Therapeutic Effects of Compound Zhenzhu Tiaozhi Capsules in Nonalcoholic Fatty Liver disease-a Randomized Controlled Study
Phase: NA
Conditions: Nonalcoholic Fatty Liver Disease
Sponsor: The First Affiliated Hospital of Guangdong Pharmaceutical University

 
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Therapeutic effects of Compound 
Zhenzhu Tiaozhi capsules in 
nonalcoholic fatty liver disease-a 
randomized controlled study 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Date: November 11, 2017  
Version: 1.0  
Institution: First Affiliated Hospital of Guangdong Pharmaceutical 
University 

 
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I. Background 
Nonalcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver 
disease in China now. It comprises a wide spectrum of disorders that range from 
simple steatosis to nonalcoholic steatohepatitis (NASH), advanced fibrosis, related 
cirrhosis, and even hepatocellular carcinoma (HCC). Currently there are no effective 
medications to cure NAFLD. Simple steatosis can progress into NASH, irreversible 
cirrhosis, and even HCC. Recently, extensive studies have emphasized that NAFLD is 
closely associated with metabolic syndrome, type 2 diabetes mellitus, dyslipidemia, 
insulin resistance, and oxidative stress/lipid peroxidation injury. In addition to the 
liver pathogenesis, both simple steatosis and NASH are well-established risk factors 
for many diseases, including diabetes, arteriosclerosis, hypertension, and colorectal 
cancer. Therefore, it is urgent to develop effective therapies for the early stages of 
NALFD and thereby reduce the risk and morbidity of cirrhosis, HCC, diabetes, 
hypertension, colorectal cancer, and other serious diseases. Current treatments for 
NAFLD include weight reduction through lifestyle modification and exercise, 
insulin-sensitizing agents, lipid-lowering agents, and hepatoprotective drugs. (Cited 
from: Hu Yinxia, Li Lan, Yuan Yu, Wu Lihao, He Xingxiang. Therapeutic effect of 
teduglutide on non-alcoholic fatty liver disease in rats[J]. World Chinese Journal of 
Digestology, 2016 (7): 1009-1016.). However, the efficacy of these therapies is not 
satisfactory, some drugs may even induce liver toxicity. Thus, it is necessary to 
develop novel therapies that are effective and safe for the treatment of NAFLD. 
 
The pathogenesis of NAFLD is complicated, which is mainly based on a 2-hit 
hypothesis, first proposed by Day and James in 1998. Essentially, the “first hit” refers 
to the development of hepatic steatosis via hepatic triglyceride accumulation and 
insulin resistance. The “second hit” is the shift from hepatic steatosis to NASH, 
including oxidative stress, inflammatory cytokines, and chronic intermittent hypoxia. 
Insulin resistance is the critical factor for hepatic steatosis. Insulin resistance is an 
abnormal metabolic state, in which peripheral tissue has impaired sensitivity to insulin 

 
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and attenuated biological response. In the stage of simple steatosis, improvement of 
insulin resistance could prevent the development of hepatic steatosis, reduce the 
accumulation of hepatic lipid, and effectively preclude the emergence of second hits. 
The liver and intestine are functionally connected in several ways. The interactions 
between the liver and the gut, called the "gut-liver axis," has been a gradually 
accepted theory and provides a powerful rationale for the treatment of many chronic 
liver diseases. Both animal experiments and clinical studies have demonstrated that 
the intestinal microflora are closely associated with the onset and progression of 
chronic liver diseases. Liver diseases can cause increased intestinal permeability, in 
which widened tight gap junctions lead to microbial translocation. On the other hand, 
microbial translocation and dysbiosis in the gut can further promote the development 
and progression of liver diseases. (Cited from: Chen Meihui, He Xingxiang, CaiJiyi. 
Progress on liver dysfunction and small intestinal diseases[J].Journal of Guangdong 
Pharmaceutical University, 2015, 31 (4): 558-560). A previous study found a 
NASH-like pattern of liver injury in a mouse model of small intestinal bacterial 
overgrowth (SIBO). Furthermore, the authors observed that SIBO mice had a higher 
incidence of NAFLD, which was related to hepatic steatosis. In the congenitally obese 
mice model, the intestinal mucosal barrier was damaged, which caused endotoxemia 
and led to the development of NASH via TNF-alpha secretion (cited from: Vanni E, 
Bugianesi E. The gut-liver axis in nonalcoholic fatty liver disease: Another pathway 
to insulin resistance, Hepatology, 2009, 49 (6): 1790-1792). Intestinal flora disorders 
and increased intestinal permeability were induced through immune responses, which 
furtherly promoted the development and progression of NAFLD. In contrast, the 
application of antibiotics, prebiotics and probiotics that regulate intestinal microbiota 
can prevent the progress of NAFLD (Cited from: Wang Junyao, Liu Yulan. The roles 
of gut-liver axison the development and progression of nonalcoholic fatty liver 
disease. Journal of Clinical Hepatology, 2012,15 (4): 276 -278).In addition, some 
studies reported that the changes in intestinal microbiota were significantly related to 
insulin resistance. Alterations in intestinal microbiota not only induced elevated 
lipopolysaccharides (LPS) in the intestinal tract, but also damaged the tight junctions 

 
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of the intestinal epithelium, leading to increased flow of LPS into the blood 
circulation, causing a series of non-specific inflammatory responses. In consequence, 
the alterations in intestinal microbiota interfered with insulin signaling and induced 
insulin resistance. 
Some studies have shown that traditional Chinese medicine (TCM) has potentially 
effective therapies for chronic diseases. Additionally, TCM has unique therapeutic 
effects on some diseases that are particularly related to metabolic syndromes like 
hyperlipidemia, diabetes and fatty liver. For example, berberine was reported to 
improve insulin resistance, have glucose-and lipid-lowering effects, and was even 
used for the treatment of NAFLD. Researchers found in a NASH mice model that oral 
administration of berberine can change the intestinal microbiota, improve biochemical 
markers, attenuate liver histological changes, and reduce the intrahepatic 
inflammatory responses. The functions of berberine on whole-body energy 
metabolism and NAFLD were exerted by adjusting the ecological structure and 
microenvironment of the intestinal flora(Cited from: Cao Yi, Xu Leiming, PanQin, 
Wang Xiaoying, Shen Feng, Chen Guangyu, Fang Jiangao. Modulation of gut 
microbiota with berberine improves nonalcoholic steatohepatitis in mice[J]. Journal of 
Clinical Hepatology, 2013, 16 (2): 137-140).Compound Zhenzhu Tiaozhi capsules 
consist of eight Chinese herbal medications, such as Ligustrum lucidum, Atractylodes 
macrocephala, 
Radix 
Salviae 
Miltiorrhizae, 
pseudo-ginseng, 
Astragalus 
membranaceus. Previous studies found that Compound Zhenzhu Tiaozhi capsules can 
reduce serum triglyceride (TG), total cholesterol (TC), fasting blood glucose, insulin, 
and the insulin resistance index. In addition, it can increase serum high-density 
lipoprotein cholesterol (HDL-C) levels, reduce weight, and improve obesity in a rat 
model, which means that it can be used to treat metabolic syndrome. It was also 
reported that Compound Zhenzhu Tiaozhi capsules upregulated PI-3Kp85 mRNA 
expression in rat adipose tissue, which might be one of the mechanisms involved in 
insulin resistance (cited from: Hu Xuguang, Guo Jiao, Bai Weijian, He Wei, Wang 
Shou, Liu Shasha. Effect of Fufang Zhengzhu Tiaozhi Capsules on Insulin Resistance 
and Insulin Signal PI-3K in Metabolic Syndrome Rats [J].Traditional Chinese Drug 

 
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Research and Clinical Pharmacology, 2012, 23 (2): 140-143).Additionally, 
Compound Zhenzhu Tiaozhi capsules can reduce serum TG, TC, and low-density 
lipoprotein cholesterol (LDL-C) and increase serum HDL-C levels in a 
hyperlipidemic rat model (Cited from: Tang Chunping, Gong Mengjuan, Guo Jiao, 
Yang Chaoyan, Chen Fangchao, Yao Hongxia, Chen Yuefeng. The effects of Fufang 
Zhenzhu Tiaozhi capsules on blood lipid level and hemorheology of rats with 
hyperlipemia [J]. Journal of Guangdong Pharmacy College, 2010, 26 (6): 612-616). 
Based on the above studies, the investigators hypothesize that Compound 
Zhenzhu Tiaozhi capsules have the potential to effectively improve NAFLD and 
related biomedical markers. The possible mechanisms include the improvement of 
SIBO and the function of the intestinal mucosal barrier. The investigators thus 
designed the following clinical study to test this hypothesis. 
 
II. Objectives 
To evaluate the efficacy of Compound Zhenzhu Tiaozhi capsules in the 
treatment of NAFLD and analyze the relationship between improvement in NAFLD 
parameters and changes in intestinal functions. 
 
III. Content of the study 
1. To evaluate the relationship between the function of intestinal mucosal barrier, 
SIBO, and NAFLD; 
2. To assess the effects of Compound Zhenzhu Tiaozhi capsules on intrahepatic 
fat content, hepatic noninvasive score, liver biochemical parameters, blood lipid, and 
insulin resistance. To evaluate the therapeutic effects of Compound Zhenzhu Tiaozhi 
capsules on the treatment of NAFLD, and determine whether the risk stratification of 
atherosclerotic cardiovascular disease (ASCVD) and quality of life (using the 36-Item 
Short Form Health Survey [SF-36]) can be improved in NAFLD patients with 
Compound Zhenzhu Tiaozhi capsules. 
3. To compare the therapeutic effects of Compound Zhenzhu Tiaozhi capsules 
with conventional medications in the treatment of NAFLD, and to compare their 
efficacies in improving the risk stratification of ASCVD and quality of life, as 

 
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assessed by the SF-36. 
4. Finally, to analyze the relationship of Compound Zhenzhu Tiaozhi capsules in 
improving NAFLD and parameters related to the function of the intestinal mucosal 
barrier and SIBO. 
 
IV. Methods 
This is an interventional and randomized parallel control study. The study will be 
conducted from December 31 2017 to December 31 2019. The recruitment and 
observation period will start in December, 2017 and continue until the required 
sample size is reached. Research venue: First Affiliated Hospital, Guangdong 
Pharmaceutical University, Guangdong province, China. Institute level: a 
third-grade-A-class hospital. 
1. Ethical approval: The study will be conducted after obtaining ethical approval from 
Ethics Committee of the First Affiliated Hospital, Guangdong Pharmaceutical 
University. The investigators will inform all prospective subjects of the study 
background, purpose, design, and medication contents. If a subject is willing to 
participate in the study, he or she will sign the consent form (Supplementary 1). 
 
2. Enrollment: 
The subjects will be enrolled following the design of a prospective study. The 
inclusion criteria will be strictly in accordance with the diagnostic criteria of simple 
steatosis and NASH, as stipulated in the Chinese Guidelines for the Diagnosis and 
Treatment of Nonalcoholic Fatty Liver Disease (Revised version 2010). The subjects 
should have a diagnosis of simple steatosis or NASH (age distribution: 18 to 75 years 
old), with serum alanine transaminase (ALT)≤2 times the upper limit of normal, with 
no other chronic liver diseases or serious complications in other organs, with on 
malignant tumors. 
2.1. Diagnostic Criteria of NAFLD (2010): NAFLD can be diagnosed if the following 
items 1–4,plus item 5 or item 6 are present:  
i. No alcoholic history (alcohol consumption <140 g/week for male;or <70 

 
7 
g/week for female);  
ii. No other diseases that can lead to fatty liver, such as viral hepatitis, 
drug-induced liver disease, total parenteral nutrition, hepatolenticular 
degeneration diseases, or autoimmune liver diseases; 
iii. Overweight 
and/or 
visceral 
obesity, 
hyperglycemia, 
dyslipidemia, 
hypertension and other metabolic syndrome;  
Serum transaminase and GGT levels are normal or elevated to a mild to 
moderate degree;  
iv. Liver imaging findings show diffuse hepatic steatosis; 
v. A liver biopsy meets the histologic diagnostic criteria of fatty liver diseases; 
2.2. Diagnostic criteria of simple steatosis: Simple steatosis can be diagnosed if the 
following item 1-2, plus item 3 or item 4 are present. 
i. Presence of item1-3 of the clinical diagnosis criteria of NAFLD;  
ii. Liver biochemical tests are normal (blood biochemical analysis);  
iii. Liver imaging findings meet the diagnostic criteria for hepatic steatosis 
(Fibro touch test); 
iv. A liver biopsy meets the histologic diagnostic criteria for simple hepatic 
steatosis. 
2.3. The diagnostic criteria of NASH: NASH can be diagnosed with the following 
items 1 to 3 or item 4. 
i. Presence of items 1–3 of the clinical diagnosis criteria of NAFLD; 
ii. Serum ALT levels are persistently elevated for more than 4 weeks; 
iii. Liver imaging findings show diffuse hepatic steatosis;  
iv. A liver biopsy meets the histologic diagnostic criteria for NASH. 
2.4. The following individuals will be considered as lost to follow-up: first, patients 
who suddenly present serious physical or mental diseases (not related to the study) 
during the study period, such as malignant tumors, severe mental illness, traffic 
accidents, etc.; second, patients who have serious side effects in which the symptoms 
can be relieved by discontinuing the study medication; third, patients who actively ask 
to stop using the study medication or refuse to come to follow up visits. Fourth, 

 
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patients who need to take other medications due to the development of other new 
diseases, where the new medications may interfere with the study results, such as 
hypoglycemic agents, including insulin. 
 
3. Assignment and intervention: 
 3.1. Calculation of the sample size and the method of assignment: 
Sample size calculation: A total of 196 patients will be enrolled in this study. The sample 
size is estimated as follows: 
Previous studies have reported that the efficiency rate of metformin and simvastatin in 
the treatment of NAFLD is 36-52%. The investigators hypothesize that the efficiency rate of 
Compound Zhenshu Tiaozhi capsules will increase by 25% compared to standard drugs. 
Statistical calculation: P1 = 69%, P2 = P3 = 44%, P4 = 10%, δ = P1-P2 = 25%, with α = 
0.05, power = 0.80, and a bilateral test, of which 
 , thus n will be equal to 
44. Assuming that the rate of loss to follow up is 10%, each group will require 49 cases. 
Therefore, a total of 196 cases will be required for the four groups. 
3.2 The method of assignment: Zhang Min, a staff member working in the 
Department of Statistics in Guangdong Pharmaceutical University, is responsible for 
generating random sequences using the SAS software. A total of 196 numbers from 1 
to 196 will be randomly generated (Supplement 2). Newly diagnosed patients who 
meet the inclusion criteria will have a case number according to their order of 
enrollment (for example: case number of the first patient: 001, second patient: 002). 
They will then obtain the corresponding therapeutic regimens based on their case 
number with 1:1:1:1 ratio randomized tables. The first group is the TLC group; the 
second group is the TLC + metformin group (0.5 g, PO tid); the third group is the 
TLC + Compound Zhenzhu Tiaozhi capsules group (2.52 (four tablets), PO tid); the 
fourth group is the TLC + simvastatin group (20 mg, PO qn). 
4. Endpoints: 
4.1 Primary endpoints 

 
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Fat attenuation index: The fat attenuation index will be used to assess the therapeutic efficacy. 
A normal fat attenuation index is defined as < 240 db/m, mild is 240-264 db/m, moderate is 
265-294 db/m, and severity is > 295 db/m. The investigators will detect the fat attenuation 
index at 0, 1, 3, 6 months following treatment. Effective: fat attenuation index is reduced by a 
level or more (example: moderate to mild). Invalid: fails to meet the effective standard. 
 
4.2 Secondary endpoints: 
(1) Serum triglyceride 
(2) Serum cholesterol 
(3) Serum lipoproteins: chylomicron (CM), very-low-density lipoprotein (VLDL), low-density 
lipoprotein (LDL), and high-density lipoprotein (HDL). 
(4) Hydrogen/methane breath testing: breath testing for SIBO. 
(5) Intestinal mucosal barrier: detect levels of diamine oxidase (DAO), D-lactic acid (D-lac), and 
lipopolysaccharide (LPS), and combine with the clinical manifestations to evaluate the function of 
the intestinal mucosal barrier.  
(6) Quality of life (SF-36 Scores): The quality of life of the patients before and after treatment will 
be assessed using SF-36 scores and compared to assess the improvement. 
 
4.3. Analysis of the relationships among the function of intestinal mucosal barrier, 
SIBO, and NAFLD. 
1) To analyze the relationship of DAO, D-lac, LPS, intestinal mucosal barrier 
dysfunction grade I, II, III, IV within trahepatic fat content in NAFLD 
patients(possible results: unrelated, positively related, or negatively related). 
2) To evaluate the relationship between SIBO and intrahepatic fat content in NAFLD 
patients (possible results: unrelated, positively related, or negatively related). 
3) To assess the relationship of DAO, D-lac, LPS, intestinal barrier function 
dysfunction grade I, II, III, IV with insulin resistance in NAFLD patients (possible 
results: unrelated, positively related, or negatively related). 
4) To detect the relationship between SIBO and insulin resistance in NAFLD patients 
(possible results: unrelated, positively related, or negatively related). 

 
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5) To explore the relationship of DAO, D-lac, LPS, intestinal barrier dysfunction 
grade I, II, III, IV and peripheral blood lipidsprofile and liver biochemical parameters 
(ALT, AST, GGT) (possible results: unrelated, positively related, or negatively 
related). 
6) To investigate the relationship of SIBO and lipids profile and liver biochemical 
parameters (ALT, AST, GGT) in NAFLD patients (possible results: unrelated, 
positively related, or negatively related). 
7) To estimate the relationship of DAO, D-lac, LPS, intestinal barrier dysfunction 
grade I, II, III, IV and NAFLD-FS in NAFLD patients (possible results: unrelated, 
positively related, or negatively related). 
8) To determine the relationship between SIBO and NAFLD-FS in NAFLD patients 
(possible results: unrelated, positively related, or negatively related). 
4.4. To evaluate the therapeutic effects of Compound Zhenzhu Tiaozhi capsules on 
NAFLD 
1)To assess whether the content of intrahepatic fat will decrease after 1 month, 3 
months, and 6 months of Compound Zhenzhu Tiaozhi capsule therapy. 
i. Compare the therapeutic effects of Compound Zhenzhu Tiaozhi capsules at 1 
month, 3 months, and 6 months with the baseline (pre-vs.post-therapeutic 
effects of Compound Zhenzhu Tiaozhi capsules). 
ii. Compare the therapeutic effects in the Compound Zhenzhu Tiaozhi 
capsulegroup with that of the other three groups atparallel time points 
(Compound Zhenzhu Tiaozhi capsules vs.Conventional drugs). 
 
2) To assess whether NAFLD-FS will be alleviated after 1 month, 3 months, and 6 
months of Compound Zhenzhu Tiaozhi capsuletherapy. 
i. Compare the therapeutic effects of Compound Zhenzhu Tiaozhi capsules at 1 
month, 3 months, and 6 months with the baseline (pre- vs. post- 
therapeutic effects of Compound Zhenzhu Tiaozhi capsules). 
ii. Compare the therapeutic effects in the Compound Zhenzhu Tiaozhi capsule 
group with those of the other three groups at parallel time points 

 
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(Compound Zhenzhu Tiaozhi capsules vs.Conventional drugs). 
 
3) To assess whether insulin resistance will be improved after 1 month, 3 months, 6 
months of Compound Zhenzhu Tiaozhi capsule therapy. 
i. Compare the therapeutic effects of Compound Zhenzhu Tiaozhi capsules at 1 
month, 3 months, and 6 months with the baseline (pre- vs. post- 
therapeutic effects of Compound Zhenzhu Tiaozhi capsules). 
ii. Compare the therapeutic effects in the Compound Zhenzhu Tiaozhi capsule 
group with those of the other three groups at parallel time points 
(Compound Zhenzhu Tiaozhi capsules vs. Conventional drugs). 
 
4) To assess whether the lipids profile in peripheral blood and liver biochemical 
parameters(ALT, AST, GGT) will be reduced after 1month, 3months, and6months of 
Compound Zhenzhu Tiaozhi capsule therapy. 
i. Compare the therapeutic effects of Compound Zhenzhu Tiaozhi capsules at 1 
month, 3 months, and 6 months with the baseline (pre- vs. post- 
therapeutic effects of Compound Zhenzhu Tiaozhi capsules). 
ii. Compare the therapeutic effects in the Compound Zhenzhu Tiaozhi capsule 
group with those of the other three groups at parallel time points 
(Compound Zhenzhu Tiaozhi capsules vs. Conventional drugs). 
 
4.5 In the Compound Zhenzhu Tiaozhi capsules group, firstly calculate the changes in 
the related parameters of NALFD and the changes in the function of intestine 
microbiota, then evaluate whether the two changes are related to each other. 
1)Analyze the relationship between the changes in DAO, D-lac, LPS, intestinal barrier 
dysfunction grade I, II, III, IV and the reduction in intrahepatic fat at 1 month, 3 
months, and 6 months of Compound Zhenzhu Tiaozhi capsuletherapy (possible results: 
unrelated,positively related, or negatively related). 
2)Analyze the relationship between the change in SIBO and the reduction in 
intrahepatic fat at 1 month, 3 months, and 6 months of Compound Zhenzhu Tiaozhi 

 
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capsule therapy (possible results: unrelated,positively related, or negatively related). 
3)Analyze the relationship between the changes in DAO, D-lac, LPS, intestinal 
mucosal barrier dysfunction I, II, III, IV and the improvement of insulin resistance 
at1month, 3months, and 6months of Compound Zhenzhu Tiaozhi capsule therapy 
(possible results: unrelated, positively related, or negatively related). 
4)Analyze the relationship between the change in SIBO and the improvement of 
insulin resistance at 1 month, 3 months, and 6 months of Compound Zhenzhu Tiaozhi 
capsule therapy (possible results: unrelated, positively related, or negatively related). 
5) Analyze the relationship between changes in DAO, D-lac, LPS, intestinal mucosal 
barrier dysfunction I, II, III, IV and the changes in the lipids profile in peripheral 
blood and liver biochemical parameters(ALT, AST, GGT) at 1 month, 3 months, and 6 
months of Compound Zhenzhu Tiaozhi capsule therapy (possible results: unrelated, 
positively related, or negatively related). 
6) Analyze the relationship between the change in SIBO and the changes in the lipids 
profile in peripheral blood and liver biochemical parameters(ALT, AST, GGT) at 
1month, 3months, and 6months of Compound Zhenzhu Tiaozhi capsule therapy 
(possible results: unrelated, positively related, or negatively related). 
7) Analyze the relationship between changes in DAO, D-lac, LPS, intestinal mucosal 
barrier dysfunction I, II, III, IV and the change in NAFLD-FS at 1month, 3months, 
and 6months of Compound Zhenzhu Tiaozhi capsuletherapy (possible results: 
unrelated, positively related, or negatively related). 
8)Analyze the relationship between the change in SIBO and the change in NAFLD-FS 
at 1 month, 3 months, and 6 months of Compound Zhenzhu Tiaozhi capsule therapy 
(possible results: unrelated, positively related, or negatively related). 
 
4.6. To estimate the ASCVD risk for all NAFLD patients, and to evaluate whether the 
therapy of Compound Zhenzhu Tiaozhi capsules improves the risk stratification of 
ASCVD for NALFD patients, as well as to compare the therapeutic effects of the 
study capsules and other conventional drugs. The investigators will estimate the 
ASCVD risk for NAFLD patients after 1month, 3 months, and 6 months of 

 
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Compound Zhenzhu Tiaozhi capsule therapy, according to Guidelines for Prevention 
and Treatment of Dyslipidemia in Adults. 
ASCVD populations with different risk stratifications have different 
target values of lowering LDL-C / non-HDL-C, as shown in the table 
below. 
 
Risk Stratification 
LDL-C 
Non-HDL-C 
High-risk 
<2.6mmol/L(100mg/dl) 
<3.4mmol/L(130mg/dl) 
Very high-risk 
<1.8mmol/L(70mg/dl) 
<2.6mmol/L(100mg/dl) 
Note: ASCVD: atherosclerotic cardiovascular disease; LDL-C: low density 
lipoprotein cholesterol; 
Non-HDL-C: non-high density lipoprotein cholesterol 
 
If the baseline of LDL-C is high, as it is difficult to reduce LDL-C level to the basic target value after 3 
months of standard treatment with existing lipid-lowering drugs, we propose to lower LDL-C level to at 
least 50% of baseline as the surrogate target value(IIa recommendation, Level B evidence). There are 
also some patients at very high-risk who have a baseline LDL-C within the basic target value; for these 
patients the investigators propose to reduce LDL-C levels to about 30% of baseline (I recommendation, 
A-level evidence). 
 
Those who meet any of the following criteria can be directly classified as high-risk or very 
high-risk populations, as indicated: 
1. Very high-risk: ASCVD patients (acute coronary syndrome, stable coronary heart 
disease, revascularization, ischemic cardiomyopathy, ischemic stroke, transient ischemic 
attack, peripheral atherosclerosis, etc.); 
2. High-risk: (1) LDL-C ≥4.9 mmol/L or TC ≥7.2 mmol/L, (2) Diabetics:1.8 mmol/L≤ 
LDL-C <4.9 mmol/L (or) 3.1 mmol/L≤ TC <7.2 mmol/L, and age ≥ 40 years old. 
3. For those who do meet the above criteria, the investigators will evaluate their 10-years 
ASCVD risk as follows 
 
 
Number of risk Stratification of Serum Cholesterol (mmol/L) 

 
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factors 
3.1oTC<4.1(or) 
1.8≤LDL-C<2.6 
4.1.TC<5.2(or) 
2.62LDL-C<3.4 
5.2≤TC<7.2(or)3
.42LDL-C<4.9 
Without 
hypertension0–1 
Low-risk (<5%) 
Low-risk (<5%) 
Low-risk (<5%) 
2 
Low-risk (<5%) 
Low-risk (<5%) 
Moderate 
risk 
(5%–9%) 
3 
Low-risk (<5%) 
Moderate risk (5%–9%) 
Moderate 
risk 
(5%–9%) 
  
  
  
  
hypertension 
0 
Low-risk (<5%) 
Low-risk (<5%) 
Low-risk (<5%) 
1 
Low-risk (<5%) 
Moderate risk (5%–9%) 
Moderate 
risk 
(5%–9%) 
2 
Moderate 
risk 
(5%–9%) 
High-risk (≥10%) 
High-risk 
(≥10%) 
3 
High-risk (≥10%) 
High-risk (≥10%) 
High-risk 
(≥10%) 
 
4. If the 10-year ASCVD risk is moderate and the age is less than 55 years old, then the 
investigators propose to do the remaining life assessment. Those who meet any two of the 
following items can be classified as high-risk: 1. ≥160 mmHg or diastolic blood 
pressure≥100mmHg; 2. non-HDL-C ≥5.2 mmol/L (200 mg/dl); 3. HDL-C <1.0 mmol/L 
(40mg/dl); 4. BMI≥28kg/m2; 5. history of smoking. 
3.5. To assess the quality of life using the SF-36 questionnaire for all NAFLD patients, 
and to evaluate whether the therapy of Compound Zhenzhu Tiaozhi capsules improves the 
quality of life for NALFD patients, as well as to compare its therapeutic effects with other 
conventional drugs. 
Patients will be required to fill in the SF-36 questionnaire at the time of enrollment, and 
after 1 month, 3 months, and 6 months of treatment. SF-36 is one of the most widely 
used and standardized tools for quality of life assessment. SF-36, as a simple health 
questionnaire, recapitulates the quality of life of the respondents based on eight 
categories: physiological function, physical pain, general health status, energy, social 
function, emotional function, and mental health. The first four categories describe 
physical health, the latter four describe mental health. 

 
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The early symptoms of NAFLD are not obvious and almost have no impact on daily life. 
As it is difficult to effectively assess whether patients have benefited from treatment by 
evaluating symptoms, the investigators will use the SF-36 questionnaire. The 
questionnaire will be administered at the time of patient enrollment, and at 1month, 
3months, and 6 months after treatment. To determine whether drug intervention during 
the early stage of NAFLD can improve symptoms, the scores of theSF-36 before and after 
treatment will be compared in the following groups: TLC + metformin, TLC +Compound 
Zhenzhu Tiaozhi capsules, TLC + simvastatin, and TLC. The four groups will also be 
compared to one another. 
4. Prescriptions: 
4.1: Medications: the dose of Compound Zhenzhu Tiaozhi capsules is 2.52 (4 tablets) tid; 
metformin is 0.5, tid, and simvastatin is 20mgqn. In general, subjects will be in the 
hospital for approximately 1 week and continue to receive treatment after discharge from 
the hospital. The total period of treatment will be 6 months. 
 
4.2:Biochemical assessments: Patients will undergo all the following tests upon 
admission: 
Routine blood test, tumor biomarker tests (alpha-fetoprotein [AFP], carcinoembryonic 
antigen [CEA], carbohydrate antigen 125 [CA125], and carbohydrate antigen 19-9 
[CA19]), 
uric 
acid, 
liver 
function 
(representing 
hepatocyte 
injury: 
alanine 
aminotransferase, 
aspartate 
aminotransferase, 
leucyl 
aminopeptidase, 
glutamyl 
transpeptidase; representing liver secretory function: total bilirubin (TBS), direct bilirubin 
(DBIL),indirect bilirubin (IBIL); representing liver reserve function: total protein (TP), 
albumin (ALB), globulin (GLB), and the albumin to globulin ratio (A/G), serum lipid 
profile (triglyceride, very low density lipoprotein, total cholesterol, low density 
lipoprotein, high density lipoprotein), FINS (fasting insulin), fasting blood glucose, serum 
biomarkers of HBV ([i]Hepatitis B surface antigen [HBsAg]; [ii]Hepatitis B surface 
antibody [anti-HBs]; [iii]hepatitis B e antigen [HBeAg]; [iv]Hepatitis B e antigen 
antibody[anti-HBe]; [v]Hepatitis B core antibody [anti-HBc]), TP-Ab, syphilis serological 
test (TRUST), human immunodeficiency virus(HIV), hepatitis C virus antibody, 

 
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glycosylated hemoglobin, intestinal mucosal barrier function, coagulation test 
(prothrombin time [PT], activated partial thromboplastin time [APTT], thrombin time 
[TT], and fibrinogen [FIB]), Fibrotouch, hydrogen and methane breath test. 
 
4.3 Medical advice on discharge: Physicians need to inform patients regarding the 
severity of complications in the late stage of NALFD and the importance of early 
treatment. All the patients should continue treatment after discharge and follow-up in the 
clinic.They will be required to come back to hospital one month later for re-assessment of 
the above mentioned items. The appointment telephone number is 15819544433. The 
contact doctor is Dr. Lin. 
 
4.4 All patients will be treated with TLC. 
4.4.1 Dietary advice: On the basis of daily nutrient and energy requirements, if the total 
amount of saturated fatty and trans fatty acids exceeds the recommended upper limit, 
unsaturated fats can be used to replace saturated fats. Recommended daily intake of 
cholesterol is less than 300 mg, especially for ASCVD high-risk patients.The total fat 
caloric intake should be limited to 20%–30%of the total caloric intake. 
4.4.2 Controlling weight: Obesity is an important risk factor for dyslipidemia. Energy 
intake in overweight or obese people with dyslipidemia should be lower than energy 
consumption to prevent weight increase.The goal is to reduce weight gradually until a 
normal state is reached. Reducing the total amount of daily caloric intake(daily reduction 
of 300–500 kcal), improving the diet style, and increasing physical activities can reduce 
weight by more than 10%for overweight and obese people. It is recommended that 
patients maintain a healthy weight. 
4.4.3 Physical activity advice: It is recommended that patients do moderate-intensity 
exercises, at least 30 minutes per day for 5 days per week. ASCVD patients should do 
loading activities first to assess whether they are tolerable, and then do physical activities. 
4.4.4 Smoking cessation: Smoking cessation and effective reduction of exposure to 
second-hand smoke are beneficial to prevent ASCVD and increase serum HDL-C levels. 

 
17
All smokers can be assisted in the clinics, inquire about smoking cessation through 
hotlines, or take medication to help with smoking cessation. 
4.4.5 Restrictions on drinking: Moderate drinking (male: 20-30 g of ethanol per day, 
female:10 - 20 g of ethanol per day) can increases serum HDL-C level. However, even a 
small amount of drinking can increase TG in patients with high TG. There is no 
convincing evidence that alcohol can increase cardiovascular disease. It is recommended 
that patients limit drinking. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 TLC + simvastatin group: 
20 mg, PO qn 
     All patients should continue treatment after discharge and follow-up in the clinic. 
Patients with simple steatosis or NASH, with serum alanine transaminase (ALT) 
≤ 2 times the upper limit of normal, without other chronic liver diseases or 
malignant tumors; patients who use insulin to lower blood glucose and use 
simvastatin to lower blood lipids will be excluded. 
TLC + Compound Zhenzhu 
Tiaozhi capsule group: 2.52 
(four tablets), PO tid 
Monitoring lipids and insulin 
resistance-related parameters 
Biochemical tests  
Physical 
examinations  
Imaging 
The function of 
the 
intestinal 
mucosal barrier 
hydrogen 
and 
methane 
breath 
tests 
Gut-function assessment 
TLC + metformin 
group: 0.5 g, PO tid 
All patients should have follow-up visits in the clinic 
after 1 month, 3 months, and 6 months of treatment.
Noninvasive 
liver 
scoring 
system 
to 
predict liver cirrhosis 
TLC 
group: 
only TLC 
Perform risk stratification of ASCVD and SF-36. 
 
 
Biochemical tests: routine blood test, liver function tests including transpeptidases, uric 
acid, FBS,FINS, lipid profile. 

 
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Physical examinations: waist circumference, hip circumference, blood pressure, heart 
rate, BMI 
Imaging: abdominal color doppler ultrasound, Fibrotouch 
 
5. Expected results: 
1) The liver biochemical parameters and other clinical parameters in NAFLD 
patients will positively correlate with SIBO and intestinal mucosal barrier function. 
2) Compound Zhenzhu Tiaozhi capsules will effectively improve liver function 
(intrahepatic fat content, liver biochemical markers, NAFLD-FS) and related 
serological parameters(improvement of dyslipidemia and insulin resistance) in 
NAFLD patients;Compound Zhenzhu Tiaozhi capsules will effectively improve the 
SF-36 score and reduce the risk of ASCVD in patients with NAFLD. 
3) Compound Zhenzhu Tiaozhi capsules will be superior to conventional drugs 
(simvastatin and metformin) in terms of improving liver biochemical markers and 
related serological parameters, SF-36 score, and risk stratification of ASCVD. 
4) All assessed items of NAFLD patients (liver: intrahepatic fat content, liver 
biochemical parameters, NAFLD-FS; and its related serological indicators: 
dyslipidemia, insulin resistance; SF-36 score; and risk stratification of ASCVD) will 
be positively associated with SIBO and improved function of the intestinal mucosal 
barrier.