Epoxy resins: A complete guide to their uses, benefits, and applications

How to extract allicin from garlic ????

Allicin is an organosulfur compound derived from garlic (Allium sativum). It is formed when garlic cells are damaged, for example by crushing or chopping, activating the enzyme alliinase which converts alliin, a stable precursor, to allicin. The chemical formula for allicin is C6H10OS2, and it is responsible for the characteristic odor of fresh garlic as well as many of its beneficial health effects.

Properties of allicin

• Antimicrobial: Allicin is known for its powerful antimicrobial properties, capable of killing or inhibiting the growth of a wide range of bacteria, viruses, fungi and parasites.
• Antioxidant: It also has antioxidant properties, helping to protect cells against damage caused by free radicals.
• Anti-inflammatory: Allicin can reduce inflammation and modulate immune responses.
• Cardioprotective: It can help reduce blood pressure, cholesterol and prevent platelet aggregation, thus contributing to cardiovascular health.

Uses of allicin

Medical and therapeutic

• Treatment of infections: Used to treat various bacterial, fungal and viral infections, especially when the pathogens are resistant to traditional antibiotics.
• Dietary Supplements: Marketed as capsules, tablets or powders, allicin is taken as a dietary supplement for its general health benefits, including immune support and reduction of inflammation.
• Cardiovascular health: Used to lower blood pressure, reduce LDL cholesterol and increase HDL cholesterol, and prevent atherosclerosis.

Food industry

• Natural preservative: Thanks to its antimicrobial properties, allicin is used as a natural preservative to extend the life of foods.
• Aroma and flavor: Used as a natural flavor to give food products the characteristic flavor of garlic.

Cosmetics

Skincare: Incorporated into skincare products for its antibacterial and antioxidant properties, helping to treat conditions like acne and protect the skin from oxidative damage.

Source of allicin

Allicin is a bioactive organosulfur compound that mainly comes from garlic (Allium sativum). It is not present in free form in intact garlic, but is produced when it is damaged, for example by crushing, chopping or chewing.

Formation of allicin

• Alliin: Garlic contains a stable precursor called alliin. Alliin is a derivative of the amino acid cysteine and is stored in garlic cells.
• Alliinase: Garlic also contains an enzyme called alliinase, which is compartmentalized separately from alliin in intact cells.
• Enzymatic reaction: When garlic cells are damaged, alliin comes into contact with alliinase. Alliinase catalyzes the conversion of alliin to allicin through a rapid chemical reaction.

Alliin + Alliinase → Allicin

Chemical structure of allicin

• Chemical formula: C6H10OS2
• IUPAC name: (E)-2-propene-1-sulfinothioate of prop-2-ene

Characteristics of allicin

• Odor: Allicin is responsible for the pungent and characteristic odor of fresh garlic.
• Instability: Allicin is relatively unstable and can break down into other sulfur compounds over time or during cooking.

Allicin Extraction Methods

Extraction of allicin with supercritical carbon dioxide (CO2)

Preparing the garlic

Peel and clean the garlic cloves

• Start by removing the outer skins from the fresh garlic cloves. This can be done manually or using a mechanical peeling device for large quantities.
• Wash the pods thoroughly to remove any dirt or impurities. This step ensures that only pure garlic components are used for extraction.

Grind the garlic cloves into a fine paste to increase the contact surface

• Use a mechanical crusher or blender to grind the garlic cloves into a smooth paste. This operation increases the contact surface, thus facilitating the extraction of allicin.
• Reducing garlic cloves into a paste allows for more efficient conversion of alliin to allicin via the enzyme alliinase, which is essential for maximizing allicin yield (Nguyen et al., 2021).

Loading the extractor

Loading the garlic

Once the garlic paste is prepared, it must be loaded into the supercritical extractor. This step is crucial to ensure uniform distribution of garlic material and maximize allicin extraction.

 Preparation of the extractor

Before loading the garlic paste, the extractor should be cleaned and checked to ensure that it is ready for operation. This includes checking seals, valves and connections to prevent leaks.

 Loading garlic paste

• The crushed garlic paste is placed in the extraction container. The container must be filled evenly to avoid air pockets and ensure optimal contact between the supercritical CO2 and the garlic paste.
• Homogeneous filling helps avoid agglomeration phenomena, which can cause blockages and reduce extraction efficiency (Valle, Glatzel, & Martínez, 2012).

 Closing and securing the container

• After loading, the extraction container is closed tightly. All connections must be secure to withstand the high pressures of supercritical extraction.
• It is important to check that the container is properly sealed to avoid any CO2 leaks during the extraction process.

 Preparation for extraction

The container containing the garlic paste is now ready for extraction with supercritical CO2. The system is configured to achieve optimal extraction conditions (temperature, pressure and CO2 flow) (He, 2013).

Introduction of CO2

CO2 supply system

 Connect the CO2 cylinder to the extractor

• The CO2 cylinder, containing pure carbon dioxide under pressure, is connected to the supercritical extractor via high pressure pipes. These connections should be checked to prevent gas leaks.
• The use of robust and secure connections is essential to maintain the security of the extraction process (Ding, 2009).

 Adjust the temperature and pressure of the extractor to reach supercritical conditions

• Adjust the temperature of the extractor to approximately 35°C to reach the supercritical state of CO2. This temperature allows the CO2 to have both gas and liquid properties, thus facilitating the extraction of allicin.
• Adjust the extractor pressure to approximately 20 MPa (200 bar). This pressure is crucial to maintain the CO2 in a supercritical state, optimizing its extraction capacity (He et al., 2007).
• The combination of temperature and pressure must be carefully controlled and monitored throughout the process to ensure efficient and stable extraction (Sapkale et al., 2010).
• The correct introduction of CO2 is a crucial step to ensure the efficiency and safety of supercritical allicin extraction.

Static extraction of allicin

Maintain supercritical conditions

• Set and maintain the extractor temperature at 35°C and the pressure at 20 MPa. These conditions are optimal for CO2 to reach the supercritical state, where it has both liquid and gaseous properties, allowing better dissolution of allicin.
• Accurately maintaining these parameters is crucial to ensure efficient and reproducible extraction (He, 2013).

Allow supercritical CO2 to saturate the garlic paste for 75 minutes without circulation

• Once supercritical conditions are reached, allow the CO2 to saturate the garlic paste statically. This means that the CO2 is maintained in the extractor without being in active circulation, allowing optimal interaction between the supercritical solvent and the garlic matrix.
• This static saturation step is crucial to allow the CO2 to fully penetrate the garlic paste and effectively dissolve the allicin. A duration of 75 minutes was determined as optimal to maximize extraction yield (Cai, 2007).

Dynamic extraction of allicin

Circulate the supercritical CO2 through the garlic paste continuously

• Once the static extraction phase is complete, the supercritical CO2 is circulated continuously through the garlic paste. This step ensures that the supercritical solvent constantly passes through the garlic material, dissolving and transporting the dissolved allicin to the collector.
• Supercritical CO2 in continuous circulation makes it possible to efficiently extract soluble compounds by maintaining them in the solvent flow (He, 2013).

Collect the CO2 enriched with allicin in the separator

• The allicin-enriched CO2 is directed to a separator where it is depressurized back to the gaseous state, leaving the allicin in collected solution.
• The depressurization process makes it possible to efficiently separate the allicin from the CO2, with optimal extraction yield. This separation is essential to recover allicin in pure form (Cai, 2007).

Dynamic extraction optimizes allicin extraction by ensuring continuous circulation of supercritical CO2 through the garlic paste, enabling efficient allicin collection.

Separation of allicin

Depressurize the CO2 to return it to the gaseous state

• After dynamic extraction, the supercritical CO2 enriched in allicin is directed to a separator.
• In the separator, the CO2 is depressurized, causing it to return to a gaseous state. This change of state makes it possible to separate the allicin dissolved in the CO2.
• Precise control of pressure and temperature is essential to ensure effective separation. Controlled depressurization avoids loss of volatile compounds and ensures maximum allicin recovery (He, 2013).

Separate the allicin which is collected in the separator

• Once the CO2 has become gaseous again, the allicin remains in liquid form in the separator. This liquid is then collected and can be purified if necessary.
• The separation process may include additional steps like filtration or distillation to obtain high purity allicin (Cai, 2007).
• Modern separation systems make it possible to recover a large part of the allicin with high efficiency, often above 90%, depending on the specific extraction conditions and the quality of the equipment used (Rybak et al., 2004 ).

Separation is a crucial step in the supercritical extraction process, allowing allicin to be recovered efficiently and purely after its initial extraction with supercritical CO2.

Recovery of allicin

Collect the separated pure allicin

• After depressurization of the CO2 in the separator, the allicin remains in liquid form and is collected in a suitable container.
• This collection must be done immediately after depressurization to avoid any loss or degradation of the allicin. The recovery efficiency of allicin can reach up to 96% under optimal supercritical CO2 conditions (Rybak et al., 2004).

Store allicin under appropriate conditions to maintain its stability

o Allicin is a relatively unstable compound, sensitive to temperature, light and oxygen. To maintain its stability, it is crucial to store it under controlled conditions.

o Storage conditions for allicin:

• Allicin should be stored at a low temperature, ideally between 4°C and -20°C, to slow down degradation reactions.
• Use opaque or amber containers to protect allicin from exposure to light, which can accelerate its degradation.
• Store allicin under an inert atmosphere (e.g., under nitrogen) to minimize exposure to oxygen and prevent oxidation (Tao, 2009).

Main solvents and co-solvents used for allicin extraction

Supercritical CO2 (CO2-SC) extraction is primarily valued for its ability to operate without toxic organic solvents. However, in some cases, co-solvents can be used to improve extraction efficiency, especially for difficult-to-extract compounds. The main solvents and co-solvents used for allicin extraction are:

 Carbon dioxide (CO2)

• Description: CO2 is the main solvent used in this method. In the supercritical state, it has unique properties that allow it to penetrate solid matrices and effectively dissolve various compounds.
• Advantages: Non-toxic, non-flammable, easily available and inexpensive. Supercritical CO2 is also considered a "green" solvent due to its low environmental impact (Beckman, 2012).

 Ethanol

• Description: Often used as a co-solvent to improve the solubility of polar compounds in supercritical CO2.
• Usage: Added in small proportions (usually between 5-20%) to facilitate the extraction of specific compounds, such as essential oils and phenolic compounds (Paes et al., 2014).

 Methanol

• Description: Used as a co-solvent to extract bioactive compounds that are difficult to solubilize with CO2 alone.
• Use: Particularly effective for the extraction of lipids and other organic compounds (Ambrosino et al., 2004).

Why use co-solvents?

• Improved solubility: Some compounds, particularly polar compounds or those with high molecular weights, are not readily soluble in supercritical CO2 alone. Co-solvents like ethanol and methanol increase the solubilization capacity of supercritical CO2.
• Extraction efficiency: Adding co-solvents can increase extraction yield and reduce the time required to extract a given amount of bioactive compounds.
• Process flexibility: Co-solvents allow the properties of the supercritical solvent to be adjusted to target specific compounds, providing greater flexibility in industrial and research applications.

Although supercritical CO2 extraction can do without organic solvents, the use of co-solvents such as ethanol and methanol can significantly improve the extraction efficiency for certain bioactive compounds.

Safety and precautions

 Use appropriate personal protective equipment (PPE)

Operators should wear personal protective equipment including chemical-resistant gloves, safety glasses and laboratory clothing to avoid direct contact with supercritical CO2. This equipment protects against the risk of frostbite caused by the rapid expansion of CO2 under pressure (Clavier & Perrut, 1996).

 Ensure adequate ventilation in the work area

CO2, although non-toxic, can cause asphyxiation if accumulated in confined spaces. It is essential to ensure that the work area is well ventilated to prevent the accumulation of CO2. Effective ventilation systems and CO2 detection alarms should be installed to alert of high gas levels (Soares & Coelho, 2012).

 Follow safety protocols for handling gases under high pressure

Working with supercritical CO2 involves the use of high pressures, which poses risks of pressure vessel rupture. It is crucial to follow strict safety protocols, including regular inspections of equipment, the use of safety valves, and training operators on emergency procedures in the event of a leak or explosion (Eggers & Green, 1990).

Compliance with these safety measures ensures safe and efficient handling of supercritical CO2, minimizing risks to operators and the working environment.

Method for extracting allicin by enzymatic hydrolysis

Enzymatic hydrolysis is a biochemical process where enzymes catalyze the breakdown of complex molecules into simpler components through the addition of water. In the context of allicin extraction, enzymatic hydrolysis involves the use of the enzyme alliinase, which converts alliin, a stable precursor found in garlic, to allicin, the active compound.

Enzyme used: Alliinase

• Alliinase occurs naturally in garlic and is activated when garlic cells are crushed or damaged.
• This enzyme catalyzes the conversion of alliin to allicin.

Optimal conditions for hydrolysis

• pH: 6.0
• Temperature: 30°C

These conditions are chosen to maximize alliinase activity, ensuring efficient conversion of alliin to allicin.

Extraction with 95% ethanol

Solvent extraction is a technique used to isolate specific compounds from a material. 95% ethanol is an effective polar solvent for extracting bioactive compounds such as allicin. 95% ethanol is used to dissolve the allicin released during enzymatic hydrolysis. It is an excellent solvent for allicin due to its polarity and ability to penetrate plant matrices.

 Optimal conditions for the extraction of allicin

• Material/ethanol ratio: 1:6.4:

This ratio means that for every gram of garlic paste, 6.4 milliliters of ethanol are used, ensuring complete dissolution of the allicin.

•  Extraction time: 60 minutes

This duration allows sufficient contact between the ethanol and the garlic paste, ensuring maximum extraction of allicin.

• Temperature: 30°C

Maintaining this temperature helps preserve the stability of the allicin and improve extraction efficiency.

 Extraction Yield

The extraction yield of 2.81‰ means that for every kilogram of raw material (garlic paste) processed, approximately 2.81 grams of pure allicin are extracted. This yield is considered effective for extraction processes of bioactive compounds.

Ultrasound-assisted allicin extraction method

Optimal conditions for the extraction of allicin

• Temperature: 35°C
• Ultrasonic power: 48 W
• Duration: 32 minutes
• Extraction yield: 2.897 mg/g

Details of the ultrasound-assisted allicin extraction method

Preparation of garlic

• Peel the fresh garlic cloves and clean them carefully.
• Grind the garlic cloves into a fine paste to increase the contact surface.

Setting up the ultrasonic extraction system

• Equipment: Ultrasound-assisted extraction device

• Settings:

• Set the temperature to 35°C.
• Set the ultrasonic power to 48 W.

Allicin extraction procedure

 Mixing the sample

• Place the garlic paste in a container suitable for ultrasonic extraction.
• Add 95% ethanol respecting a material/solvent ratio of 1:6.4 (for example, 1 g of garlic paste for 6.4 ml of ethanol).

 Ultrasonic extraction

• Subject the mixture to ultrasonic power of 48 W at 35°C for 32 minutes.
• Ultrasonic agitation creates cavitations in the solvent, thereby increasing the efficiency of extraction of bioactive compounds like allicin.

Separation and recovery of allicin

 Allicin recovery procedure

• Filter or centrifuge the mixture to separate the dissolved allicin from the garlic paste.
• Evaporate the ethanol under vacuum or by distillation to concentrate the allicin extract.

 Result of allicin extraction

Under these optimal conditions, the maximum extraction yield achieved is 2.897 mg of allicin per gram of raw material (Li, 2013).

Vacuum microwave allicin extraction method

The optimal conditions for the extraction of allicin by microwave under vacuum are:

• Temperature: 35°C
• Microwave power: 48 W
• Duration: 32 minutes

The yield of allicin extraction by microwave under vacuum can reach 2.897 mg/g

Details of the allicin extraction method by microwave under vacuum

Preparation of garlic

• Peel the fresh garlic cloves and clean them carefully.
• Grind the garlic cloves into a fine paste to increase the contact surface.

Setting up the vacuum microwave extraction system

• Set the temperature of the microwave vacuum extraction device to 35°C.
• Set the power of the microwave vacuum extraction device to 48 W.

Allicin extraction procedure

 Mixing the sample

• Place the garlic paste in a container suitable for vacuum microwave extraction.
• Add 95% ethanol respecting a material/solvent ratio of 1:6.4 (for example, 1 g of garlic paste for 6.4 ml of ethanol).

 Vacuum microwave extraction

• Subject the mixture to microwave power of 48 W at 35°C for 32 minutes.
• The vacuum environment reduces the boiling temperature of solvents, thereby improving the extraction efficiency of bioactive compounds like allicin.

Separation and recovery of allicin

• Filter or centrifuge the mixture to separate the dissolved allicin from the garlic paste.
• Evaporate the ethanol under vacuum or by distillation to concentrate the allicin extract.

Under these optimal conditions, the maximum extraction yield achieved is 2.897 mg of allicin per gram of raw material (Li, 2013).

Extraction of allicin by ultrasound-assisted two-phase aqueous system

The optimal conditions for extraction of allicin by ultrasound-assisted two-phase aqueous system are:

• Mass fraction of PEG: 19.4%
• pH: 3.0
• Ultrasonic treatment time: 55 minutes
• Extraction efficiency: 88.89%

Preparation of garlic

• Peel the fresh garlic cloves and clean them carefully.
• Grind the garlic cloves into a fine paste to increase the contact surface.

Setting up the two-phase aqueous system

• Use polyethylene glycol (PEG) with a mass fraction of 19.4%.
• Adjust the pH of the solution to 3.0 using appropriate acid (e.g. citric acid).

Ultrasound-assisted extraction procedure

 Mixing the sample

• Place the garlic paste in a container containing the two-phase aqueous system (PEG and aqueous phase).
• Add 95% ethanol respecting a material/solvent ratio of 1:6.4 (for example, 1 g of garlic paste for 6.4 ml of ethanol).

 Ultrasonic extraction

• Subject the mixture to ultrasonic treatment with a power of 48 W at 35°C for 55 minutes.
• Ultrasonic agitation creates cavitations in the solvent, thereby increasing the efficiency of extraction of bioactive compounds like allicin.

Separation and recovery of allicin

• Filter or centrifuge the mixture to separate the dissolved allicin from the garlic paste.
• Evaporate the ethanol under vacuum or by distillation to concentrate the allicin extract.

Under these optimal conditions, the maximum extraction efficiency achieved is 88.89%, which means that the majority of the allicin present in the raw material is recovered (Wang, 2014).

This ultrasound-assisted two-phase aqueous system extraction method is efficient, providing rapid extraction and high recovery of allicin under optimized conditions.

Allicin extraction can be carried out efficiently by several methods, each having specific optimal conditions. The most appropriate method depends on available resources and the specific requirements of the application.

Chemical synthesis of allicin

The chemical synthesis of allicin is an effective alternative to its natural extraction. Here is an example of a detailed chemical procedure for the synthesis of allicin:

Necessary reagents

• Alliin or a similar precursor compound: Alliin is a natural precursor of allicin found in garlic.
• Mild oxidant: Hydrogen peroxide (H22O22) is commonly used as an oxidant.
• Suitable solvent: Ethanol or water can be used as a solvent to dissolve the alliin and facilitate the reaction.

Procedure

 Dissolution of alliin

Dissolve the alliin in a suitable solvent such as ethanol or water. This step prepares the substrate for the oxidation reaction.

 Addition of the oxidant

Slowly add the hydrogen peroxide (H22O22) with constant stirring. Slow addition and stirring helps control the reaction and avoid unwanted side reactions.

 Temperature control

Maintain the reaction at a controlled temperature, generally around 0-5°C. This prevents the breakdown of allicin, which is a relatively unstable compound.

 Separation and purification of allicin

Once the reaction is complete, extract the allicin from the reaction mixture using an appropriate separation technique, such as liquid-liquid extraction or chromatography. This step makes it possible to obtain pure allicin for subsequent applications.

Efficacy and purity

• Yield: The chemical method makes it possible to obtain allicin with a high yield and a purity of more than 98%.
• Analysis: The identity and purity of allicin can be verified by analytical techniques such as high-performance liquid chromatography (HPLC) and nuclear magnetic resonance (NMR) spectroscopy (Albrecht et al., 2017 ).
• The chemical synthesis of allicin provides a convenient and efficient method to produce this important bioactive compound, essential for various medical and industrial applications.

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