Chapter III

Methodology

The study will use the experimental method of research. The method is a highly controlled
procedure in which manipulated treatment or actions from a factor or condition, called the
experimental variable (the Achuete ink) will be applied upon another factor or condition, and
called the dependent variables (absorption, stick consistency and color) to determine the effect
of the former upon the latter. All other factors or variables will be kept constant or equal so that
any change in the dependent variables is attributable only to the independent variable.

This study also focused on how the Achuete seed extract is capable of being utilized as
ingredient for the permanent marker ink. Additionally, the Achuete seed extract will be tested
to determine its effectiveness and through experiments, observations will be gathered and
utilized as a collective in making conclusions.

The experimental set-up consists of three (3) sub set-ups and the controlled set-up. Each set-up
has to undergo in the same process, same environment and same tests in order to avoid
alteration and confusion of observation. The set-up with three (3) replications will be tested
with the use of a standardized test, the Waterproof Test.

This study will also use another method in data gathering. Thirty respondents will be asked to
rate the ink in terms of absorption, color and over-all acceptability.
INSTRUMENTATION

The study used the following method of instrumentation in data gathering:

Questionnaire. The questionnaire will be framed by the researcher in order to gather data in
determining a) the effectiveness of Achuete ink in terms of the absorption, stick consistency,
color and over-all acceptability of the ink and b) the significant difference between the standard
ink and Achuete ink. This had two parts, the respondent’s profile, and question-scale part.

Part I will be composed of personal information of the respondents such as name, section and
sex. It also includes the date of observation. The respondents were identified through random
sampling.

Part II was divided into two parts: a) the acceptability of the Achuete ink and b) significant
difference between the Achuete ink and standard ink. Respondents will rate the Achuete ink
and standard ink in terms of color, odor, and rate of evaporation and absorption of ink.

In order to rate the variables, the Likert’s scale will be used in which 5 means exceptional, 4
means very good, 3 means good, 2 means fair and 1 means poor. Instructions will be given that
served as guide for the respondents on how to rate the variables.

Waterproof Test. The waterproof test will be also used in the study to test the stick consistency
of the ink.

LIST OF MATERIALS

Vials

Test Tubes

450ml and 100 ml Beakers

10ml , 20ml , 50ml, and 100ml graduated cylinders

Stirring Rod

Mortar and pestle

Filter Paper

Double beam balance

Laboratory gloves

LIST OF INGREDIENTS

Glycerol/Glycerin

Benzalkonium chloride
Achuete Gall extract

Ferrous Sulphate

GENERAL PROCEDURE

A. PREPARATION OF ACHUETE SEED EXTRACT

The process in extracting achuete seeds was based on the study SUBSTITUTE INDELIBLE INK
USING Bixa orellana L. (ANNATTO) AND Corcus sativus L. (KASUBHA) EXTRACTS

100 gram of achuete seeds , 15 mL water and 50 mL Benzalkonium chloride will be collected for
extracting the achuete seeds. The prepared achuete will be boiled in 15 mL water and will be
placed in a casserole and heated on a stove. The extract will be placed in small bowl and waited
until it cools down. The extracted Achuete seed will be mixed with Benzalkonium chloride and
will be filtered.

B. PREPARATION OF THE INK

Other substances known as additives such as glycerol or glycerin and ferrous sulfate will be
added with three different proportions. The Benzalkonium chloride prevents the ink from
solidifying. This helps in reducing the surface tension of the ink solution, allowing the ink to
soak more quickly into the paper fibers. It also prevents molds from growing in the ink.

The glycerol will be also used because of its viscous nature and high absorbency on paper. It
increases the rate of extraction of tannin in the pigment.The ferrous sulfate is a disinfectant that
aids in the inhibition of growth of microorganisms in the ink. It is responsible for the black color
of Achuete ink.

The formulized ink from the extract of Achuete seeds will be transferred into ink bottles and will
be left undisturbed for 24 hours before undergoing in a series of tests. The recycled permanent
marker containers will be refilled with the Achuete ink. The table below shows the proportion of
ingredients that was used in making the Achuete ink. The proportion of ingredients was based
on the study Narra (Pterocarpus indicus) Bark Extract as a Permanent Marker Ink.
Table 1. Proportions of Achuete Ink Ingredients

Sample Achuete Glycerol BENZALKONIUM FERROUS WATER

Seeds CHLORIDE SULPHATE
(mL) (mL)
(gram) (mL) (mg)

A 100 1.0 93 1500 15

B 100 2.25 93 1500 15

C 100 3.5 93 1500 15

C. EVALUATION OF SAMPLES

The researchers will evaluate each sample to determine the difference between the Achuete ink
and standard ink using a waterproof test for quality inks:

i. Waterproof Test of Ink

Both the Achuete and standard ink will be subjected to the same test on the same sheet of
paper.

Samples will be written in the paper and the page was left out overnight (about 9 hours) before
doing the test. The water test will include holding the page under runningwater for about 30
seconds on each side of the page. To dry, the page will be putted between two paper towels
with a lightweight box placed on top to aid in drying.

For the judging factors, the results will be observed carefully for clarity of stroke, retention of
original color, and leftover 'halos' of ink around the letters. The paper towels that were used to
dry the papers were also examined for ink transfer. The judging factors will determine the score
or rating on each ink with 5 as the highest, indicating the ink’s retention on the paper, and 1 as
the lowest, indicating that the words were unreadable after test.4

STATISTICAL TOOL

The data that will be gathered from the questionnaire would be classified, tabulated, computed
and analyzed statistically using chi-square and weighted mean to test the hypothesis of the
study.

The chi-square test of homogeneity shows the significant difference between the Achuete ink
and the standard ink. The weighted mean tests will be the acceptability of the Achuete ink.

The formula to be used for the chi-square test of homogeneity is:

Wherein O is the observed frequency, E is the expected frequency and is the p-value.

To compute for E, use: where RT is the total in a row, CT is the total in a

column and GT is the grand total.4