LABORATORY TECHNIQUES AND METHODS

Aside from doing the Scientific Method as a general method of conducting any Science-
related activities, laboratory methods and techniques are essentially introduced since
experiments are mostly done in laboratories. Cambridge dictionary defines a laboratory as “a
room or building with scientific equipment for doing scientific tests for teaching Science,
or a place where chemicals or medicines are produced.” On the other hand, laboratory
techniques are procedures and practices that are required in utilizing various equipment
inside the laboratory.
Here are some of the laboratory techniques and methods that you need to learn as you
conduct your research study:

A. Microbiological Techniques
In microbiology, microorganisms like archaea, bacteria, fungi, protozoa, viruses, etc. are
being studied.

The common microbiological techniques are discussed in detail below:

1) Sterilization
Microbiological media, a medium for growing bacteria or any other microorganisms, must
be sterilized by heating it in an autoclave (like a pressure cooker) at 121°C for 15 minutes.
This will kill all living organisms, i.e., spores.

2) Aseptic Techniques
This is done to lessen the chance of bacterial contamination. Furthermore, this also
involves disinfecting working area 4exposed to microbiological media to minimize the
possible contact by bacteria from the air and use flames to kill bacteria that might enter the
vessels after opening them.

3) Inoculation
This is the process by which bacteria are introduced to a media in various means. For
example (bacteria), from a drop in a heat-sterilized loop are dispersed on the surface of the
agar. This is the same with broth cultures. For bacteria in a liquid, a sterile pipette is
introduced to a Petri dish before the agar is poured on the top (“pour plates”).

4) Incubation

This is the process of maintaining favorable conditions for growth and development, i.e.,
microbiological culture.

The Petri dishes that contain the agar or tubes contain broth are incubated, for example,
a special apparatus is placed in a room with a fixed temperature (usually at 37 °C, for
pathogens, while for bacteria is 35 °C from the environment). In school, incubation
temperatures are set lower to minimize the growth of potential pathogens.

B. Chemical

1) Spectrophotometry
This is a method by which you measure the amount of absorption and transmittance of
light. This is used widely for qualitative analysis in various fields of Sciences (e.g.,
biochemistry, biology, chemistry, material and chemical engineering, clinical applications,
 industrial applications, etc. ). A spectrophotometer is a device that measures the number of
photons (the intensity of light) passing through a sample solution. This consists of two
devices: a spectrometer and a photometer. A spectrometer is a device that disperses and
measures light. On the other hand, a photometer consists of a photoelectric detector that
measures the intensity of light.

2) Extraction

This is the conversion of compound/s from a solid or liquid state into another phase or
state.

Preparation of Crude Extracts from Plants

Studies about the medicinal properties of plants require the extraction of crude
compounds from plant parts such as leaves, flowers, and roots. Knowledge on how to do
the extraction process is necessary.

Step 1: Air Drying of Samples. After collecting the adequate amount of plant samples,
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wash with clean water and dry at room temperature. The efficiency of extracting crude
compounds can be achieved when the plant samples are airdried.

Step 2: Grinding the Dried Samples into Powder. The purpose of this step is to turn
samples into small pieces (powder) using a Wiley Mill or if you do not have one, you can
use a pair of scissors. Compounds can easily dissolve in the solvent when the plant sample
is powdered.

Step 3: Soaking the Ground Plant Samples in a Solvent. Soaking the ground samples in
a specific solvent will make the compounds go to the solvent or will dissolve in the solvent.
This is already the start of the extraction process. The kind of solvent you want to use
depends on your study particularly on the specific compound you want to extract from the
plants. Most commonly used are ethanol, acetone, petroleum ether, and hexane.

Step 4: Collecting the filtrate. The dissolved compounds from the plant samples can be
found in the filtrate, the liquid mixture. Filter the mixture using a filter paper (usually
Whatman #1 filter paper or as required by the procedure).

Step 5: Evaporating the solvent. This is the last step in preparing the crude compounds
of crude extract from the plant sample. You can evaporate the solvent using a Rotatory
Evaporator (Rotavap) or you can simplify by evaporating the solvent by heating using an
alcohol lamp. The crude extract should be in semi-solid form.

Step 6: Recover the extract from the rotary evaporator container. The extract should be
semi-solid or sticky in appearance to ensure that the solvent ethanol or other solvent
chemicals are thoroughly removed.

3) Titration
This is the process of determining the concentration of a solution. Medwick &
Kirschner (2010) defined titration as “a common laboratory method of quantitative
chemical analysis that used to determine the unknown concentration of an identified
analyte.” The common types of titration are acid-base titration and redox titration. An
acid-base titration depends on the neutralization between an acid and a base when mixed
in a solution. When there is a change in color in the acid-base indicator signals the endpoint
of the titration.
 C. Physical

1) Aeration
This is the interchange of various gases (CO 2 & O2) between the atmosphere and soil and
the various reactions that either consume or produce gases in the soil.

For example, the oxygen gas (O 2) moves from the atmosphere to soil and is consumed by
plant roots and microorganisms. While carbon dioxide (CO 2) moves from soil, where it is
produced by the plant and microbial respiration, to the atmosphere.

2) Centrifugation
This is the process of separating substances applying centrifugal force through the use of
a centrifuge. A centrifuge is a device used to separate particles or macromolecules (i.e.,
cells, nucleic acids, proteins, and sub-cellular components).

3) Chromatography
The word chromatography means “color writing”, which chemists used to test liquid
mixtures. This is the process of separating organic and inorganic compounds used by
scientists to analyze and study these compounds.

There are four main types of chromatography: Gas Chromatography, Liquid

Chromatography, Paper Chromatography, and Thin-layer Chromatography.

The table below presents a detailed comparison of these types.

Types of Chromatography Description Application(s)

- Used to analyze volatile o Detects bombs in
gases. airports
o Identifies and
quantifies drugs, i.e.,
a) Gas Chromatography
in alcohols o
(Forensics) Compares
fibers found on a
victim
- Used to analyze the o Detects pollution by
metal ions and organic testing water
compounds in samples
solutions.
b) Liquid Chromatography - uses liquids that may
incorporate hydrophilic,
insoluble molecules
- common type of o Separates amino
chromatography acids and anions
- paper is the stationary o RNA fingerprinting o
c) Paper Chromatography phase Separates and tests
- uses capillary action to histamines and
pull the solutes up antibiotics
through the paper and
separate the solutes
 o Detects pesticide or
- simple and quick insecticide residues
d) Thin-layer method to check the in food
Chromatography purity of the organic o Analyzes the dye
compounds composition of fibers
(forensics)

4) Distillation

This is the process by which organic compounds (contain carbon) are purified. 7 This is
advantageous in the sense that two different compounds have different boiling points. Take,
for example, two different liquids are present in a homogeneous mixture (completely
miscible, e.g., H2O & alcohol). If these compounds have different boiling points, one of the
compounds will evaporate first (volatile) before the other will.

5) Drying

This is defined as the process of vaporizing and removing water or other liquids in a certain
material to form a dry solid. Physiochemical transformations result from the simultaneous
heat and mass transfer (which are critical factors in the drying process).

6) Grinding

This is the process of turning a material into small pieces from fine to coarse using a
certain device, i.e., Wiley Mill or a grinding wheel.

7) pH measurement
A pH is a measurement of how weak and strong a particular acidic or basic substance. A
substance with a pH of 7 is neutral, i.e., H2O. A pH lower than 7 means acidic, i.e., acetic
acid found in vinegar. A pH higher than 7 is a base, i.e., sodium hypochlorite found in
bleaching products.

In many studies, pH is an important parameter or variable. For example, pH is an

important parameter or factor that is measured to determine the condition of water because
of pollution. How do you relate pH measurement with pollution or water quality? With the
use of pH, one can measure how acidic or basic water is to determine its water quality. Pure
water has a pH of 7, a little lower or higher than that can be considered normal. But when
the pH goes significantly lower, it means the substance is already acidic and basic when it
goes far beyond pH 7. If drinking water for example becomes acidic or basic only indicates
that other components are visible. A pH of 7.4. of water in rivers means good quality. Any
higher or lower than pH 7 means harm to the organisms living in the water.

8) Weighing
This procedure shows how o use a balance to make a precise and accurate measurement,
i.e., mass. To do this, the use of measuring devices is very essential.

Example (Measuring devices)

o Solids – triple balance, top-pan electric balance, analytical electric balance, etc.
o Liquids or solids that do not holder together (i.e., powder, ice, etc.) – receptacle,
graduated cylinder, etc.