1

INTRODUCTION
1. The urgency of the study
Gout is a common disease, due to the deposition of monosodium
urate (MSU) crystals in synovial fluid or tissues. Between 1 - 4% of
the adult population worldwide has gout. In developed countries, the
incidence of gout in men is between 3 - 6% of the population, and
between 1 - 2% of the population. In some developing countries, the
incidence of gout may increase to 10%. Gout usually develops in
stages. About 25% of gout patients have joint damage, irreversible
joint deformity. The quality of mental and physical life of gout
patients decreased, due to the frequent affects of acute gout attacks,
the number of swollen, painful joints increased over time, the
progression and spread of tophus, using colchicin, corticosteroids...
To research into new herbal medicine for gout treatment that can
limit unwanted effects is meaningful and necessary. Therefore, over
the past decade, scientists have studied a variety of herbal and
traditional medicines for the gout treatment. Vietnam traditional
medicine has many effective formulas in treating gout, Si Miao Yan
formula is one of them. The composition of Si Miao Yan formula
consists of Semen Coicis, Phellodendron sp Rutaceae, Achyranthes
bidenta, Rhizoma Atractylodis. Some experimental studies have
shown that polysaccharides in Rhizoma Atractylodis and baicalein,
oroxylin A in Cortex Oroxyli (replaced Phellodendron sp Rutaceae)
have anti-inflammatory effect. Stigmasterol and p-coumaric acid in
Semen Coicis are involved in the elimination of uric acid. We
adjusted the dosage, the drug mix, the drug form then evaluated the
effects of Tu dieu tan granule on two objectives:
2. Study objectives
1.

activities function according to VAS2, VAS3. Improving median
function assessed by HAQ was 0,96 ± 0,32 point; Its efficacy of
improving traditional symptoms was 98,33%. The improvement of
these indicators in the study group were statistically significantly
higher compared to the control group (p < 0,05). Tu dieu tan granule
had effect in treating both the wind damp heat impediment pattern
and the phlegm blood stagnation pattern with SUA levels decreased
222,26 ± 114,84μmol/l; 194,97 ± 96,69μmol/l and Nimodiping score
improvement was 10,92 ± 2,31 points; 10,62 ± 2,83 points; the
difference between two patterns was not statistically significantly. No
adverse effects of Tu dieu tan granule in clinical or subclinical were
observed. These findings are scientific indications of the safety, antiinflammatory and analgesic, hypouricemic effects of Tu dieu tan
granule in treating chronic gout.
4.Thesis structure. Apart from the Introduction and Conclusions
chapters, the thesis consists of four chapters.
Chapter 1: Literature review
35 pages
Chapter 2: Study subjects and methods
25 pages
Chapter 3: Results
37 pages
Chapter 4: Discussion
34 pages
And 38 tables, 6 graphs, 5 images, 13 appendixes, and 151
references (34 Vietnamese references, 93 English references, 24
Chinese references).


3
Chapter 1

alkalinization (sodium bicarbonate solution) may be considered.
1.2. The concept of gout according to traditional medicine
1.2.1. Cause and pathogenesis
According to traditional medicine, gout has the name is “Tong Feng”.
Tong Feng is used to refer the pain causes by wind. Gout is cateogrised as
“Tong bi” according to traditional medicine (Bi means standstill, not
cleared). The causes of “Tong bi” include six external climatic factors,
seven emotional factors, and other pathogenic factors.


4
1.2.2. Treatment of gout
Depending on the clinical types, there are appropriate treatments
principles and specific formulas. According to Hao Xian Gou and Jie
Tian Ping – China (2008), Tong Feng is divided into five patterns:
Wind cold damp impediment pattern, wind damp heat impediment
pattern, phlegm blood stagnation pattern, spleen and kidney yang
deficiency pattern, liver and kidney yin deficiency pattern. For the
wind cold damp impediment pattern, the principles include of
dispelling wind, scattering cold, eliminating dampness, circulating
meridians; the appropriate herbal formula are Tigao Tai Tang or
Fangjian Fengge Tang. For the wind damp heat impediment pattern,
the principles include of dispelling wind, clearing heat, eliminating
dampness; the appropriate herbal formula are Er Miao San combined
with Bai Hu Gui Zhi Tang; for the damp heat pattern include
numbness, the principles include of clearing heat, eliminating
dampness, circulating meridians, analgesic, the appropriate herbal
formula are Si Miao San combined with Long Dan Da Gan Tang. For
the phlegm blood stagnation pattern, the principles include of
circulating blood, dispelling stasis, transforming phlegm; the

anakinra) on anti-inflammatory and analgesic effects in gout flares. In
Vietnam, there has been no research on new medicine or gout treatment
medication in the last decade. Most studies of Western medicine focused
on the medicine abuse (corticosteroid). Some studies assessed the
knowledge of patients in preventing and using gout medicines.
1.2.2. Review on gout treatment studies according
to traditional medicine
In experimental, many models were built up to evaluate the
effectiveness of gout medication on anti-inflammatory, analgesic, and
lowering SUA level such as: Xanthin oxidase inhibatory in vitro and in
vivo models, rat arthritis after intrasynovial injection of sodium urate,
evaluating hypouricemic effects in potassium oxonat induced
hyperuricemia in rat... Shen Wei Zeng, Pang Xue Feng used rat arthritis
after intrasynovial injection of sodium urate in evaluating antiinflammatory and analgesic effects of Dang Gui Nian Tong Tang, Zhi
Tong Qu Feng Tang. In clinical, many scientists have built up and
studied the effectiveness of new herbal formulas such as: HA1 herbal
formula, GLP hypouricaemia... Other authors have investigated the
association between Western medicine and traditional medicine in order
to have higher therapeutic efficacy than either Western medicine or
traditional medicine. All researchs have the same procedure is controlled
trial, comparison study (Studying on the effectiveness of Tong Feng
Wan, or the combined of Narcaricin and Si Wu Tang).
1.3. Review of Tu dieu tan granule
1.3.1. Origin and traditional efficacy
Tu dieu tan granule is derived from the traditional medicine Si Miao
in Fang Ji Xue book, with changing doses and replacing Phellodendron
sp Rutaceae and Achyranthes bidenta with Cortex Oroxyli and Radix
Achyranthes asperae. The increased dosage of Tu dieu tan granule was



- Rhizoma Atractylodis has effective on scattering cold and
circulating cuticle, tonifying spleen qi, detoxifying, eliminating
phlegm. Chemical composition: Mostly are hinesol or β-eudesmol,
atractylon. Polysaccharide has effective on regulating the intestine
immune system against candida albicans. β-eudesmol and atractylon
protect the liver cells.


7
Chapter 2
STUDY MATERIALS, SUBJECTS AND METHODS
2.1. Study materials
2.1.1. Baseline medicines
- Hypouricemic medicine: Allopurinol oral tablet contains 300mg
allopurinol, excipients just enough. This product is distributed by
Domesco Medical Import Export Joint Stock Corporation.
- NSAIDs: Mofen oral tablet contains 400mg ibuprofen, excipients
just enough. This product is distributed by Medopharm – India.
2.1.2. Studied medicines
- Tu dieu tan granule 7,5g with the composition of 1,13g dried extract
Radix Achyranthes asperae; 0,56g dried extract Cortex Oroxyli; 1,5g
dried extract Semen Coicis; 0,56g dried extract Rhizoma Atractylodis;
excipients enough for 7,5g. Tu dieu tan granule is produced by
Department of Pharmacy/Military Institute of Traditional Medicine
(MITM), has been granted certification basic-level standard.
- Placebo: Presenting format like Tu dieu tan granule. One package
with the composition of 7,5g lactose, excipients enough. Placebo is
produced by Department of Pharmacy/Military Institute of Traditional
Medicine.
2.2. Study subjects

controlled study with the following stages.
2.3.1.1. Investigating the acute and sub-chronic toxicity effects
* Acute toxicity: was assessed in white mice using Litchfield Wilcoxon method based on OECD and WHO guidelines. The mice
received the medication under investigation with increasing doses
through oral use. The aim was to identify the non-lethal highest dose
(0%), the lethal lowest dose (100%) and the intermediate doses. The
mice were observed for 72 hours to monitor their mortality rate and
overall health condition; and were observed for 7 days since
medication administration to monitor mortality rate.
* Sub-chronic toxicity: According to WHO guideline, Wistar rat
were devided into 3 groups: Controll group (distilled water);
Treatment group 1 (Tu dieu tan granule 1,8g/kg/day - corresponding
to the clinical dose); Treatment group 2 (Tu dieu tan granule
5,4g/kg/day - three times higher than the clinical dose). Rats were
drunk water and Tu dieu tan granule for 8 weeks continuously. The
rats were observed to monitor their weight, eating, sleeping, activity,
digestion, hematology, biochemistry, liver and kidney function, liver
and kidney histopathology. Comparisons were made between preand post-treatment, and between the treatment and control groups.
2.3.1.2. Acute anti-inflammatory effect
Evaluating anti-inflamatory effect in experimental models such as the
carrageenin-induced rat paw oedema, rat peritonitis inflammatory, rat
arthritis after intrasynovial injection of sodium urate. The carrageenininduced rat paw oedema model was conducted according to Winter
method. The rat peritonitis inflammatory model was conducted according
to Patel method. The rat arthritis after intrasynovial injection of sodium
urate model was conducted according to Faires and McCarty method.
2.3.1.3. Analgesic effect


9
Evaluating analgesic effect in experimental models such as the

patients. The medication under investigation and the placebo were
presented in the same format.
During 30 treatment days, patients in the study group used
allopurinol and Tu dieu tan granule, patients in the control group used
allopurinol and placebo. Patient was treated on the first day of the
study (D0): Allopurinol (01 tablet taken once per day, after meal); Tu
dieu tan granule (1 package per time taken twice per day, 1 hour
before meal); placebo (1 package per time taken twice per day, 1 hour
before meal). During the study, patients did not use other medicines.
In case of patient with gout flare was painful and ask for analgesic


10
medicine, he/she would be allowed to use Mofen 400mg in 5 days
maximum (01 tablet per time, taken twice or three times per day).
The patients’ identities were only revealed after the course of
treatment was completed, then data collection, analysis and report
were carried out.
2.3.2.2. Study indicators and evaluation methods
- Describe characteristics of the study subjects: Age, gender,
historical symptoms (disease level, risk factors, traditional patterns).
- Clinical indicators: Study indicators was used to evaluate the
effectiveness of Tu dieu tan granule in clinical and subclinical and
compare the effectiveness between the control and study group, preand post-treatment. These indicators were assessed in the first day
(D0) and the 30th day (D30) of the treatment course: Anti-inflammatoy
effect (reduced the number of swollen joints); analgesic effect
(reduced the number of painful joints, improved VAS 1 scores);
improved active function (improved VAS2, VAS3, HAQ scores);
improved traditional symptoms according to Nimodiping score.
Evaluating the treatment effective in sub-clinical (D0, D30); evaluating

the treatment did not change the rats’ liver and kidney function
indicated by biochemiscal as well as histopathological tests. Tu dieu
tan granule at the dose of 5,4g/kg after 8 weeks of continuous
administration showed the changes in hematological indicators but
still in normal standards.
3.1.2. Acute anti-inflammatory effects
* The carrageenin-induced rat paw oedema model: Tu dieu tan
granule at the dose of 1,8g/kg and 5,4g/kg had no anti-inflammatory
effect at all times after causing inflammation.
* The rat peritonitis inflammatory model
Table 3.1. Acute anti-inflammatory effects of Tu dieu tan granule on
the volume and components of rat peritonitis inflammatory
Tu dieu tan Tu dieu tan
Biological
Aspirin
control
granule
granule
Indicators
200mg/kg
group
(n = 10)

(n = 10)

1,8g/kg
(n = 10)

5,4g/kg
(n = 10)


** p ≤ 0,01 and *** p ≤ 0,001 compared to control group
The results showed that Tu dieu tan granule at the two doses
reduced the volume and protein content of rat peritonitis
inflammatory statistically significantly compared to the biological


12
control group (p < 0,001), reduced the number of white blood cells (p
< 0,01); the effect was equivalent to aspirin 200mg/kg.
* The rat arthritis after intrasynovial injection of sodium urate
- The effects of Tu dieu tan granule on the inflammatory levels, size
and temperature of the knee joint: Tu dieu tan granule at the dose of
2,1g/kg had effect on reducing knee oedema, synovial fluid, lowering
the temperature of the inflammatory site, reducing the infiltration of
inflammatory cells after 6 hrs and 24 hours. The difference was
statistically significantly compared to the biological group (p < 0,05).
- Knee pathology in rats treated Tu dieu tan granule: The synovial
fluid has a sloughed area which is easy to observe. The synovial fluid
had less oedema fluid, urate crystals and infiltration of inflammed
cells than the biological control group.

1

2

1

2
(HE x 40)

Induction control group
allopurinol 20mg/kg group

10
10

Tu dieu tan granule 3,6g/kg group

10

Tu dieu tan granule 10,8g/kg
group

10

The reduction
compaired to
control group

Uric acid
(mmol/L)
168,90 ± 33,59
39,40 ± 4,43∆∆∆
54,80 ±
10,04∆∆∆
50,80 ±
16,96∆∆∆

76,67%
67,55%


9.4

Group
Induction
group

Tu dieu tan
3,6g/kg

Tu dieu tan
10,8g/kg

Chart 3.1. The effects of Tu dieu tan granule on urinary serum uric
acid level in mice
Lô nghiên cứu


14
Tu dieu tan granule at the dose of 3,6g/kg and 10,8g/kg increased
uric acid excretion in mice, the difference was statistically
significantly compared to the biological control group (p < 0,05).
3.2. Results of clinical studies
3.2.1. Characteristics of study subjects
A comparison between the study group and the control group
regarding age, sex, occupation, disease severity indicated (number of
swollen and painful joints, average duration of chronic gout, VAS1,
VAS2, VAS3, HAQ scores) at D0 showed no significant difference (p
> 0,05).
3.2.2. Results of supporting gout treatment in clinical and

> 0,05

D30
p (D0 - D30)

< 0,001

< 0,001

After treatment, a statistically significant decrease in the average
number of swollen joints was found in both groups (p < 0,001). At D30,
the reductions in the study group was more pronounced than in the
control group (p < 0,05).
* Analgesic effects
Table 3.4. Improvement in the pain intensity assessed by VAS1
Study group
Control group p
VAS1 index (point)
study-control
D0
D30
Average
improvement D0 D30

± SD (n = 60)

± SD (n = 60)

6,90 ± 0,82
2,19 ± 1,10

± SD (n = 60)
D30
VAS2 index (point)
4,56 ± 1,53
2,97 ± 1,12
< 0,001
VAS3 index (point)
4,58 ± 1,25
2,68 ± 1,14
< 0,001
HAQ index (point)
- 0,96 ± 0,32
- 0,42 ± 0,22
< 0,001
After treatment, both two groups had statistically significant
improvement in physical activities fucntion according to VAS2, VAS3,
HAQ scores better than pre-treatment (p < 0,001). The improvement
in the study group being more pronounced than the control group (p
< 0,001).
* Hypouricemic effects
Table 3.6. SUA levels changed in the two groups pre- and posttreatment
SUA levels
(μmol/l)

Study group
± SD (n = 60)

Control group
± SD (n = 60)


After treatment, both two groups had statistically significant
decreased in SUA levels more than pre-treatment (p < 0,001); the
reduction in the study group being more pronounced than the control
group (p < 0,001).


16
The percentage of patients in the study group who had the
hypouricemic treatment efficacy was 98,33%, higher than the control
group (71,67%), the effectiveness in the study group being more
pronounced than in the control group (p < 0,001).
3.2.2.2. Supporting gout treatment in clinical and subclinical of
Tu dieu tan granule according to traditional medicine
* The effect of Tu dieu tan granule on Tong Feng treatment
according to traditional medicine: Patient was assessed the Tong Feng
treatment efficacy according to Nimodiping score. After treatment, the
percentage of patients in the study group had completed and
pronounced results (98,33%) higher than the control group (15%), the
difference was statistically significantly (p < 0,001).
* The effect of Tu dieu tan granule on Tong Feng treatment in
two traditional patterns
Table 3.7. Changes in treatment evaluation indicators of two
traditional patterns in the study group
Average
improvement D0 D30
Nimodiping index
(point)
Number of swollen
joints (joint)
Number of painful

> 0,05

0,42 ± 0,52

0,44 ± 0,58

> 0,05

> 0,05
2,52 ± 0,97
2,83 ± 0,80
> 0,05
4,00 ± 1,65
4,7 ± 1,48
> 0,05
4,08 ± 1,08
4,71 ± 1,27
> 0,05
0,88 ± 0,22
1,03 ± 0,32
222,26 ± 114,84
194,97 ± 96,69 > 0,05
After treatment, the study indicators were used to evaluate the
anti-inflammatory, analgesic, hypouricemic effects, Nimodiping
score in both two traditional patterns improved better than pretreatment, the difference between two patterns was not statistically
significantly (p > 0,05).
3.3. Adverse effects


17

liver and kidney function indicated by biochemiscal as well as
histopathological tests. Tu dieu tan granule at the dose of 5,4g/kg
after 8 weeks of continuous administration showed the changes in
hematological indicators but still in normal standards. The bias of
reducing number of red blood cells, hemoglobin levels, and
hematocrit may be due to the action of saponin in Radix Achyranthes


18
asperae. Saponin used in high dose for long time can rupture the red
blood cells, thus affects hemoglobin level and hematocrit. This
indicator needs to be monitored in clinical studies.
4.1.2. Acute anti-inflammatory effects
* The carrageenin-induced rat paw oedema model: The results
showed that Tu dieu tan granule at the dose of 1,8g/kg and 5,4g/kg
had no anti-inflammatory effect at all times after causing
inflammation. This finding is not consistent with the previous studies
on the anti-inflammatory effects of Rhizoma Atractylodis and Radix
Achyranthes asperae on the carrageenin-induced rat paw oedema
model. Thus, we continue to conduct the rat peritonitis inflammatory
model to evaluate the mechanism of Tu dieu tan granule.
* The rat peritonitis inflammatory model: The results on table 3.1
showed that Tu dieu tan granule at both two doses reduced the volume
and protein content of rat peritonitis inflammatory statistically
significantly compared to the biological control group (p < 0,001),
reduced the number of white blood cells (p < 0,01). This result is
consistent with some studies on the anti-inflammatory effect of Radix
Achyranthes asperae, Rhizoma Atractylodis, Cortex Oroxyli, which
were found in Tu dieu tan granule component. Nguyen Huong Giang
(2014) found that the hydroxyl at position 2 of benzoxazinone in

stimulation in rat by the Thermal Plantar Test Instrument showed that
Tu dieu tan granule at the dose of 3,6g/kg and 10,8g/kg had effects on
prolonging reaction time for temperature, increasing the endurance for
painful force, increasing the response time for painful stimulation, the
difference was statistically significantly compared to the biological control
group and pre-treatment (p < 0,05). The response time for painful
stimulation was prolonged shows the analgesic effects of Tu dieu tan
granule.
This effect may be due to β-eudesmol in Rhizoma Atractylodis
and other substances in Radix Achyranthes asperae. β-eudesmol has
analgesic effect in the hot plate model and the acetic acid writhing
and colorectal distention models.
4.1.4. Hypouricemic effects
* The potassium oxonat induced hyperuricemia in rat
The results on table 3.2 showed that Tu dieu tan granule at the dose of
3,6g/kg and 10,8g/kg had hypouricemic effects, the difference was
statistically significantly compared to the induction control group (p

VAS3, HAQ scores) were comparable between the study group and
control group. This is a major criterion in controlled experimental
studies to ensure the study objectivity.
4.2.2. Results of supporting gout treatment in clinical and
subclinical


21
4.2.2.1. Supporting gout treatment in clinical and subclinical of
Tu dieu tan granule according to Western medicine
* Acute anti-inflammatory effects: The results on table 3.3 showed
that all patients had the reductions of swollen joints. At D30, the
reductions in the study group was more pronounced than in the
control group (p < 0,05). Thus, Tu dieu tan granule had antiinflammatory effect in clinical. These effects were demonstrated in
experimental models. Capably, saponin in Radix Achyranthes
asperae, polysaccharide in Rhizoma Atractylodis and baicalein,
oroxylin A in Cortex Oroxyli involved in inhibiting gouty arthritis
with different mechanisms.
* Analgesic effects: The results showed that Tu dieu tan granule
improved the average number of painful joints at D30 and improved
the pain intensity assessed by VAS 1 higher than the control group and
pre-treatment, the difference was statistically significantly (p < 0,05).
According to traditional theory, the painful on the lower limb occurs
when the climate changes, which is related to the wind damp
stagnation factors. Semen Coicis has effect on tonifying spleen qi,
diuretic, clearing heat, analgesic which is the king ingredients of Tu
dieu tan granule. Radix Achyranthes asperae supports Semen Coicis
on diuretic, eliminating dampness, clearing heat, detoxifying that
treats the numbness pattern with painful joints, difficulty in
movement. Rhizoma Atractylodis has effect on tonifying spleen qi,

with the aim to qualitative quantification of traditional symptoms in order
to diagnose the pattern exactly and improve the clinical efficacy. After
treatment, the percentage of patients in the study group had completed and
pronounced results (98,33%) higher than control group (15%), the
difference was statistically significantly (p < 0,001).
The symptoms made patient tired were the feeling of thirsty, heat
inside heart, anorexia, full stomach, difficulty in sleeping, heavy in
the lower limbs, body aches. Improving the symptoms of Tong Feng
will improve the quality of life, increase the quality and effectiveness
on treatment, while minimizing the unexpected effects.
The results on table 3.7 showed that after treatment, the study
indicators were used to evaluate the anti-inflammatoy, analgesic,
hypouricemic effects, Nimodiping score in both two traditional
patterns improved better than pre-treatment, the difference between
two patterns was not statistically significantly (p > 0,05). Tu dieu tan
granule has the same effects on the wind damp heat impediment
pattern and phlegm blood stagnation pattern.
4.2.3. Adverse effects
* In clinical: During treatment, the percentage of patients in the
control group had unexpected symptoms such as digestive disorders
were 5% (3/60), itchy rash was 1,67% (1/60). Patients with
gastrointestinal disorders were treated with Berberin, an itchy rash
was treated with anti-histamin. Patients were stability after taking the


23
medications and continued to participate in the study. Patients in the
study group did not have unexpected symptoms.
Thus, the combination of Tu dieu tan granule and allopurinol did not
cause adverse effects. In our view, the unexpected effects in the control

- Tu dieu tan granule had anti-inflammatory effects in the rat
peritonitis inflammatory, the rat arthritis after intrasynovial
injection of sodium urate models.


24
-

Tu dieu tan granule had analgesic effects in the hot plate model,
the hyperalgesia to thermal stimulation in rat by the Thermal
Plantar Test Instrument.
- Tu dieu tan granule had hypouricemic effect in the potassium oxonat
induced hyperuricemia in rat. Tu dieu tan granule had hypouricemic
effect by excreting uric acid through the urinary system.
2. Tu dieu tan granule had supported the chronic gout treatment,
had effect in treating two patterns of Tong Feng. Initially, no
adverse effects of Tu dieu tan granule on clinical were observed.
2.1. Supporting the chronic gout treatment
- The hypouricemic effects: In the study group (the combined
treatment of Tu dieu tan granule with allopurinol): SUA levels
decreased 200,42 ± 100,14μmol/l after 30 days of treatment. The
effectiveness of lowering urate decreased by 98,33%.
- The anti-inflammatory and analgesic effects: In the study group, the
number of swollen, painful joints decreased after treatment. Mean
pain intensity on VAS1 scale, physical activities function according to
VAS2, VAS3 scale improved higher than pre-treatment. Median
function assessed by HAQ score was improved 0,96 ± 0,32 point.
- The improvement of traditional symptoms: Its efficacy of
improving traditional symptoms was 98,33%.
- The improvement of these indicators in the study group were