

an enzyme so let's say this enzyme is

important for your

biological research so in order to get

the enzyme you have to disrupt the cell

and you need a cell disruption system

and there are huge

industrial grade pistons which actually

breaks apart the cell

by mechanical shearing and try to get

all this product out in the solution

now once the product is out in the

solution you can so and the product is

soluble if it's an enzyme

then you can get a get rid of all of

these debris using this centrifuge

whereas my product would be in the

dissolved state right

in the supernatant so the supernatant is

collected and followed by several other

downstream processing

such as passing through a column so

this broth which has our protein of

interest has to be concentrated because

there is a huge volume in order of

thousands of liters of culture could be

inside a tank

so that is why this broth need to be

concentrated in order to recover the

product

efficiently now that is why dewatering

step could be a essential step where you

remove the excess amount of water

with the application of vacuum drying

process

now there could be initial purification

of the metabolites for example you use

hplc based columns or any kind of column

chromatographic high throughput column

chromatographic method

to purify your protein and depending

upon the nature of your

product what you want to purify you can

use your

columns such as hydrophobic interaction

column or ion exchange column any kind

of column you can use in these

hplc setups again these hplc setups

differ a lot from

that we use in our day-to-day lab work

because everything here is a industrial

skill process right so it's a macro

level process lastly there is

polishing and quality control so this

polishing polishing ensures that

this particular product would be 98 to

100 percent pure and all the purified

product should be mixed with inert

ingredients

and that ensures the product would be

purified and it would be preserved for a

long time it won't be degraded quickly

and ultimately there is a quality

control step which is very important for

this product development

and after these things these products

would be packaged nicely

and distributed where where the demand

is right

so overall in this video we learned

about the steps of

bio bio processing we looked at what is

downstream what is upstream process we

kind of had an

overview of downstream and upstream

process so in subsequent videos we would

look at all of these things in nitty

gritty details but this was just an

overview

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