Integrated Science Education: Analytical Processes - Compound Analysis

This assignment comprehensively explores the extraction, purification, and identification of carbohydrates, essential oils (specifically coconut oil), and alkaloids. It details methods for carbohydrate extraction using alcoholic solutions and clarifying agents like heavy metal salts and ion-exchange resins, alongside purification strategies like silica gel and C18 reverse phase. The identification of carbohydrates is explained using Benedict’s test. For essential oils, the assignment outlines the extraction of coconut oil, including expeller techniques and hand methods, followed by purification processes such as refined and unrefined processing, and identification based on purity, aroma, and flavor. The extraction of alkaloids is discussed using methods like Soxhlet extraction, Stas-Otto Process, and Kippenberger’s Process. The document offers a deep dive into analytical processes, making it a valuable resource for students studying integrated science education.

NAME: CHIMEZIE APRIL
MATRIC NO: F/BS/16/3800004
DEPARTMENT: INTEGRATED SCIENCE EDUCATION 4/4
COURSE CODE: ANALYTICAL PROCESSES (ISC 459)
TOPIC: Enumerate extensively on the extraction processes,
purification procedures and mode of identification of Carbohydrates,
Essential oils and Alkaloids.
INTRODUCTION
Carbohydrates are one of the most important components in many foods. Carbohydrates maybe
present as isolated molecules or they may be physically associated or chemically bound to other
molecules. Individual molecules can be classified according to the number of monomers that
they contain, such as Monosaccharides, Disaccharides, and Polysaccharides. Molecules in which
carbohydrates are covalently attached to proteins are known as GLYCOPROTEINS, whereas
those in which the carbohydrates are covalently attached to lipids are known as GLYCOLIPIDS.
Some carbohydrates are digestible by humans and therefore provide an important source of
energy, whereas others are indigestible and therefore do not provide energy. Indigestible
carbohydrates form a group of substances known as DIETARY FIBER, which also includes
lignin. Carbohydrates also contributes to the sweetness, appearance and textual characteristics of
many foods. It is important to determine the type and concentration of carbohydrates in foods for
a number of reasons.
Standards of Identity- foods must have compositions which conform to government regulations.
Nutritional Labeling- To inform consumers of the nutritional content of foods.
Detection of adulteration- each food type has a carbohydrate “fingerprint”
CLASSIFICATION OF CARBOHYDRATES
1. Monosaccharides
Monosaccharides are water-soluble crystalline compounds. They are aliphatic aldehydes or
ketones which contain one carbonyl group and one or more hydroxyl groups. Most natural
monosaccharides have either five (pentoses) or six (hexoses) carbon atoms. Commonly occurring
hexoses in foods are glucose, fructose, and galactose, whilst commonly occurring pentoses are
arabinose and xylose.

2. Disaccharides
These are relatively low molecular weight polymers of monosaccharides that are covalently
bonded. Disaccharides consist of two monomers examples include sucrose, maltose and lactose.
3. Polysaccharides
The majority of carbohydrates found in nature are present as polysaccharides. Polysaccharides
are high molecular weight polymers of monosaccharides. (e.g, starch, cellulose, and glycogen
which are formed from only glucose).
EXTRACTION OF CARBOHYDRATES
One of the most commonly used methods of extracting low molecular weight carbohydrates
from foods is to boil a defatted sample with an 80% alcohol solution. Monosaccharides and
Disaccharides are soluble in alcoholic soutions, whereas, proteins, polysaccharides and dietary
fibers are insoluble. The soluble components can be separated from the insoluble components by
filtering the boiled solution and collecting the filtrate (the part which passes through the filter)
and the retentante (the part retained in the filter). These two fractions can then be dried and
weighed to determine their concentration. In addition to monosaccharides and disaccharides,
various other small molecules may also be present in the alcoholic extract that could interfere
with the subsequent analysis e.g., amino acids, organic acids, vitamins, pigments, minerals etc. it
is usually necessary to remove these components prior to carrying out a carbohydrate analysis.
This commonly achieved by treating the solution with clarifying agents or by passing it through
one or more ion-exchange resins.
Clarifying agents:- water extracts of many foods contain substances that are colored and thus
interfere with spectroscopic analysis or endpoint determinations. For this reason solutions are
usually clarified prior to analysis. The most commonly used clarifying agents are heavy metal
salts (such as lead acetate) which form insoluble complexes with interfering substances that can
be removed by filtration or centrifugation. However, it is important that the clarifying agent does
not precipitate any of the carbohydrates from solution as this could cause an underestimation of
the carbohydrate content.
Ion- exchange;- many monosaccharides and disaccharides are non-charged molecules and can
therefore be separated from charged molecules by passing samples through ion-exchange
columns. By using a combination of a positively and a negatively charged column, it is possible
to remove most charged contaminants. Non polar molecules can be removed by passing a
solution through a column with a non-polar stationary phase.

PURIFICATION OF CARBOHYDRATES
Carbohydrate compounds are purified in similar manner as other small molecules. They are often
reacted with protecting groups that greatly modify their polar character and allow purification
with silica gel.
Carbohydrates are UV transparent, but groups attached to the carbohydrate generally absorbs UV
light allowing the use of UV detectors or All-Wave-Length Collection (AWC). The AWC
monitors the absorbance across a user-specified wavelength range. The program applies a signal
processing algorithm that suppresses interference from solvent absorbance that causes drifting
baselines that can “hide” peaks or interfere with fraction collection. Detecting all wavelengths
ensures that no compound absorbing light in a single wavelength range is missed during
collection. All-Wavelength detection converts the absorbance across a rang e of wavelengths into
a single signal for the fraction collector while suppressing absorbance changes due to the eluting
solvent. Purification strategies of carbohydrates include Silica gel strategies, C18 Reverse Phase
strategies, and ion pairing.
IDENTIFICATION OF CARBOHYDRATES
A Benedict’s test can be used to identify reducing sugars. The Benedict’s solution (a chemical
reagent) contains a blue soluble form of copper ions (Cu++) that can undergo reduction (that is,
the copper ions gain electrons) when heated in the presence of a reducing sugar. When the blue
copper ions are reduced, they change from the soluble blue color to reddish color copper ions
(Cu+) that are insoluble. The color of the test solution changes from
BlueGreenOrangeRed-brown or Rust color.
As more reduced copper ions are formed, you will use the Benedict’s solution to test for
presence of reducing sugar in various substances.
INTRODUCTION (Essential Oils- Coconut oil)
Coconut is high in fat, with a fine, edible oil that is used in cooking, cosmetics and other
products. There are several methods of extracting coconut oil from the meat of the nut, known as
COPRA. One method of extracting coconut oil is the expeller technique, in which dried copra
passes through a special screw press that compresses the coconut copra, squeezing about 75
percent of the oil from the coconut meat. Small, hand-operated or motor-driven presses are
relatively affordable and available online. For smaller amounts, a hand method using cheese
cloth will also work.
EXTRACTION OF COCONUT OIL
To prepare for extraction, first crack open fresh coconuts and remove the copra by carefully
prying the meat away from the shell with a knife. Cut the copra into ¼ inch pieces. There is no

need to remove the brown outer lining, called the Testa, on the coconut meat, allow the copra to
dry thoroughly to remove all moisture from the meat.
Press Preparation
Before you extract any coconut oil, clean the oil press thoroughly to ensure there is no residue
remaining from the previous pressing. Assemble the press according to manufacturer’s
specifications. Install a plastic container under the press to catch the extracted oil. Place another
container at the end of the press to catch the spent coconut meat.
Press Operation
Now that the press is clean and ready to operate, insert the dried copra into the hopper in the top
of the oil press. Turn the press handle or start the motor that runs the press. As the impeller
within the press turns, it will compress the copra to extract the oil. Push the copra into the press
with a wooden plunger. Check the hopper regularly and add more copra as needed to keep the
press full. Examine the containers: when full, replace them and continue to extract the oil until
all of the copra is pressed.
Storage
Place the lid on the newly extracted coconut oil and put it in a warm place to settle for 24 hours.
Once the oil has been separated from any solids, pour the oil off into a bottle for storage and
place a cap on it tightly to seal it. Keep the oil away from light and in a cool place to prevent it
from becoming rancid.
PURIFICATION
 Refined Processing
Whether coconut oil becomes refined or unrefined depends on what processing method the oil
undergoes. Refined coconut oil starts out as dried coconut meat. Because coconut oil producers
frequently dry the meat in open air, it can gather hazardous bacteria and pathogens- so any oil
extracted from copra needs further purification before it is safe to consume. During the
refinement process, copra undergoes bleach filtration to screen out impurities, as well as heat
treatment to remove its distinctive odor.
 Unrefined Processing
Unrefined coconut oil, on the other hand, starts with fresh coconut meat rather than dried copra-
ensuring the extracted oil is sanitary without the need for further purification. To produce
unrefined coconut oil, the meat goes through one of two methods: “quick drying” or “wet
milling”. Quick-dried coconut meat receives a small amount of heat to remove moisture, and
then mechanical extraction separates the unrefined oil from the meat. With the wet-milling,
machines press the liquid out of fresh coconut meat-and then use boiling, refrigeration,

centrifugation, fermentation or enzymes to isolate the oil from the extracted liquid. With both the
quick drying and wet milling methods, the resulting coconut oil is safe for consumption without
bleaching or deodorization.
IDENTIFICATION
The first thing to look for is a Virgin coconut oil. Virgin coconut oil is recommended over non-
virgin or RBD oil. RBD stands for refined, bleached, and deodorized. Processing can remove and
destroy nutrients.
In RBD oil, all the phytonutrients have been removed so it is tasteless and odorless. Virgin
coconut oil is generally superior to RBD oil, but not all Virgin oils are of equal quality.
The second thing you look for in high quality coconut oil is purity. Pure coconut oil naturally has
high melting point. At 24 degrees C and lower it becomes solid, at higher temperatures it turns
liquid.
High quality virgin coconut oil should be snow- white in color when it is solid and water clear
when liquid. Discoloration is a sign of contamination or excessive heating during processing.
The third thing to look for is aroma and flavor. Virgin coconut oils should have a mild coconut
smell and taste. If not, they have been highly refined. If they have no flavor, they are essentially
RBD oil, even if they did come from fresh coconut. Some virgin coconut oils have a very strong
flavor or smell. This usually indicates a poor quality oil because smell and taste comes mainly
from contaminates and not coconut. Many virgin coconut oils use some type of heat in
processing, often smoke from thee heating process contaminates the oil giving it a roasted or
smoky smell and taste. Some of these contaminated oils smell like roasted coconut and taste like
ash. The very best virgin coconut oils do not have strong roasted or smoky taste or smell. They
should have a very mild coconut aroma and flavor. The flavor should not be overpowering, but
just mild enough to enjoy without overpowering the flavor of the foods its used with.
INTRODUCTION (ALKALOIDS)
Alkaloids are a group of naturally occurring chemical compounds that mostly contain basic
nitrogen atoms. This group also includes some related compounds with neutral and even weakly
acidic properties. Some synthetic compounds of similar structures are also termed Alkaloids. In
addition to carbon, hydrogen, and nitrogen, alkaloids may also contain oxygen, sulfur, and more
rarely other elements such as, chlorine, bromine and phosphorus.
EXTRACTION OF ALKALOIDS
Because of structural diversity of alkaloids, there is no single method of its extraction from
natural raw materials.

Alkaloids are separated from their mixture using different solubility in certain solvents and
different reactivity with certain reagents or by distillation. In general, the alkaloids may be
extracted by any of the following three well-defined and widely accepted processes, namely;
a. Soxhlet Extraction Process
b. Stas-Otto Process, and
c. Kippenberger’s Process.
Soxhlet Extraction Process- The Soxhlet assembly is a continuous extractor which is generally
suitable for the extraction of alkaloids from powdered plant materials with the help of organic
solvents. In this instance, the powdered drug is usually moistened with dilute ammonia solution
and then packed loosely in the thimble of the Soxhlet apparatus; and the organic solvent affords a
deep penetration of the moist drug thereby allowing the greatest possible extraction of the
alkaloids from the exposed surfaces of the cells and tissues of the crude drug. Once the
extraction is ascertained to have been completed, the solvent is filtered and evaporated in a
Rotary Thin-Film Evaporator and the residue is treated further for the isolation of individual
alkaloids.
Stas-Otto Process- The Stas-Otto process essentially consists of treating the powdered and
sieved drug substance with 90-95% (v/v) ethanol, previously acidified with tartaric acid. The
proportion of crude drug to solvent should be maintained as 1kg to 1L. the alcohol is distilled off
under vacuum and the resulting aqueous residue is treated with petroleum ether (60-80C) to
remove the fatty components completely. If any alkaloid is removed by the petroleum ether, it
must be recovered by treating it with dilute mineral acid. Thus, the resulting aqueous extract is
filtered and evaporated to dryness preferably in a Rotary Thin-Film Evaporator under vacuum.
The residue is extracted with absolute ethanol thereby dissolving the total alkaloids.
Kippenberger’s Process- In Kippenberger’s process the powdered and sieved plant substance is
first and foremost digested with solution of tannin(100g) in glycerol (500g) at a constant
temperature of 40C for a duration of 48 hours. The resulting mixture is further heated to 50C so
as to help in the complete coagulation of proteinous substances, cooled to ambient temperature
and finally filtered. The resulting filtrate is thoroughly shaken with petroleum ether to get rid of
faulty materials (oils, fats and waxes), and the last traces of petroleum ether is removed from the
extract by heating either or on a water-bath (electric) or exposure to infrared lamp. The fat-free
crude plant extract is subsequently acidified and shaken with chlorofoam, successively to remove
the bulk of the alkaloids, namely, atropine, codeine, colchicine, narcotine, nicotine, papaverine,
spartenine, and thebaine.
The resulting residual extract may still contain narceine, curarne, and morphine. Howerer,
narceine and morphine may be isolated by passing freshly generated CO2 directly into extract so
as to convert the alkali hydroxide into their corresponding carbonate, which is then ultimately
subjected to solvent extraction using a mixture of alcohol and chlorofoam. Finally, the third

alkaloid, curarine, may be extracted by agitation with a mixture of equal volumes of ether and
chlorofoam.
PURIFICATION OF ALKALOIDAL EXTRACT
Purification of alkaloids can be done by the combination of the following methods;
a) Extraction with Acid Solution- The extraction of the alkaloid from the bulk of the
crude alkaloid solution in immiscible organic solvent is invariably carried out by
shaking with an acid solution. In usual practice, the use of HCL is restricted when
chlorofoam remains as the solvent because of the fact that quite a few alkaloidal
hydrochlorides are distinctly soluble in the latter. However, dilute H2SO4 is always
preferred over HCL for general use in the extraction of alkaloids. Subsequently, the
acid solution is rendered alkaline with dilute NH4OH solution to liberate the alkaloids
which is then extracted with an organic solvent. The solvent is removed under reduced
pressure and the traces of moisture is removed with anhydrous sodium sulphate.
b) Precipitation of Alkaloid with Precipitating Reagent – The usual precipitation of the
alkaloid as a complex compound is accomplished by the addition of a suitable
precipitation reagent. The resulting alkaloidal complex is further purified by filtration,
recrystallization and ultimately decomposed to obtain the desired free alkaloid(s).
c) The purification of alkaloids may also be accomplished by the formation of its
crystallized alkaloidal salt by the addition of an appropriate mineral or organic acud,
such as: hydrochloric, hydrobromic, perchloric, sulphuric, oxalic and tartaric acids.
IDENTIFICATION OF ALKALOIDS
The qualitative chemical tests used for detection of alkaloids are dependent on the character of
alkaloids to give precipitate as salts of organic acids or with compound of heavy metals like Hg,
Au, Pt, etc.
Test by Dragendorff reagent (Potassium-bismuth-iodide solution):- Alkaloids give reddish-
brown precipitate with this reagent.
Test by Mayer reagent (Potassium-mercuric-iodide solution):- Alkaloids gives cream color
precipitate with this reagent.
Test by Wagner reagent (Iodine-potassium-iodide solution):- Alkaloids give brown color
precipitate with this reagent.
Test by Tannic acid:- Alkaloids gives buff color precipitate with this acid.