Preliminary Assessment of the Anticoagulant Potential
of Turnera subulata (Passiﬂoraceae)
´ryo Duarte da Luz,
Thayse Evellyn Silva do Nascimento,
Leandro Vinicius Fernandes de Morais,
Ana Katarina Menezes da Cruz,
Adriana Augusto de Rezende,
Marcela Abbott Galva
Jorge A. Lo
Hugo Alexandre Oliveira Rocha,
and Maria das Grac¸as Almeida
Postgraduation Program in Health Sciences, Health Sciences Center,
Federal University of Rio Grande do Norte, Natal, Brazil.
Multidisciplinary Research Laboratory, DACT, Health Sciences Center,
Federal University of Rio Grande do Norte, Natal, Brazil.
Department of Biochemistry, Biosciences Center, Federal University of Rio Grande do Norte, Natal, Brazil.
Department of Clinical Medicine, Health Sciences Center, Federal University of Rio Grande do Norte, Natal, Brazil.
Tiradentes University/Institute of Technology and Research, Aracaju, Brazil.
ABSTRACT Cardiovascular and thromboembolic disturbances are the main causes of disease-related deaths worldwide.
Regardless of the etiological factors involved in thrombus formation, coagulation is mainly activated by thrombin, one of the
most important blood clotting molecules. Thus, this study evaluated the Turnera subulata leaf crude extract, its ethyl acetate
fraction effect on the coagulation cascade, and its possible side effects. Their phytocomposition indicated polyphenols, mainly
ﬂavonol-3-O-glycosylate and a ﬂavone glycoside, without in vitro and in vivo toxicity. Regarding their potential anticoag-
ulants, results displayed partial thromboplastin and prothrombin time activation, and Xa and IIa, and thrombin inhibition by
heparin II cofactor, indicating signiﬁcant anticoagulant activity, suggesting direct and indirect thrombin inhibition as the main
mechanism of action. Therefore, T. subulata leaf active compounds exhibit therapeutic potential required to develop phy-
totherapeutic formulations to assist conventional anticoagulants in clinical treatments.
KEYWORDS: clinical therapy coagulation cascade plant extract polyphenols
Nowadays, cardiovascular and thromboembolic
disorders are leading causes of death worldwide.
Arterial thrombosis is the most common cause of acute
myocardial infarction, stroke, and ischemia, while deep
venous thrombosis complications include pulmonary
embolism and post-thrombotic syndrome.
coagulation system and its interaction with platelet ag-
gregation are responsible for arterial and venous thrombus
Regular indications for anticoagulant uses include pro-
phylaxis and venous thromboembolism treatments, cardi-
oembolic prevention in patients with cardiac arrhythmia or
mechanical valve prostheses, as well as secondary pre-
vention in patients with acute coronary syndromes or un-
dergoing percutaneous coronary intervention.
point, nonfractionated and low molecular heparins are
used as anticoagulants, even causing side effects, such as
thrombocytopenia and a high risk of systemic bleeding.
This stimulates the search for new substances to aid in
prolonged anticoagulant therapy.
Thus, plant extracts exhibit a proven ability to inhibit the
blood coagulation cascade, especially regarding intrinsic and
extrinsic pathway factors.
In this context, Turnera sub-
ulata (family Passiﬂoraceae) is widely distributed in tropical
and subtropical regions and used in folk medicine
due to its
pharmacological properties, such as anti-inﬂammatory,
and antioxidant properties.
Phytochemical studies with species of this genus revealed the
presence of ﬂavonols, alkaloids, tannins, cyanogenic gly-
cosides, fatty acids, triterpenoids, and various phenolic
compounds related to bioactivities.
Several studies describe the use of the Turneraceae genus
in inﬂammatory processes.
Thus, considering that thrombin
is closely related to coagulation and inﬂammatory systems,
Manuscript received 18 September 2018. Revision accepted 28 January 2019.
Address correspondence to: Maria das Grac¸as Almeida, PhD, Laborato
disciplinar em Pesquisa, Faculdade de Farma
´cia, Universidade Federal do Rio Grande
do Norte, R. Gen. Gustavo Cordeiro de Farias, s/n–Petro
´polis, Natal 59012-570, Brasil,
JOURNAL OF MEDICINAL FOOD
J Med Food 00 (0) 2019, 1–9
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this genus represents a potential reservoir for discovering
compounds for clinical treatment associated with conven-
tional anticoagulants to minimize their side effects. There-
fore, this study evaluated the anticoagulant potential and toxic
and hemorrhagic effects of the T. subulata leaf crude extract
and its fraction of ethyl acetate.
MATERIALS AND METHODS
Plant material and leaf extract preparation
T. subulata leaves, collected in Natal, Rio Grande do
Norte, Brazil, were taxonomically identiﬁed by Dr. Jomar
Gomes Jardim, depositing a voucher specimen (Herbarium
No: 0674/08) at the Herbarium of the Department of Botany
and Zoology, Federal University of Rio Grande do Norte,
Natal, RN, Brazil. Leaves were air-dried at 40C for 48 h
and powdered, before preparing its T. subulata crude extract
(CETS) by ethanol:water (50:50, v/v) maceration for 4 days,
ﬁltered, and then lyophilized.
To characterize CETS-active compounds, an extract
portion, resuspended in methanol, was subjected to liquid–
liquid partition using increasing polarity solvents: n-hexane
(3 ·300 mL) and ethyl acetate (3 ·300 mL), obtaining two
fractions hexanic fraction of T. subulate (HFTS) and
acetate fraction of T. subulate (AFTS). Phenolic compound
contents were determined in all fractions by the Folin–
Ciocalteu method described
(data not shown). No poly-
phenols were detected in HFTS.
High performance liquid chromatography with diode
array chromatographic proﬁle
Chromatographic analyses in triplicates were performed on
a Phenomenex C18 chromatography column (4.6 ·100 mm,
particle size 2.6 lm; Torrance, CA, USA) coupled to the
HPLC system (VARIAN ProStar, Walnut Creek, CA, USA)
equipped with a ProStar 240 quaternary pump, autosampler
(ProStar 410), and a detector (mod. 355 PDA UV/V). CETS
and AFTS (5 mg/mL) were dissolved in methanol.
microliters were injected and elution was conducted at
room temperature at a ﬂow rate of 1.3mL/min, using 0.1%
formic (A phase) and acetonitrile (B phase) in the mobile
phase, under following gradient conditions: 0–3 min, 5% B;
3–7 min, 5–20% B; 7–9 min, 20% B; 9–10 min, 2–23%, B;
10–15 min, 23% B; 15–19min, 23–50% B; and 19–20min,
50–5% B, monitoring at 280 nm. Phenolic compounds were
identiﬁed by comparison with external standards (gallic and
chlorogenic acids, epigallocatechin, rutin, hyperin, quercetin,
apigenin, and kaempferol). All solutions were ﬁltered using a
0.22-lm membrane (Millipore, Billerica, MA, USA).
Three-month-old Wistar rats of both sexes, weighing
250–300 gm, were kept under standard environmental
conditions with food and water ad libitum. All animal pro-
cedures were performed according to the Brazilian National
Health Surveillance Agency (ANVISA),
for Economic Cooperation and Development (OECD)
guidelines, and Protocol No. 035/2015, approved by the
Committee on Ethics in Animal Use, Federal University of
Rio Grande do Norte.
Activated partial thromboplastin time assay
This assay was performed in accordance with the activated
partial thromboplastin time (aPTT) information kit (CLOT
Bios Diagnostica, Sa
˜o Paulo, SP, Brazil), and the coagulation
time was measured in triplicate using a clot timer coagulometer
(Drake Electronica Commerce Ltd., Sa
˜o Paulo, Brazil).
Prothrombin time assay
The prothrombin time (PT) assay was performed accord-
ing to the manufacturer’s instructions (CLOT Bios Diag-
˜o Paulo, SP, Brazil), while the coagulation time
was measured in triplicate using a clot timer coagulometer
(Drake Electronica Commerce Ltd., Sao Paulo, Brazil).
Assay for anti-Xa activity
Regarding anti-Xa activity, the assay was performed on a
96-well microplate using the Biophen Heparin Anti-Xa kit
(Ref: 221010; HYPHEN Biomed, Paris, France) according
to the manufacturer’s instructions. Absorbance was mea-
sured at 405 nm using an Epoch microplate spectropho-
tometer (Epoch BioTek, Winooski, VT, USA).
Direct thrombin inhibition assay (anti-IIa activity)
This assessment was conducted in a 96-well microplate
using the Biophen Heparin Anti-IIa kit (ref: 221025; HY-
PHEN Biomed, Paris, France) according to the manufac-
turer’s instructions, measuring absorbance at 405 nm using a
microplate reader (Epoch BioTek, Winooski, VT, USA).
Indirect thrombin inhibition mediated
by heparin cofactor II
This parameter was determined spectrophotometrically
using standard kit assays, according to Yoon et al.,
suring absorbance at 405 nm using an Epoch microplate
spectrophotometer (Epoch BioTek, Winooski, VT, USA),
using a blank containing all the reagents without the test
Residual hemorrhagic effects
The CETS and AFTS compound residual hemorrhagic
effect was analyzed according to the rat topical scariﬁcation
model described by Brito et al.
After anesthesia with ke-
tamine and xylazine at 1:1 (v/v), an incision was performed
with a surgical blade in the distal tail portion, dipping the
tail in physiological saline to observe bleeding. In the
following stages, the tail was dipped in CETS or AFTS or
heparin solution at 100 lg/mL for 2 min and washed with
saline solution before immersing in a fresh physiological
saline solution for 40 min. Blood was quantiﬁed by the
Drabkin test and the result expressed as the hemoglobin
2LUZ ET AL.
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sum of each tube, subtracting the hemoglobin value ob-
tained before exposure to substance test.
Cytotoxicity by 3-(4,5-dimethylthiazol-2-yl)-2,
5-diphenyltetrazolium bromide assay
Mouse (3T3) and human embryonic epithelial kidney (HEK
293) cells were cultured under standard conditions in Dul-
becco’s modiﬁed Eagle’s medium (DMEM), supplemented
with 10% fetal bovine serum at 37C, 5% CO
, and 95%
humidity. Cells (1 ·10
cells/well) were cultured for 24 h in
96-well microplates to promote adhesion. Thereafter, cells
were exposed in triplicate at different CETS and AFTS con-
centrations (0.1, 1, 10, 100, and 1000 lg/mL) and incubated at
37Cfor24h.Then,100lL of 3-(4,5-dimethylthiazol-2-yl)-
2,5-diphenyltetrazolium bromide (5 mg/mL) dissolved in
DMEM was added to each well and cells incubated for 4 h.
After culture medium removal, 100 lL of dimethyl sulfoxide
was added to each well to assess cell viability at 570nm using
a microplate reader (Epoch BioTek, Winooski, VT, USA).
Acute oral toxicity. The acute oral toxicity was evalu-
ated according to ANVISA
a period of 14 days. Animals randomized into ﬁve groups
(n=5) received, by gavage, doses of 500 and 2000 mg/kg, as
recommended by OECD guidelines.
The control group
received only distilled water. Experimental design for each
group is described as follows: Group 1: normal control rats
received only distilled water (vehicle); Group 2: received a
single dose of 500 mg/kg of CETS; Group 3: received a
single dose of 2000 mg/kg of CETS; Group 4: received a
single dose of 500 mg/kg of T. subulata ethyl acetate frac-
tion (AFTS); and Group 5: received a single dose of
2000 mg/kg of AFTS.
During the ﬁrst 12 h, systematic behavioral observations
were performed (e.g., vocal tremor, piloerection, hyperac-
tivity, tremors, abdominal cramps, diarrhea, and deaths).
Behavioral parameters were evaluated in all animal experi-
mental groups. On the 14th day, animals were euthanized
with thiopental sodium (100mg/kg, i.p.), then laparotomized
for cardiac puncture blood collection and evisceration. The
liver, kidney, spleen, lung, heart, intestine, stomach, esoph-
agus, and brain were removed for macroscopic and relative
Biochemical and hematological parameters
Biochemical parameters (alanine aminotransferase, as-
partate aminotransferase, gamma-glutamyl transferase,
total protein, cholesterol, glucose, urea, creatinine, tri-
glycerides, amylase, and bilirubin) were evaluated by
commercial kits according to respective manufacturers’
instructions (Labtest kits, Lagoa Santa, MG, Brazil) on the
LabMax Plenno automated analyzer (Lagoa Santa, MG,
Brazil). Hematological parameters were determined by
ABX Micros 60 OT Equipment (ABX Diagnostics,
France). Biochemical and hematological parameters were
evaluated in all experimental groups of animals.
Results are expressed as mean –SD, analyzed by a one-
way analysis of variance (ANOVA) and Tukey’s post hoc
test, with GraphPad Prism, version 5.0. Statistical signiﬁ-
cance was considered at P<.05.
CETS and AFT chromatographic analyses revealed the
presence of peaks consistent with reference standards (Fig. 1),
identiﬁed by comparison with ultraviolet spectra (UV)
spectra and retention times (RT) of the two extracts and
external patterns, as depicted in Table 1. Based on this
analysis, rutin (ﬂavonol-3-O-glycosylate) with values of
121.51 and 262.36 lg Eq/g for CETS and AFTS and values
for the apigenin-like compound (ﬂavonoglycoside) corre-
sponding to 80.57 and 252.48 lg Eq/g for CETS and AFTS,
respectively, were identiﬁed with regard to spectral simi-
larity considering RT and UV standards.
CETS and AFTS anticoagulant activity was demonstrated
by activated partial thromboplastin and PT evaluation, such
as activated X and II factors, as well as by indirect thrombin
inhibition mediated by the heparin II cofactor.
CETS, AFTS, and heparin (standard positive control)
revealed a signiﬁcant aPTT anticoagulant activity over 240 s
(negative control: 36.05 –0.03 s) at 5 lg/mL, while PT >60 s
(negative control: 16.65 –0.33 s) was revealed at 30 lg/mL,
as expected and displayed in Figure 2A and B, respectively.
Both CETS and AFTS inhibited clot formation through in-
trinsic and extrinsic pathways in concentrations >100 lg/mL.
Based on these data, the heparin and extract ability to
directly inhibit the activity of factors Xa and IIa (thrombin)
was evaluated. Figure 2C shows that heparin hindered factor
Xa activity in a dose-dependent concentration, requiring
1lg/mL for total factor inhibition, while 100 lg/mL of ex-
tracts completely hampered thrombin activity (Fig. 2D).
Regarding CETS and AFTS, despite their lower activities,
both extracts showed an ability to inhibit factor Xa activity in
a dose-dependent concentration, reaching a value of *40%
and 80% at *100 lg/mL to hinder thrombin activity.
To evaluate the extract’s anticoagulant action mecha-
nisms, an indirect inhibition assay was performed by heparin
cofactor II (HCII) on thrombin. Results displayed a signif-
icant extract capacity (at 100 lg/mL) to inhibit thrombin
(Fig. 2E), revealing that AFTS through HCII showed an
inhibition rate of 70%, whereas CETS showed only 30%.
Heparin’s adverse effects restrict its clinical use, such as
thrombocytopenia and hemorrhagic complications, which
interfere with the hemostatic balance. Considering the rel-
evance of these events, CETS and AFTS effects on hemo-
stasis were investigated.
Concerning the heparin assay (100 lg/mL), results dis-
played a marked hemorrhagic effect, evaluated by the high
residual bleeding level determined using the rat tail scariﬁ-
cation model after hemoglobin dosage in treating animals.
Regarding CETS and AFTS treatments with 100 lg/mL, both
showed anticoagulant potential with lower hemorrhagic rates,
ANTICOAGULANT POTENTIAL OF TURNERA SUBULATA 3
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FIG. 1. HPLC proﬁle of phenolic
compounds from Turnera subulata
leaves detected at 280 nm. Proﬁle
(A) HPLC-UV chromatogram of stan-
dard phenolic compounds, (a) gallic
acid; (b) chlorogenic acid; (c) catechin;
(d) (-)epigallocatechin gallate; (e) rutin;
(f) hyperin; (g) quercetin; (h) apigenin;
and (i) kaempferol. Proﬁle (B) Crude
extract of T. subulata with two major-
itarian peaks: (1) related to ﬂavonol-3-
O-glycosylate such as rutin and (2)
related to ﬂavone glycoside such as
apigenin. Proﬁle (C) Ethyl acetate
fraction of T. subulata with two ma-
joritarian peaks: (1) related to ﬂavonol-
3-O-glycosylate such as rutin and (2)
related to ﬂavone glycoside such as
apigenin. HPLC-UV, high perfor-
mance liquid chromatography coupled
to an ultraviolet detector.
*50% and 10%, respectively, compared with clinical hepa-
rin (Fig. 2F).
With respect to cytotoxicity of CETS and AFTS, no cy-
totoxic effect was evidenced on normal ﬁbroblast (3T3) and
human embryonic kidney (HEK-293) ﬁbroblast cells
(Fig. 3A, B). No statistically signiﬁcant differences were
observed relative to the negative control (DMEM). During
the in vivo toxicity assessment, no animal exhibited be-
havioral abnormalities over the trial period.
However, the biochemical parameter analysis in CETS-
and AFTS-treated animals showed statistically signiﬁcant
differences with regard to glucose, triglyceride, and total
cholesterol levels. These parameters exhibited a mean de-
crease of 50% after treatment compared with the control
group (Table 2).
Regarding hematological parameters and organ weights,
no signiﬁcant differences were observed relative to the
control, indicating no toxicity signs. Water and food intake
displayed no signiﬁcant difference. Data regarding hema-
tological parameters and relative organ weights are attached
as Supplementary Tables S1 and S2.
Although anti-inﬂammatory, antidiabetic, and antiobesity
properties have been reported for the Turnera genus,
data regarding its anticoagulant activity have been described
Overall, plant extracts have been assessed for their ability
to inhibit blood clotting factors.
With regard to Passi-
ﬂoraceae species, studies have shown their potential to treat
cardiovascular diseases. The Passiﬂora nitida Kunth extract
showed anticoagulant activity by partially activating
thromboplastin, suggesting an inhibitory effect on intrinsic
factors of the coagulation pathway VIII, IX, XI, and XII.
Concerning T. subulata, both CETS and AFTS promoted
efﬁcient inhibition of intrinsic and extrinsic pathways of the
blood coagulation cascade. This was indicated by the PT
test, also suggesting an anticoagulant effect due to thrombin
inhibition (common pathway cascade). Therefore, extracts
directly inhibited thrombin, assessed by the factor IIa inhi-
bition assay, and indirectly by HCII as a blood coagulation
These parameters play a key role in coagulation inhibi-
tion. Thus, thrombin, a multifactorial enzyme, acts on the
coagulation system converting soluble plasma ﬁbrinogen
into insoluble ﬁbrin molecules and activating factor VIII.
This factor binds to cross-linked ﬁbrin polymers constituting
a stable clot, amplifying the coagulation cascade with sub-
sequent factors V, VII, VIII, and XI and platelet activation
and stimulating granule release and platelet aggregation.
On the other hand, the long-term anticoagulant prescrip-
tion such as heparin in clinical therapy causes side effects
(e.g., bleeding, thrombocytopenia, hypersensitivity, skin
necrosis, and intestinal toxicity).
This has encouraged
the anticoagulant substance prospection to assist in clinical
treatment. However, research concerning the medicinal
plant anticoagulant effect is scarce, hence the relevance of
Thus, the inhibitory effect of CETS and AFTS on the
blood coagulation cascade may be due to the chemical
composition rich in glycosylated ﬂavonoids, mainly rutin
and apigenin, besides other phenolic compounds. This
phytocomposition is consistent with that previously reported
for Turnera ulmifolia leaf extract, exhibiting signiﬁcant
polyphenolic content, such as rutin and apigenin.
have demonstrated the signiﬁcant anticoagulant effect of
Besides anticoagulant and low hemorrhagic activities,
CETS and AFTS exhibited no toxic effects in vitro using
human ﬁbroblasts and human embryonic kidney cells. An
analogous result was reported with the Turnera diffusa
methanolic extract, exhibiting low cytotoxicity in normal
Concerning the acute oral toxicity, no toxic effects were
observed regarding evaluated biochemical and hematolog-
ical parameters, especially those relating to hepatic and re-
nal functions. There are no data regarding the T. subulata
toxicity potential. However, the low acute oral toxicity level
of T. diffusa Willd corroborates the results of the present
study since no behavioral change, mortality, renal, and he-
patic histopathology, as well as biochemical, parameters
Although AFTS displayed side effects
lower than CETS due to higher polyphenol content, both
extracts can be considered safe for clinical use.
though no acute toxic effects have been observed, anato-
mopathological analyses of different tissues or organs such
as the liver or kidney are required to corroborate the absence
of damage to the liver and/or kidneys. This evaluation will
be carried out in the continuation of the present study.
A relevant aspect regarding the biochemical parameters
evaluated in this study was the decrease in glucose, tri-
glyceride, and total cholesterol values after the CETS and
AFTS treatments. These results suggest hypoglycemic
and hypolipidemic properties, which indicate the phy-
totherapeutic potential of these extracts for cardiovascu-
lar disease treatment. These effects have been reported in
Table 1. Quantiﬁcation Parameters of Nine Phenolic
Compounds Identiﬁed by Comparison Between UV Spectra
and Retention Times of the Extracts and External
Standards Based on the Chromatographic Method
Gallic acid 1.71 1.5–50 271 0.9945
Chlorogenic acid 6.47 1.5–50 326 0.9973
Catechin 7.19 1.5–50 278 0.9993
8.18 1.5–50 275 0.9989
Rutin 9.20 1.81–100 275–354 0.9976
Hyperin 9.24 1.81–100 256–354 0.9995
Quercetin 16.03 1.5–50 255–371 0.9991
Apigenin 18.51 1.5–50 266–337 0.9989
Kaempferol 18.85 1.5–50 263–367 0.9984
UV, ultraviolet spectra.
ANTICOAGULANT POTENTIAL OF TURNERA SUBULATA 5
FIG. 2. Anticoagulant activity of CETS, AFTS, and heparin by activated partial thromboplastin time assay (A), prothrombin time assay (B),
inhibition of factor Xa activity (C), inhibition of thrombin activity (D), indirect thrombin inhibition activity mediated by heparin cofactor II (E),
and bleeding activity of CETS, AFTS, and porcine intestinal mucosa heparin (100 lg/mL) applied topically (F). AFTS, acetate fraction of
T. subulate; CETS, T. subulata crude extract.
several Passiﬂoraceae species, especially in the Turneraceae
for example, in T. diffusa
and T. ulmifolia,
whose extracts were evaluated in animal models.
Overall, results regarding CETS and AFTS anticoagulant,
nontoxic, and low hemorrhagic effects are due to their sig-
niﬁcant polyphenolic compound content. These compounds
are associated with several pharmacological activities
widely described in the literature.
In conclusion, T. subulata CETS and AFTS results dis-
played anticoagulant activity, inhibiting intrinsic and ex-
trinsic pathways in the blood coagulation cascade. This
suggests direct and indirect thrombin inhibition as the main
action mechanism. Moreover, extracts showed low hem-
orrhagic and toxic effects in vitro and in vivo, while the
biochemical parameters showed possible hypoglycemic
and hypolipidemic activities. These biological effects can
be attributed to the signiﬁcant polyphenol content. Overall,
experimental results are promising due to the T. subulata
therapeutic potential to develop herbal formulations to
assist in anticoagulant therapy, although further studies are
required, considering that this is the ﬁrst report evaluating
the anticoagulant capacity of this plant.
This research was supported by the Conselho Nacional de
Desenvolvimento de Cientı
´ﬁco e Tecnolo
(Protocol No.478652/2010-0) and Banco do Nordeste
(Protocol No.912011) grants. The authors would like to
˜o de Aperfeic¸oamento de Pessoal de
FIG. 3. Cell viability (cytotoxicity effects) of CETS and AFTS on mouse ﬁbroblast cells (3T3) (A) and epithelial embryonic human kidney cells
(HEK 293) (B), measured by MTT assays. Culture medium DMEM was used as a negative control of cytotoxicity. Comparisons between groups
were analyzed with an ANOVA and Tukey’s post hoc test. ANOVA, analysis of variance; DMEM, Dulbecco’s modiﬁed Eagle’s medium; MTT,
Table 2. Biochemical Parameters of Rats After 14 Days of Treatment with CETS and AFTS
Biochemical parameters Control 2000 mg/kg CETS 500 mg/kg CETS 2000 mg/kg AFTS 500 mg/kg AFTS 2000 mg/kg
Glic (mg/dL) 122 –21.9 60 –20.1* 61.4 –21.6* 60.1 –19.2* 62 –21.4*
Trig (mg/dL) 53 –5.45 25 –3.89* 28.6 –4.38* 25 –2.32* 28.7 –4.34*
Col (mg/dL) 54.5 –6.05 24.5 –0.24* 22 –0.19* 25.5 –0.19* 21.5 –0.10*
ALT (U/L) 97 –4.24 86.1 –2.30 89 –2.20 89.9 –4.34 97 –5.29
AST (U/L) 250 –23.3 238 –25.3 249 –15.3 228.5 –24.8 240.3 –14.1
c-GT (U/L) 12 –1.4 11 –1.5 10.66 –0.05 12 –0.23 11.5 –0.04
TB (mg/dL) 0.99 –0.01 0.90 –0.01 0.93 –0.01 0.90 –0.01 0.99 –0.02
DB (mg/dL) 0.99 –0.07 0.98 –0.05 0.97 –0.09 0.99 –0.05 0.110 –0.1
IB (mg/dL) 0.28 –0.01 0.13 –0.03 0.12 –0.09 0.15 –0.03 0.15 –0.08
Urea (mg/dL) 54.5 –7.7 54.5 –7.5 57.3 –7.04 55.9 –7.5 58.3 –7.75
Cret (mg/dL) 0.65 –0.07 0.65 –0.07 0.7 –0.03 0.59 –0.19 0.70 –0.07
TP (g/dL) 6.6 –0.1 6.2 –0.5 6.3 –0.64 6.1 –0.5 6.9 –0.6
ALB (g/dL) 3.1 –0.42 3.9 –0.42 2.9 –0.34 3.1 –0.42 3 –0.12
Glo (g/dL) 3.5 –0.42 3.8 –0.35 4.3 –0.54 3.1 –0.32 4.82 –0.54
Ami (U/L) 900 –3.53 900 –3.44 903 –2.09 902.5 –3.43 913.8 –2.07
Results are expressed as mean –SD (n=5); control group treated with vehicle (distilled water). Comparisons between groups were analyzed with an ANOVA and
Tukey’s post hoc test.
*P<.05 compared with the control group.
AFTS, acetate fraction of T. subulate; ALT, alanine aminotransferase enzymes; ALB, albumin; Ami, amylase; ANOVA, analysis of variance; AST, aspartate
aminotransferase; CETS, T. subulata crude extract; Col, cholesterol; Cret, creatinine; DB, direct bilirubin; c-GT, gamma-glutamyl transferase; Glo, globulin; Gli,
glucose; IB, indirect bilirubin; TB, total bilirubin; TP, total proteins; Trig, triglycerides.
ANTICOAGULANT POTENTIAL OF TURNERA SUBULATA 7
´vel Superior (CAPES) and CNPq for providing a post-
graduation fellowship and the Department of Biochemistry
(UFRN) for cell culture technical assistance.
AUTHOR DISCLOSURE STATEMENT
No competing ﬁnancial interests exist.
Supplementary Table S1
Supplementary Table S2
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