Isolating Helminth Eggs using Gasoline as a

Substitute to Ether in the Formalin-Ether

Concentration Technique

Erylle P. Cadiente
Rhubilyn L. Landero
Rhea F. Suberon
Maria Rheena Flor C. Zamora

Bachelor in Medical Laboratory Science, University of Immaculate Conception, Father

Selga St., Davao City

Abstract

The Formalin- Ether Concentration Technique is a very useful and effective

procedure in the diagnosing parasitosis of man but is seldomly used in the clinical
laboratory since ether can only be obtained in small due to existing Laws regulating its
production and distribution because it is expensive, flammable, volatile, produces
anesthetic vapor and explosive (Robert et al., 1996).

Gasoline (Regular, Special, Unleaded) were used and proven viable for the
substation of Ether in the said Concentration Technique introducing the Formalin-
Gasoline Concentration Technique. 200 clinical known positive fecal specimens were
used in this comparative descriptive, experimental type of study wherein four solvents
were being compared in the natural setting of the procedure.

Results as treated with F-test, ANOVA, Duncan’s and Scheffe’s in the statistics
show that there is no significant difference in the effectiveness in the parasitic recovery
rate and macroscopic clarity rate of the sample preparation between ether and the three
types of gasoline. There is a significant difference in terms of microscopic clarity rate for
Gasoline- Regular compared the effectiveness of other solvents. Thus, Gasoline- Special
and Gasoline- Unleaded viable in substituting ether in the concentration technique
considering the parameters in assessing their effectiveness.

Key Words: FECT, FGCT, Ether, Gasoline, Concentration Technique

 Introduction

In isolating parasites that can be found in the fecal material, several procedures
may be used in order to diagnose the presence of a certain underlying pathologic
conditions due to parasitosis. The procedures to be used may range from the simple
Direct Fecal Smear (DFS) to the more complex procedures of fecal concentration
techniques. The simplicity or the complexity of the procedure used will greatly affect the
amount of parasites recovered (Zeibig, 1997).
Gasoline is a promising substitute for ether since gasoline is of the same type of
compound as that of ether. Both gasoline and ether are organic hydrocarbon compounds,
meaning they are made up of Carbon, Hydrogen, and Oxygen atoms. Furthermore, they
are both of almost the same Density or Specific Gravity, which is very important in the
separation of the Fecal Plug formed in concentration techniques. Gasoline is also far
more available than Ether, thus making it a very viable substitute.
In the Philippine setting, FECT is one of the most effective procedure used to
diagnose different types of intestinal parasitosis. It is a sedimentation method to separate
parasitic elements from fecal debris through centrifugation (Robert and Jonovy, 1996).
Although FECT is the best procedure in isolating parasites present in the feces, since this
technique provides less or very minimal alterations to the organisms present in the stool
sample and increased recovery of Helminth eggs as well as protozoan cysts, it is seldom
performed because of the unavailability of ether. Ether can only be obtained in small
amounts since there are Laws that regulate the production, distribution, and trade of ether,
because it is extremely flammable, is highly volatile, produces anesthetic vapor, and
forms explosive peroxides when exposed to light (Robert and Jonovy, 1996; Boswell and
Collins, 1996).
Medical Technologists working in the Parasitology Section of the Clinical
Laboratory opt to find substances to substitute ether or use the less effective method,
Direct Fecal Smear, to overcome these disadvantages, since pursuing the performance of
FECT in diagnosing parasitosis implies an additional cost to the laboratory, as well as
rendering it unavailable to everyone (Robert and Jonovy, 1996).
In this research paper, the study routinely used Formalin-Ether Concentration
Technique procedure and the Formalin Gasoline Concentration Technique for isolating
parasites from known samples, and compare which among the three types of gasoline
(Regular, Special, Unleaded) will yield the best result in terms of parasitic recovery rate
and sample clarity rates (macroscopic and microscopic) were studied.
 Materials and Method

This research is a comparative descriptive, because there were four kinds of

solvents whose characteristics are being compared in the natural setting of the procedure.
Experimental that it examines the effect of substituting different types of gasoline to ether
in the FECT.
This research is a non-probability type of study. The subjects of this research
were the respondents or the ten individuals from which the positive samples came from.
Ten individuals with known cases of parasitosis were selected from the selected families
of Brgy. 5A- Bankerohan, Davao City used in the analysis of the viability of the gasoline
types as substitutes to ether.
Collection of specimen was done from ten individuals in Barangay 5-A,
Bankerohan, Davao City with known positive fecal samples. The experimental
procedures involved in the study were conducted at the 4th floor Microbiology
Laboratory, University of the Immaculate Conception, Fr. Selga st., Davao City, which
provided a highly controlled laboratory setting performing the Formalin-Ether
Concentration Technique and Formalin-Gasoline Concentration Technique. The
laboratory was provided with all the materials needed in the study except the three types
of gasoline which were provided by the proponents.

Results and Discussion

Table 1.1
Summary of Mean for Recovery Rate
ETHER REGULAR SPECIAL UNLEADED
Sample (%) (%) (%) (%)
1 100 100 100 100
2 100 80 20 20
3 100 100 100 100
4 80 80 100 100
5 100 100 80 80
6 100 100 100 100
7 100 100 100 100
8 100 100 100 100
9 100 100 100 100
10 100 100 100 100
Mean 98 96 90 90
 Table 1.1 shows the summary of the means of the recovery rate for each type of
solvent used in each sample. In each sample, there are five repetitions of each solvent
used, and the means of the trials are plotted under their corresponding solvent. After
which, a mean is derived from the means of the trials in each sample under each solvent.
These final means are those that are found plotted under table 1.2 and are tested using the
F-test for computed F-ratio and probability F-ratio under the significance level of 0.05 in
order to decide whether to accept or reject the null hypothesis or if there is a significant
difference or not.

Table 1.2
F-Test for the Significance of the Difference Between the Four Formalin-Concentration
Techniques when Analyzed in Terms of Recovery Rate

Variables Mean Computed F- Probability F- Decision on Ho

ratio ratio @ 0.05

Ether 98

Regular 96 Accept Ho

0.486 0.694
Special 90 Not Significant

Unleaded 90

Table 1.2 presents the test for the significance of the difference between the Four
Formalin-Concentration Technique when analyzed in terms of recovery rate. From the
table, the mean for ether is 98 %, regular gasoline is 96 %, special 90 % and unleaded is
90 %. Since there are four variables to be compared, one way Analysis of Variance was
used. Further analysis would show that the computed F-ratio yielded a value of 0.486
with a probability F-ratio of 0.694. For the computed F-ratio to be significant, the
probability F-ratio must be less than or equal to 0.05 level of significance. In this case,
the probability F-ratio is greater than 0.05 level of significance, therefore, there is enough
evidence to accept the null hypothesis of no significant difference. This means that there
is no significant difference between the recovery rates of the four Formalin Concentration
Techniques. This implies that the four concentration techniques have the same effect in
terms of recovering parasite eggs from fecal samples. Although there is a difference
between the values of the four means that are being compared, these values are not
statistically significant.

Table 2.1
Summary of Mean for Clarity Rate (Macroscopic)
ETHER REGULAR SPECIAL UNLEADED
Sample (%) (%) (%) (%)
1 86.67 86.67 80 66.67
2 100 33.33 33.33 53.33
3 66.67 33.33 66.67 33.33
4 93.33 66.67 66.67 100
5 80.00 80.00 100 66.67
6 66.67 66.67 33.33 33.33
7 100 100 73.34 93.33
8 86.67 93.33 93.33 100
9 93.33 93.33 73.34 93.33
10 93.33 86.67 93.33 80.00
Mean 86.67 74.00 71.33 72.00

Table 2.1 presents the summary of the means of the macroscopic clarity rate for
each type of solvent used in each sample. In each sample, there are five repetitions for
each solvent used, and the means of the trials are outlined under their corresponding
solvent. After which, a final mean is derived from the means of the trials in each sample
under each solvent. These final means are those that are found plotted under table 2.2 and
are tested using the F-test for computed F-ratio and probability F-ratio under the
significance level of 0.05 in order to resolve whether to accept or reject the null
hypothesis or if there is a significant difference or not.

Table 2.2
F-Test for the Significance of the Difference between the Four Formalin-Concentration
Techniques when analyzed according to Clarity (Macroscopic)

Variables Mean Computed F- Probability F- Decision on Ho

ratio ratio @ 0.05
Ether 86.67

Regular 74.00 Accept Ho

1.081 0.370
Special 71.33 Not Significant

Unleaded 72.00

Presented on table 2.2 is the test for the significance of the difference between the
four formalin concentration techniques when analyzed according to clarity
(macroscopic). The table shows that ether has the highest mean of 86.67 followed by
regular gasoline 74.00, unleaded gasoline with 72.00 and special gasoline with 71.33. To
test the significant difference between the four variables that are being compared, one
way analysis of variance was utilized. The table revealed that the computed F-ratio is
1.081 and the probability F-ratio is 0.0370 at 0.05 level of significance. Since the
probability F-ratio is greater than 0.05 level of significance, the null hypothesis of no
significant difference is accepted. This means that there is no significant difference
between the means of the four Formalin Concentration Techniques in terms of
macroscopic clarity rates. This implies that the four formalin concentration techniques
namely: formalin-ether, formalin-regular, formalin-special and formalin-unleaded have
similar effects in terms of macroscopic clarity rates. The lack of significant difference
between the four variables strongly allude that the three types of gasoline are viable for
substitution to ether in the Formalin-Ether Concentration Technique in terms of the
macroscopic clarity of sample preparation.

Table 3.1
Summary of Mean for Clarity Rate (Microscopic)
ETHER REGULAR SPECIAL UNLEADED
Sample (%) (%) (%) (%)
1 32 28 48 44
2 40 36 40 40
3 68 28 48 56
4 44 20 20 40
5 24 24 40 36
6 60 60 60 64
7 20 20 20 52
8 52 24 60 64
9 56 20 56 52
10 20 20 32 40
Mean 41.60 28.00 42.40 48.80
 Table 3.1 exhibits the summary of the means of the microscopic clarity rate for
each type of solvent used in each sample. In each sample, there are five repetitions of
each solvent used, and the means of the trials are plotted under their corresponding
solvent. After which, a mean is obtained from the means of the trials in each sample
under each solvent. These final means are those that are found mapped under table 3.2
and are tested using the F-test for computed F-ratio and probability F-ratio under the
significance level of 0.05 in order to decided whether to accept or reject the null
hypothesis or if there is a significant difference or not.

Table 3.2
F-Test for the Significance of the Difference Between the Four Formalin-Concentration
Techniques when Analyzed According to Microscopic Clarity

Variables Mean Computed F- Probability F- Decision on Ho

ratio ratio @ 0.05

Ether 41.60

Regular 28.00 Reject Ho

3.928 0.016
Special 42.40 Significant

Unleaded 48.80

Table 3.2 displays the test for the significance of the differences between the four
formalin concentration techniques when analyzed according to microscopic clarity. From
the table, unleaded gasoline has the highest mean among the four concentration
techniques with a value of 48.80, second in line is special gasoline with a value of 42.40,
followed by ether with a mean of 41.60, and the lastly, regular gasoline with 28.00. The
table shows that the computed F-ratio is 3.928 and the probability F-ration is 0.016 at
0.05 level of significance. Since the probability F-ratio is less than 0.05 level of
significance, then the null hypothesis of no significant difference is rejected. This means
that there is a significant difference between the Microscopic Clarity Rates of the four
Formalin Concentration Techniques. To determine which of the four techniques differ
significantly, Scheffe multiple comparison was applied. This technique revealed that there
is a significant difference between regular gasoline and ether in favor of ether, there is
also a significant difference between regular and special in favor of special, and there is a
significant difference between unleaded and regular in favor of unleaded.
However, there was no significant difference between the three concentration
techniques namely; formalin-ether, formalin-special and formalin-unleaded. This result
particularly entails that the two types of gasoline, Special and Unleaded are viable
substitutes to Ether in the Formalin-Ether Concentration Technique. With regards to the
Regular Gasoline, since among the four solvents it has the lowest mean value, it is
implied that it is not viable as a substitute to Ether in the Formalin-Ether Concentration
Technique in terms of microscopic clarity of the sample preparation.

Conclusion

Based on the results presented, there is no significant difference in effectiveness

in terms of parasitic recovery rate macroscopic clarity rate of the sample preparation
between ether and the three types of gasoline, namely: Regular, Special and Unleaded
when used as solvents in the FECT and FGCT in isolating helminth eggs. There is a
significant difference in effectiveness in terms of microscopic clarity rate between
Regular-Gasoline as compared to Ether, Gasoline-Special and Gasoline- Unleaded when
used as solvents in the FECT and FGCT in isolating helminth eggs.

Recommendations

In this study, the researchers would like to recommend the following:

1. Recommending the use of Gasoline-Special and Gasoline-Unleaded as a

substitute to ether in Formalin- Ether Concentration technique.
2. It is recommended to conduct research and experimentation to test the protozoan-
isolating capacity of gasoline.
3. Recommendation of the viability of the study to concerning health and/or
academic agencies for support and promotion of usage of Gasoline-Special and
Gasoline- Unleaded as substitute to Ether in FECT in the clinical laboratories.
4. The researchers would want to recommend that multiple related tests be
performed in the future to completely evaluate the helminth-isolating capacity of
gasoline.
 Acknowledgement

Sincere gratitude to all the people who greatly contributed in the success of this
research paper mostly to the Almighty God and to the parents of the authors. To Adorico
M. Aya-Ay, PhD, Avee Joy B. Dayaganon, RMT, MSMT, John Mark De Real, RMT,
Hon. Edgardo Ibuyan, Jr., S. Ma. Carolina Perales, RVM, Maritess Saco, RMT,
Micheline Phoebe Pangilinan, M.D., CFP, Ursula Quiban, PhD, Carlito Yurango, Venchie
Badong, MAT, Iven Christ Castromayor, RMT, Desiree Victorino, RMT, and Ace Ronald
Sarabia, RMT.

References

Robert, Peter J. and Jonovy, Helen M. (1996). Basic Parasitology. Manila, Philippines;
Pearson Education Asia

Markell, John S. (2002). Chemistry for Health Sciences. Manila, Philippines; Pearson
Education Asia.

Belizario, Vicente Y., M.D., MTM&H and de Leon, W.U., RMT, DAP&E,
MPH.(2004).Philippine Textbook of Medical Parasitology (2nd
Edition).Manila,Philippines; The Publications Program.