Oxalic acid content in green leafy vegetables

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a
I'id1,odus,-aJ.
of Sci
(2011)
Vol. 16, PP 7-17
Oxalic acid content
in green
leafy
vegetables
H. A. A. Premssiri and S. Ekanayuke-
Department of B ioche
mistry
Faculty of Medical Sciences
University of Sri Jayewardenepura
Nugegoda
Sri Lanka
*Corresponding
Author
E mail : s agarikae@
hotmail. com
Received on
: I6-06-201I
Accepted
on : 26-07
-20
I I
Abstract
Oxalic acid is toxic in increased
quantity due to the prornotion
of kidney stone
formation, inhibition of mineral absorption etc. Oxalic acid is a metabolic end
product naturally present in plant materials. Green leaff vegetables being an
essential
part of a Sri Lankan diet often beneficial with high amount of fiber,
B-carotene
and minerals
may
contain oxalic acid. Thus, the objective
ofthe present
study was to determine
the oxalic acid content of some green leaff vegetables
commonly consumed
in Sri Lanka since this data
is not currently
available.
Plant rnaterials
were collected mainly from the Western
province. The edible
portion was homogenized,
extracted and titrated with potassium
permanganate
solution.
According
to the oxalic acid contents
of the plant materials,
the plants
were categorized in to 3 groups. a) Leaves with high oxalic acid content
mukunuwenna white and red stemmed
(245-892 mgl100 g), sarano (creeper
and
plant) 400-1071 mgllOO
g, spinach
[326-563 mgl 100
g], thampala
red and
green
stemmed
1209-947
mgllO0gl; b) Leaves with low oxalic acid (< 200 mg/100g)
content
(passion
fruit [39-62 mgll00g], manioc
red and white stemmed
[75-200
mgllOOgl,
katurumurunga
146-71
mg/100g1,
kankun
120-54
mgl|00gl, kohila [20'
93 mg/l0Ogl,
asamodagarn
[16-78 mgll00g], anguna
leaves
[5-34 mg/I00g] )

Premasiri & Ekanayake
,H,:l *""'iJjlundetectable amount
[<l
4 ms/r00g]
of
oxaric
acid
(gorukora,
The
contribution
of oxalic
acid
to the
diet
by green
reafy
vegetabres
is rerativery
Iow
when
the
edible
portion
is
co'sidered
in ail the
reaves
anaryzed,rn
the
presentstudy' However'
inclusion
of a
variety
of leaves
in the
diet
will be
more
beneficial
instead
of frequent
consumption
of high oxalic containing
leaves
which may
decrease
the
absorption
of minerals
in the
meal. In addition,
people
with kidney
stone
diseases
or
family
history
of the
kidney
stone
diseases
shourd
avoid
frequent
consumption
of high
oxalic
acid
containing
gr."n leafr
vegetabres
such
as
nivithi,
serana,
mukunuwenna
and
thampala
Key words: oxaric
acid,
carcium
oxarate,
green
reafo
vegetabres
fnfuoduction
Green
leafu
vegetables
are
an
essential
part
of a Sri
Lankan
diet
and
are
consumedin many
ways' Addition
of green
leafy
vegetables
is often
beneficial
since
these
contain
high
amounts
of di ehryfiber,
p-carotene,
minerars
and
arso
some
vitamins
(folic acid
and
vitamin
K). However,
these
green
reafi vegetables
Premasiri & Ekanayake
also
contain
some
hannful
substances
like
oxalic
acid
(Sheelaet
a1.,2004)
which
courd
rower
the
bioavairability
of essentiar
nutrients.
oxalic acid
is a low molecular
weight
dicarboxyric
acid.
It occurs
naturary
in a
large
number
of plants'
oxalic acid
is water-sorubre
and
combines
with minerars
such
as
calcium'
magnesium
and
sodium
and
form insoruble
oxarates.
The
rever
of oxalic
acid
in plant
tissues
appears
to increase
with the
age.
This
suggests
that
oxalates
are
an
end
product
of plant
metabolism.
The
oxalic acid,
loxalate
content
vary
in plants
of the
same
'ptti", due
to the
differences
in soil
condition,
crirnate,
:T:iili#::;:'on' state
orripeness
orthe
part
that
is
being
anatyzed(personar

Oxalic
acid
content
in green
lrofy vegetables
The main problem associated
with increased
consumption
of oxalic acid is
insoluble
complex
formation
with important
minerals
such
as calcium,
magnesium
and
sodium
in the diet making these
minerals
unavailable
to the body.
In addition
to oxalic acid absorbed
from the digestive
tract the oxalic acid produced
by the
liver synthesis
and degradation
of ascorbic
acid will be excreted
via the kidney.
Since
calcium-oxalates
are not very soluble
in urine these
tend to form stones
in
the
kidney and
cause
damage
to the tissues.
It is proven
that more
than 10-20%
of
dietary
oxalates
are
excreted
in urine (Holmes
et al., 2001). Intestinal
absorption
of oxalic
acid
occurs
by simple passive
diffusion
and
absorption
rate of oxalic acid is about 22% while the rate of calcium oxalate
absorption
is < 2Yo
from the ingested
amount (Hanes
et al., l99g). The rate of
absorption
decreases
from the colon to the duodenum.
Most of the green leafu
vegetables
are
a good source
of calcium,
but its formation
of complexes
with the
oxalates
present
in the same
plant makes
this important
mineral
unavailable
to the
body (Weaver
and Heaney,
1991).
Kidney stones, a painful and a common disease
condition, are formed by
precipitation
of substances
normally excreted
in urine.
Most stones
are
composed
of calcium oxalate (65%) (Baker, 2002). More than 20Yo
of urinary oxalates
are
derived
from diet (Holmes et al., 2001).
Thus absorption
of dietary
oxalates
may
be an important factor in calcium oxalate stone formation. Hyperoxaluria
may
occur due to excessive
oxalate ingestion
by consuming
green leafu vegetables
(Talbot and Water, 1996). Calcium oxalate stones
have rough surfaces,
which
may prolong in to sharp
spikes
that cause
mucosal
bleeding
(Varley et al., lggl).
The acidity may irritate the tissues
in the digestive
tract. Thus consumption
of
extremely
high doses
of oxalic acid with food sometimes
can be fatal (Sheela
et al., 2004). Oxalic acid produces
hydrogen peroxide in the oxalate oxidase
metabolic
pathway.
This hydrogen peroxide
can promote
a "burst" of phagocytes
which helps
to engulf foreign bodies
like bacteria. Oxalic acid also
decreases
the
bioavailability
of calcium
from the calcium
rich sources.
This may lead
to calcium
deficiency.

Premasiri
& EkanaYake
Thus
the
present
study
was
carried
out
as
an
observational
study
with the
objective
of determining
the oxalic acid content of commonly consumed
green leaty
vegetables.
This
would also
contribr.rte
to some
extent
to food composition
tables
since
data
on
total
oxalic
acid/oxalate
content
of vegetables
consumed
in Sri
Lanka
are
currently
unavailable.
Materials and Methods
Materials
commonly consumed
r4 differenttypes
of green
leaff vegetables
in Sri
La'kawere
analyzed
The edible portions
of these
plants,
as collsumed
by Sri Lankans
were
analyzedto
determine
the
oxalic
acid
content.
Sample
collection
was
random
and,
most
of the
plant
samples
were
collected
from
the
market
places
and
home
gardens
in the
western
province.
Manioc and
nrukutluwenna
samples
were
collected
from
both
wet and
dry zones.
The
plant species,
variety
and
analyzed
edible
portion
are
given in Table
1. All chemical
reagents
used
were
of analytical
grade
purchased
from reputed
chemical
comPanies.
Methods
Moisture content
Moisture
content
ofthe plant samples
was
determined
by drying
in an
oven
(105"C)
until constant
weight.
The weight
Loss
on Drying (LOD) was
collsidered
as
the
moisture
content
of the
plant
samples
(AoAc, 1984a).
Determination of oxalic acid content
Standard
AOAC method
(AOAC, 1984b)
content of plant samples. The method
precipitation
and
quantifi
cation.
was used to analYze
the oxalic acid
includes 3 steps;
sample
extraction,
All the plant samples
were washed
and air-dried.
Edible portion of each
pla't
sample
was chopped
and
weighed
accurately
(a0.00g)
and
transferred
to a high
speed
blender
with distilled
water
(100 mL).
The
blended
sample
was
homogenized
(15
min, 1700
rpm)
and
mixture
cooled
to room
temperature.
The
slurry
(35'00g)
was
accurately
weighed
in to a 500
mL beaker
and
distilled
water
added
to make
a final weight of 3009.
Hydrochloric
acid (6N, 55 mL) and caprylic
alcohol
(2
10

--
e
Oxalic acid content in green leafy vegetables
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Premasiri
& Ekanayake
drops) were added
and the mixture boiled (15 min) in a boiling water bath to
increase
the efficiency
of dissolving. The sample
was carefully transferred
to the
volumetric flask once cool and volume made up to 500 mL with distilled water.
Thus
prepared
samples
were left overnight
at room temperature
(30"C).
On the following day the sample was mixed, filtered and first 100 mL discarded
to minimize contamination.
Precipitation
of oxalic acid was
achieved
by pippeting
the above
filtrate (25 mL) in to a Erlenmeyer
flask. Tungstophosphoric
acid reagent
(5 mL) was added
to puritr the filtrate from any contaminated
metals
which was
then
allowed to stand for 5 hours. After 5 hours
the sample
was
filtered and 20.0 mL of filtrate was pipetted
in to a 50 mL conical
centrifuge
tube and
ammonium
hydroxide
(NI{,OH) added till pH was 4.5. Acetate
buffer
(pH
4.5,5 mL;2.5
g
anhydrous
CaCl, in 50 mL CHTCOOH
and
33
g
NaCIICOOx3H,O
in 50 mL) was
added
to the sample, mixed well and left overnight. The
precipitate
obtained following centrifugation
ofthe sample
(15
minutes,
1700rpm)
was
washed
completely,
breaking
up into fine suspension
with fine jet stream
of filtered cold
wash
liquid (20 mL, saturated).
Centrifugation and decanting
steps were repeated
and the precipitate
was finally dissolved in l\Yo sulfuric acid (5 mL).
Both reagent
blank and sample were heated in a boiling water bath to facilitate
the liberation
of oxalic acid from calcium oxalate
precipitate. Titration of sample
nd blank was carried
out while hot, with 0.01N KMnOo. The endpoint
reading
was
the first pink color in the sample
persisting
for more
than
30 seconds. The mg
oxalic acid /1009 was calculated
using the following equation.
mg oxalic
acid I 1009
product :
(Wet
slurry
x Net wt)
Where
1350 : 0.45
(mg
anhydrous oxalic
acid equivalent
to I mL
0.0lN KMnOo)
x [(30/20)
x (500/25) (dilution
factor)]
x 100
(to
convert
to 100g
product.)
t2

Oxalic acid content in green leafi vegetables
Results
-'ralate
is an
ubiquitous
component
of plants
and hence
an
unavoidable
component
: rur diets. Contribution to dietary oxalate from green lea$ vegetables,
an
:":'ential
component
of a Sri Lankan
diet would be
an
important
'.1--tordeterminingtheavailabilityofminerals.
Inaddition,withtherecommendation
-: hieh dietary fibre intake,
major part of which would be contributed
by green
,i3rr vegetables
for people
with diseased
conditions
like
::rbetes and
hypercholesterolemia,
oxalate
content
may be an important
factor to
;':nsider
in avoiding
long term complications.
In addition
green
leafu
ponidges
re a common
dietary remedy
adopted
by many
diabetics
in our country
which may
i.so contribute
to the oxalate
intake
and further complications
in such
patients.
Table
2.
present
moisture
content
and total
oxalic
acid contents
in some commonly
;'"'nsumed
green
leaff vegetables.
The oxalic acid content
of studied
green
leaves
:rnged from 15-1071
mgll0Og edible portion.
According to the results
of this
.tudy the green
leafli vegetables
were categorized
in to three
groups
according
to
:he
total
oxalic acid content.
High oxalic acid
content
on a wet basis
was seen
in spinach
(326-563
mgl100g),
ntukwtuwenna
white stemmed
variety (437-592
mg/100g),
sarana creepers
(631-
''071mgl100g)
andamaranthus
redvariety (421-947
mgll00g).Thesaranacreeper
had
the highest
oxalic acid
content
followed by red amaranthus
andrnukunuwenna
trvhite).
Some
green
leafu vegetables
(katurumurunga,
white mannioc,
passion
fruit, kohila, kankun, anguna and
asarnodagam)
had low oxalic acid compared
to
tlre above leaves
(< 200 mgllO0g). However,
it was not possible
to detect
very
low amount
(< 14 mgl100g)
of oxalic acid
contents
using
this method.
Gotukola,
lettuce and cabbage
leaves had no detectable
levels of oxalic acid and were
categorized
as
leaves with undetectable
oxalic acid content.
13

sampte Moisture 7o Uxalrc acrd
content
expressed
as
a range
(vnoll OOo\ rvef rrleiohf*
rtants wrtn hlgh oxalic
acid
MuKunuwenna (wet zone, stem recl) 6/fl z4)-) t9
MuKunuwenna (wet zone stem whtte) u7+
l437-892
Mulrunuwenna (dry zone
stem whrte) 87+l 322-6'/E
sarana
(creeper) E6t I oJl tul
sarana $tant) 9Ut l4U0-la4
Nlvlthl (plant) 9'3+'2 326-563
lnampala (green) 88+4 209-4t8
lnampala (red) 63+7 42I-94 t
rlants wrtn low oxalrc aclo
Manloc wet zone
(whrte
stem) 75+2 tU) - ZUU
Manloc wet zone (rect
stem
)u2+5 lu-t9
Manroc Dry zone
(red stem )76+5 4J- l)U
Katurumurunsa 9'2+2 46-71
l(ohlIa u6+5 20-93
Kankun 6 /+2 '/u-54
rasslon U I+J .J9-62
Asamoclagam u6+3 I6-7E
AngunaKola 7E+4 I)-J4
rlants wlth neglrglble oxallc acid
UotuKola u6+3 Undetectable
Lemrce 92+3 Undetectable
uabbage rcaves 6 /+'2 Undetectable
Premasiri & Ekanayake
Thble 2. Moisture
and oxalic
acid mgl100g
wet weight in commonly
consumed
green
leafu
vegetables
(*n:6)
Discussion
When considering
the above results, a significant variation in the oxalic acid
content
among the different types
of green
leafu vegetables was observed. lt was
also apparent
that low oxalic acid containing
green
leafy vegetables
of the same
t4

Oralic
acid
content
in
green
ltqfy vegetables
i.*; rss
rbrained
fi-om
differe't areas
showed
a
lower
variation
in total
acid
content
.::ie I r. In
contrast.
plants
which
had
high
oxalic
acid
contents
indicated
a
high
i -.irion
in the
acid
cont
ent
(mukunuwenna,
nivithi,
sarana and
thampala)' The
:r..€a e,i high
r.ariation
in the same
species
courd
be due
to biological
variation,
:-,.a:urin
and'or
genetic
factors.
In addition,
the
ammonia
to nitrate
ratio
of the
soil
*.l. :.-'siderably,
affect
the oxaric
acid
content. If the nitrate
content
(fertilized)
* :.rl'^
the
oxalic
acid
content
of the
plantwould
be
high
and
it is recommended
:3: \\ hen
cultir
ating
green
leaff vegetables
that
fenilizer
rich
in ammonia
be
used
Pr.rnissami.2002). However,
we were
unable
to see
such
a difference
in the
:c:r.ized and unfeftilized
katurumurutxga
leaves
grown in a home garden'
The
:.lii \ariatio. i' oralic acid seen
in the commercially
cultivated
plants
such
as
n:t;:untnt'etilla.
sarano and
nivithi may
be due
to the application
of fertilizer in
i-':re instances.
to obtain
a quick
harvest'
',\ :sn the oxalic acid contents
in the white and red stemmed
manioc leaves
wefe
::;isidered,
the r,vhite
variety
had
higher
oxalic acid
content
than
the red
variety'
Ii:.. :ame
r.vas
observed
in the
white and
red mukunuv'enna
samples'
In the
present
s:-:d.r
\\
e
were
unable
to see
a
significant
variation
between
the
oxalic
acid
contents
:i marrioc and
nrukunuwenna
leaves
obtained
from western
province
and some
]:easindryZone(p:0'74)'However,beforeadefiniteconclusionisreached
:ttrt€ sarnples
from both
places
should
be
analyzed'
..r1
sfi"atlaleaves
the
content
of oxalic
acid
in the
creeper
was
much
higher
than
the
.'lralic
acid
content
in the plant
(Table
2)' Depending
on the place
of cultivation
rire
creeper
extracting
more
nitrate/oxaric
acid
from soil than
the
plant
could
be
the
reasolt
for high
oxalic
acid
content'
According
to the
riterature
the
percentage
absorption
of oxalic
acid
in the gutts22
, o
(Hanes
et
al., lggg).lf
the
daily
intake
and
absorption
is
considered,
consurnption
of 20 g of prep
ated
sarana leaves(wet
weight)
would result
in an
intake
of 33 mg
oxalic
acid.
The
intake
would be
in the
tolerable
region
even
with
sarana
which is
the
highest
oxalic
acid
containing
green
leaff vegetable
from the leaves
analyzed
in the present
study. Hence,
the other green
leaff vegetables
would have lower
oralate than
sarana intrre
edible
portion.
Therefore,
the contribution
to oxalic acid
15

Premasiri
& EkanaYake
in the diet by green
leafu
vegetables
would be relatively low'
However,
in a meal this is not the onry source
of oxarate.
Thus, the consumption
of high oxalic acid containing leaves
daily with lower intake of water may lead
to accumulation
of oxalic acid in the body.
The fractionar
excretion
of oxalate
is
reported
to be (f%higher in stone
formers
compared
to nonnal i'dividuals. Thus
the transient
renal
oxalate
load following ingestion
of oxalate
rich meal
may lead
to cerl injury in the proximal tubule (Holes et ar.,
2001) in susceptible
individuals
and
in addition
decrease
the mineral
absorption'
In concrusion,
though
addition
of green
reafy
vegetables
in the
diet
has
many
health
benefits,
variation of the leaf types included in the diet may be more beneficial
for hearth
than frequent
consumption
of high oxalic containing reaves
that may
decrease
the absorption
of minerals
in the meal' In addition' people
with kidney
stone
diseases
or family history of kidney stone
diseases
should avoid frequent
consumption
of high oxalic acid containing
green
leaff vegetables
such
as
nivithi'
s
ar
anu, mulu,muw
enna and
thampal
a'
rhe authors
are
grateful
," -IHHlt:Hilelped to collect
samples
from
different Parts
of the country'
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