BIOCHEMISTRY

2.04 Pentose Phosphate Pathway & Uronic Acid Pathway

Dr. Maria Lilia Reyes | 30 September 2016
LE 2

OUTLINE Warburg’s effect: Basis for several methods of

detecting and treating cancer. The case of abnormal
I. PENTOSE PHOSPHATE PATHWAY regulation of glycolysis is seen in cancer. The tumors of
A. Overview (definition, products, function) nearly all types carry out glycolysis at a much higher rate than
B. Oxidative Phase normal tissue, even when oxygen is available.
C. Non-oxidative phase
D. Comparison of Oxidative and Non- Otto Warburg – made two hypotheses regarding this
oxidative phase data:
E. PPP in relation to Glycolysis
F. Biomedical importance First, that the increased ratio of glycolysis to aerobic
1. Prolongation of RBC’s lifespan respiration was likely due to defects in the mitochondrial
2. G6-PD deficiency respiratory chain
G. Regulation of PPP
H. Other pathways requiring NAPDH Second, that this enhanced glycolysis enabled cancer cells to
II. URONIC ACID PATHWAY preferentially proliferate in the reduced oxygen tension often
seen in tumors. Furthermore, Warburg argued that the switch
A. Overview (definition, products, function)
from aerobic to anaerobic glucose metabolism was the
B. Steps driver of tumorigenesis. (Lehninger’s) (Harper’s)
C. Biochemical Importance
1. Man’s inability to Synthesize Ascorbic
Acid  Function
2. Pentosuria - Generation of NADPH
III. Summary o fatty acid synthesis
IV. Questions o maintaining reduced glutathione
V. Appendix (antioxidant)
- Provision of ribose residues for nucleic acid
formation
- Metabolic utilization of 5-carbon sugars
VI.
OBJECTIVES:  Sites
At the end of the lecture, the student should be able to: - Fatty acid synthesis: liver, adipose tissues,
1. VII. Penalties
Explain the role of the pathway in metabolism in terms lactating mammary glands and red blood cells
of its substrates and products - Steroid synthesis: adrenals, testes and ovaries
2. VIII. Tables,the
Enumerate Images and Figures
key features of the PPP.  Main Products : NADPH, ribose-5-phosphate and
3. Discuss the essential role of the PPP in providing key CO2 (in RBC)
biosynthesis intermediates.  Anabolic (synthetic purposes)
4. Discuss the regulation of the pathway in different  2 Phases of reactions:
metabolic states (fed/fasting state) Oxidative, irreversible phase
Products: 2 NADPH + 1 ribulose-5-phosphate + CO2
Legend: Non-oxidative, reversible phase
Remember Previous Products: F-6-P, G-3-P
Lecturer Book Trans Com
(Exams) Trans

     B. Oxidative Pathway

I. PENTOSE PHOSPHATE PATHWAY STEP 1: Dehydrogenation (aka oxidation)

A. Overview
 A cytosolic route for complete oxidation of glucose,
producing a pentose phosphate, NADPH, and CO2.
Favorite question in the exam:
This pathway produces vitamin C (ascorbic acid) but NOT in
humans.
Which is this enzyme present in lower mammals that
makes vitamin C needed in our diet?
Answer: Gulonolactone Oxidase
 Other names
- Hexose Monophosphate Shunt
- Phosphogluconate pathway
- Direct Oxidative Pathway
Figure 1: Dehydrogenation of Glucose 6-Phosphate
- Warburg-Dickens Pathway

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 2.04 PPP & Uronic Acid Pathway BIOCHEMISTRY 2020C

- Note: 3-keto-phosphogluconate = unstsable enediol

Enzyme: Glucose-6-phosphate dehydrogenase intermediate  Ribulose-5-phosphate will always
+
- NADP -dependent form
2+ 2+
- Cofactors: Mg and Ca  C3 of 6-phosphogluconate donates a hydride ion to
Reaction: at C1 NADP
+

 Rate-limiting step: major regulatory site  Spontaneous oxidative decarboxylation of 3-keto-

 “shunt”: goes directly to PPP instead of glycolysis
 No standard free energy values because it is mainly
for synthesis of required products, not energy
production
 Produces NADPH
Inhibition: NADPH (allosteric)  self-limiting
Recall: G6PD deficiency is the most common cause of
hemolytic anemia; screening for this deficiency is part of the
newborn screening program (heel stick puncture)
STEP 2: Hydrolysis
phosphogluconate  removal of CO2  ribulose-5-
phosphate (ketopentose)
 Endpoint of oxidative phase
End products for the Oxidative Phase: 2 NADPH + ribulose-
5-phosphate + CO2
NOTE: Under certain metabolic conditions, PPP can end at this
point
- NADPH  reductive biosynthetic reactions
- Ribulose-5-phosphate  ribose-5-phosphate (via
ribose-5-phosphate ketoisomerase  nucleotide
synthesis

Biochemical Application: (good to know)

Figure 2: Hydrolysis of 6-phosphogluconolactone
Enzyme: Gluconolactonase (lactonase)
2+ 2+ 2+
- Cofactor: Mg , Ca or Mn (divalent cations)
Lactone: substrate for glucolactonase which ensures that
the reaction goes into completion
STEP 3: Dehydrogenation and Decarboxylation
hexose-6-phosphate dehydrogenase
 Found in the endoplasmic reticulum,
 isoenzyme of glucose-6-phosphate dehydrogenase
 provides NADPH for hydroxylation reactions
 provides NADPH for 11-β-hydroxysteroid
dehydrogenase-1
11-β-hydroxysteroid dehydrogenase-1
 catalyzes the reduction of cortisone (inactive) 
cortisol (active) in the liver, nervous system, and
adipose tissue
 Major source of intracellular cortisol in these tissues
and plays an important role in obesity and
metabolism (Harper’s)
C. Non-Oxidative Pathway
NOTE: since all reactions are reversible, direction of
pathway depends on the needs of the cell
 Ribulose-5-phosphate  ribose-5-phosphate +
xylulose-5-phosphate  glycolysis intermediates
 Main enzymes involved are an epimerase,
isomerase, transketolase, and transaldolase

STEP 1: Isomerization

Figure 3: Dehydrogenation and Decarboxlyation of 6-

Phosphogluconate

Enzyme: 6-phosphogluconate dehydrogenase

+
- NADP -dependent
2+ 2+
- Cofactors: Mg and Ca
Figure 4: Conversion of Ribulose 5-phosphate to
aldopentose and ketopentose

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Reaction: STEP 3: Transfer of 3C dihydroxyacetone group

 ribulose-5-phosphate (aldopentose) via ribose-5-phosphate
isomerase
 xylulose-5-phosphate (ketopentose) via ribulose-5-
phosphate 3-epimerase

 Ribose-5-phosphate and xylulose-5-phosphate are

epimers at C3

STEP 2: Transfer of 2C ketol group

Figure 6: Transferring of 3-carbon dihydroxyacetone

group of seduheptulose 7-phosphate to glyceraldehyde 3-
phosphate by means of Transaldolase

Enzyme: Transaldolase (specifically

dihydroxyacetonetransferase)
Reaction: 3-carbon dihydroxyacetone group of
seduheptulose7-phosphate to glyceraldehyde-3-phosphate

Products:
1. Fructose-6-phosphate from addition
- Intermediate in glycolysis
2. Erythrose-4-Phosphate (4C-aldose sugar) from removal

Figure 5: Transferring of the two gycoaldehyde group of Remember: Number of C atoms will not change
xylulose 5-phosphate to the aldehyde carbon of ribose 5-
phosphate Ribose-5-phosphate and xylulose-5-phosphate = 10C
Seduloheptulose-7-phosphate and glyceraldehye-3-phosphate
Enzyme: Transketolase (specifically glycoaldehdyetransferase) =10 C
Reaction: glycoaldehyde group (2C ketol group) of xylulose- Fructose-6-phosphate and Erythrose-4-phosphate = 10C
5-phosphate to the aldehyde carbon of ribose-5-phosphate
STEP 4: Transfer of 2C group
Transketolase

2+
Cofactors: TPP (Thiamine pyrophosphate), Mg
o participate in the transfer of the keto group
 Measured to assay thiamine deficiency

Biochemical Application:
Erythrocyte Transketolase Study
 can be ordered in the lab and may give a clue if an
individual is thiamine deficient

Products:
1. Sedoheptulose-7-phosphate from the addition
2. Glyceradehyde-3-phosphate from the removal

Figure 7: Transferring of C1 and C2 of Xylulose 5-

phosphate to c1 of erythrose 4-phosphate

Enzyme: transketolase
2+
- Cofactors: TPP, Mg

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Reaction: transfers C1 and C2 of Xylulose-5-PO4 (donor) E. PPP in Relation to Glycolysis

to C1 of Erythrose-4-PO4 (acceptor) A. Differences
Table 3: A brief comparison between Glycolysis and
Products: Pentose Phosphate Pathway (from lecturer’s ppt)
1. Fructose 6-phosphate Features Glycolysis PPP
- Intermediate in glycolysis/gluconeogenesis Glucose-6- Substrate Substrate and end-
2. Glyceraldehyde 3-phosphate phosphate product
- Intermediate in glycolysis/gluconeogensis CO2 Not produced produced
+ +
Hydrogen acceptor NAD NADP
Table 1. Summary of the Non-Oxidative Phase ATP Generation Major function None
Substrate Enzyme Product Ribose-5- None Present
Ribulose-5-P Isomerase Ribose-5-P phosphate
(ketopentose) (aldopentose) generation
2 Ribulose 5-p Epimerase 2 Xylulose-5-P
Xylulose-5-P + Transketolase Seduheptulose-7- B. Similarities/Connections (good to know from
Ribose-5-P P+ Harper’s)
Glyceraldehyde-3- Xylulose -5-phosphate  activates the protein
P phosphatase  dephosphorylates the 6-
Seduheptulose-7- Transaldolase Xylulose-5- phosphofructokinase/fructose 2,6-biphosphate
P+ P+Erythrose-4-P functional enzyme  increased formation of fructose
Glyceraldehyde-3- 2,6-biphosphate  increased activity of
P phosphofructose kinase-1 (PFK-1)  INCREASED
Xylulose-5- Transketolase Glyceraldehyde-3- GLYCOLYTIC FLUX
P+Erythrose-4-P P+ Fructose-6-P
3 Ribulose-5-P Summary (see Glyceraldehyde-3- Xylulose -5-phosphate
appendix) P+ 2 Fructose-6-P  Initiates the nuclear translocation and DNA binding
of the carbohydrate response element binding
 A way to remember the non-oxidative pathway is protein  increased synthesis of fatty acids in
through the # of Cs per reaction: response to a high carbohydrate diet

2 5’s  5 + 5  7 + 3  6 + (4 + 5)  6 + 3 F. Regulation of PPP

The ones in red are shuttled back into glycolysis.

2 ribulose-5-P  xylulose-5-P + ribose-5-P 

sedoheptulose-7-P + G-3-P  F-6-P + (erythrose-4-P +
xylulose-5-P)  F-6-P + G-3-P

D. Comparison of Oxidative and Non-oxidative Pathway

Table 2. Overview of the PPP (oxidative and non-
oxidative phase)
Stage Reversibility General General
Process Products
OXIDATIVE Irreversible G-6-P 2 moles of
STAGE redox stage oxidation NADPH per
and mole of
decarboxylat glucose-6-P;
ion  ribulose-5-P Figure 7: A brief summary of PPP
ribulose-5- and other
phosphate pentose  Final product of PPP can change depending on
phosphatases the metabolic needs of the cell
NON- Reversible F-6-P and 2 moles of - CELL NEEDS MORE NADPH
OXIDATIVE interconversio G-3-P fructose-6-P o synthesis of fatty acids and steroids
STAGE n stage interconversi and 1 mole of o ribose-5-phosphate  F-6-P + G-3-P
on to 5C gly-3-P for - CELL NEEDS TO SYNTHESIZE
sugars every 3 moles NUCLEOTIDES AND NUCLEIC ACIDS
of ribulose-5- o in rapidly-dividing cells
phosphate o F-6-P + G-3-P  ribose-5-phosphate

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G. Biomedical Importance II. URONIC ACID PATHWAY

1.
Prolongs RBC lifespan
A. Overview
 PPP: sole source of NADPH for the reduction of
oxidized glutathione by glutathione reductase  An alternative pathway for glucose metabolism into
(flavoprotein containing FAD) glucuronic acid, ascorbic acid (not performed in
 Reduced glutathione: removes H2O2 in a reaction humans), and pentoses.
catalyzed by glutathione peroxidase (enzyme  Does not produce ATP or any other reducing
containing the selenium analog of cysteine) at the equivalents (CO2, O2, etc).
active site  Mainly occurs in the liver.
 H2O2 accumulation  SIGNIFICANCE
o oxidative damage to the cell membrane  a. Detoxification of foreign compounds and
hemolysis drugs (covaently attached glucuronyl
o oxidizing hemoglobin to methemoglobin residues impart polarity; readily excretable)
decrease lifespan, inefficient transfer of oxygen b. Synthesis of mucopolysaccharides
2. Glucose-6-Phosphate Dehydrogenase Deficiency  URONIC ACID = class of sugar acids whose terminal
 Most common cause of hemolytic anemia carbon’s hydroxyl group has been oxidized to a
 Deficient synthesis of NADPH carboxylic acid
Deficiency can be partial or complete, since some
patients do not show symptoms right away
 inherited or acquired
Can grow to full adulthood and not exhibit symptoms
until exposed to the following:
o Anti-malarial drugs (primaquine).
Chloramphenicol, Aspirin, Sulfonamide
o Food: Fava beans: generation now would be
from parents of mixed races – more
susceptible to hemolytic anemia

Figure 9: glucose and glucuronic acid structures

B. Steps
STEP 1: CONVERSION OF GLUCOSE-6-PHOSPHATE TO
GLUCOSE-1-PHOSPHATE
Enzyme: Phosphoglucomutase
Reaction: Glucose-6-phosphate is isomerized to Glucose-
Figure 8. Role of the pentose phosphate pathway in the 1-Phosphate
glutathione peroxidase reaction of erythrocytes.

H. Other Pathways that Require NADPH

1. Reductive Synthesis (Anabolic Pathways)
 Fatty acid synthesis
 Fatty acid chain elongation
 Cholesterol synthesis
 Neurotransmitter synthesis
 Nucleotide synthesis
 Superoxide synthesis
2. Detoxification
a. Cytochrome P450 monooxygenase: NADPH is
the source of reducing equivalents for cyt P450
hydroxylation of aromatic compounds, steroids,
alcohol and drugs  detoxification
b. NADPH maintains levels of reduced
glutathione important for
- RBC membrane integrity
- Defense against reactive oxygen species and
free radicals
Reduced glutathione: injecting patients with liver or
hepatocellular cancer with reduced glutathione stop tumor
spread  detoxification  better quality of life, skin,
improvement of activity of liver enzymes

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STEP 2: FORMATION OF UDP GLUCOSE - Glucuronic acid is encountered in bilirubin

Enzyme: UDP Glucose Phosphorylase
UDP is a glucose carrier. In contrast, ADP, GDP, and
CMP are carriers for other sugars.
Reaction: Glucose-1-Phosphate reacts with Uridine
triphosphate (UTP) to form UDP Glucose.
conjugation from B1/ unconjugated bilirubin to
B2/ conjugated bilirubin
Conjugated = more polar and water soluble form, in the
liver for easier excretion of bile as glucuronide conjugates.



STEP 5: OXIDATION OF D-GLUCURONIC ACID
STEP 3: OXIDATION OF UDP GLUCOSE Enzyme: L-Gulonic Dehydrogenase
Enzyme: UDP Glucose Dehydrogenase L-Gulonate is the direct precursor of ascorbate in animals
+
UDP Glucose is oxidized at Carbon 6 by NAD dependent capable of synthesizing Ascorbic Acid/Vitamin C, in an
UDP Glucose Dehydrogenase in a two-step reaction to NADPH- dependent reaction.
form UDP Glucuronic Acid (or Activated Glucuronic Acid) In humans, Ascorbic Acid cannot be synthesized
because of the absence of L-Gulonolactone Oxidase
UDP- Glucuronate: source of glucuronate for reactions Ascorbic Acid/Vitamin C
involving its incorporation - In plants and most higher animals, Glucuronic
EXAMPLES Acid is reduced to L- Gulonic acid and converted
- Formation of proteoglycans to L- Ascorbic acid by the enzyme L-
- reactions with substances such as steroid Gulonolactone Oxidase
hormones, bilirubin and a number of drugs - Humans, other primates and guinea pigs, lack the
that are excreted in urine or bile as enzyme L- Gulonolactone oxidase
glucuronide conjugates. o Due to genetic deficiency of enzyme
o No production of ascorbic acid
- Vitamin C must be supplied through the diet
o needed in iron absorbtion.
- Scurvy
o Results from Vitamin C deficiency.
o Condition of the connective tissues that
causes multiple hemorrhages and
decreased wound healing.

STEP 4: HYDROLYSIS OF UDP GLUCURONIC ACID

Reaction:
- UDP glucuronic acid is hydrolyzed to form UDP
and D-Glucuronic acid.
- Glucuronic Acid = main goal product of uronic
acid pathway.

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STEP 6: OXIDATION OF L-GULONIC ACID STEP 9: REOXIDATION OF XYLITOL

Enzyme: β-L- Hydroxy Acid Dehydrogenase Enzyme: D-Xylulose dehydrogenase
Reaction: L-gulonic acid is oxidized to 3-keto-L-gulonic
acid
NADH is generated (Madarcos Notes)

STEP 10: PHOSPHORYLATION OF D-XYLULOSE

 Enzyme: Xylulose Kinase
 D-Xylulose 5-Phosphate via xylulose kinase further
STEP 7: DECARBOXYLATION OF 3-KETO-L-GULONIC metabolized via the HMP Shunt converted to
ACID intermediates of glycolysis for energy production.
Enzyme: Beta-Keto-L-Gulonate Decarboxylase
C1 of 3-Keto-L-Gulonic Acid is releases as carbon dioxide



 C. Biomedical Importance
STEP 8: OXIDATION OF L-XYLULOSE
1. Man’s inability to Synthesize Ascorbic
Enzyme: Xylitol Dehydrogenase or Xylulose Reductase Acid
In humans, L-Xylulose is the ketopentose excreted in
essential pentosuria
SYNTHESIS OF ASCORBIC ACID
 In plants and most higher animals other than man,
Vitamin C is synthesized directly from L-gulonic acid
by enzyme L-Gulonolactone Oxidase (blocked in
humans, other primates, and guinea pigs)
 Humans must intake Vitamin C from their diet.
 Vitamin C is needed in iron absorption. Lack of
Vitamin C in the diet can lead to Scurvy
o causes multiple hemorrhage and decreased
wound healing.

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QUESTIONS
1.) Which is this enzyme present in lower mammals that
makes vitamin C needed in our diet?
a. Transketolase
b. Gulonolactone Oxidase
c. Glunolactonase
d. Transaldolase
2.) Which of the ff can increased synthesis of fatty acids in
response to a high carbohydrate diet?
a. Erythrose-4-PO4
b. Fructose 6-PO4
c. Glucose 6-Phosphate
d. Xylulose 6-Phosphate
3.) The Uronic Acid Pathway
a. Leads to the formation of ATP
Figure 9. Synthesis of Ascorbic Acid b. Catalyzes the conversion of glucose to glucoronic
acid, ascorbic acid, and pentoses
Essential Pentosuria c. Is an alternative reductive pathway for glucose
 Rare benign hereditary condition metabolism
 Due to: Lack of xylulose reductase -> d. Is a 9-step pathway
enzyme that reduces xylulose to xylitol 4.) TRUE or FALSE
+
 Effect: Xylulose in urine UDP glucose is oxidized at carbon 6 by NAD -
 Implications: can give false positive results dependent UDP glucose dehydrogenase to yield UDP
when urinary glucose is measured using glucuronic acid
alkaline copper reagents (xylulose is a 5.) L-gulonate is the direct precursor of
reducing sugar). a. Ascorbate
Alimentary Pentosuria b. Xylitol
o Occurs after consumption of relatively large c. Glucoronic acid
amounts of fruits rich in pentoses,( e.g d. 3-keto-L-gulonic acid
pears) 6.) Which of the ff is usually associated with thiamine
deficiency?
SUMMARY a. Transaldolase
b. G6P Dehydrogenase
c. Transketolase
 The pentose phosphate pathway (PPP) is a cytosolic d. Gulonolactone Oxidase
process involving complete oxidation of glucose,
producing NADPH and CO2, but not ATP. It is a 7.) The product of the oxidative pathway of the HMP
purely anabolic process. from 3 molecules of glucose 6-phosphate:
 The pathway has two phases, oxidative a. 3 ribulose 5-phosphate
(irreversible), generating NADPH, and non- b. 3 xylose 5-phosphate
oxidative (reversible) which produces ribose c. 3 glucose 6-phosphate
precursors for nucleotide synthesis. The complete d. 3 ribose 5-phosphate
pathway is present mainly in tissues requiring
NADPH; the non-oxidative phase present in all cells Answers: b,d,b,true,a,c,a
requiring ribose. REFERENCES:
 PPP has a major function on erythrocytes, preventing
hemolysis by providing NADPH to maintain 1. Lecture Notes
glutathione in its reduced state. 2. Rodwell, V.W., et al., Harper’s Illustrated
th
Biochemistry, 30 ed.
 An important enzyme in PPP, glucose-6-phosphate
3.
th
Lehinger’s Principle of Biochemistry 5 ed
dehydrogenase, when deficient, is the main cause of
4. 2019 A, B, C Transcriptions
hemolytic anemia.
 The Uronic Acid Pathway provides glucoronic acid NOTES:
needed for conjugation of many endogenous and
exogenous substances as glucuronides, before
excretion in the bile and urine. “The only person you should try to be better than is the person
 Important biomedical application in uronic acid you were yesterday.”
pathway is essential pentosuria, caused by a lack of “Student, you do not study to pass the test. You study to
xylulose reductase. prepare for the day when you are the only thing between a
patient and a grave.” –Mark Reid
 Ascorbic acid cannot be synthesized by mammals
“You are confined only by the walls you build yourself.” –
due to the blockage of the enzyme L-gulonolactone
Andrew Murphy
oxidase; loss of Vitamin C will cause scurvy.
“Be brave and learn to ganern all your pak.” (Someone, 2016)

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APPENDIX

SUMMARY OF THE PENTOSE PHOSPHATE PATHWAY AND ITS RELATION TO GLYCOLYSIS (HARPER’S)

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OVERVIEW OF URONIC ACID PATHWAY (HARPER’S)

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