The Effect of N-nitrosodimethylamine and Metabolic Activation in

the Liver on the Formation of Nasopharyngeal Carcinoma

Waiki Cen

The British School of Guangzhou, Guangzhou, 510515, China

Keywords:

Nasopharyngeal Carcinoma (NPC), N- nitrosodimethylamine (NDMA), Timecourse, Metabolic Activation.

Abstract:

Nasopharyngeal carcinoma (NPC) is a common tumor of the head and neck with originates in the nasopharynx

cancer, especially in Asia. Moreover, N-nitrosodimethylamine (NDMA) is a highly toxic, carcinogenic

nitrosamine, C

O. It is generally accepted that NDMA is the main cause of NPC. This research aims to

investigate the level of NDMA necessary to induce the formation of NPC. To achieve the purpose of this

experiment (how much NDMA will cause the formation of NPC) I first injected different amounts of NDMA

and pyrazole into the 3 Male Ydd strain mice (inject nothing into the control mice), then carry out a

timecourse, lastly use MRI to test the formation of NPC in the Male Ydd strain mice. The result obtained

from the experiment is when the level of NDMA is higher than 6.25nmole/200kd of water/mouse and the

level of metabolic activation is lowered NPC would be formed (Not found out yet). This study sheds light on

the future study about the formation of NPC, also suggests that people should eat less smoked food.

1 INTRODUCTION

Nasopharyngeal carcinoma (NPC) is a common

tumor of the head and neck with originates in the

nasopharynx (Osong Public Health and Research

Perspectives. 2016), especially in southern China.

(Cao,

Simons and Qian. 2011) There is less than one

case for every 100,000 people in most parts of the

world (including the United States) per year.

(American Cancer Society. 2018) Whereas in part of

southern China 141.7/100,000 people would be

diagnosed with NPC per year. (Cao,

Simons and Qian.

2011) A previous study (Smoked cooked meat as a

risk factor for Nasopharyngeal Carcinoma: A case-

control study among Saudi populations (Saudi

Journal of Oto-Rhino-Laryngology Head and Neck

Surgery. 2018) has reported that smoked food is one

of the biggest causes of NPC. And in the newest

study, it’s suggested that NDMA in smoked food is

the main cause of NPC. (Khalid Hakami, N

Prepageran, Talal Al Thubaity, Eidah Al Juaid, Nawaf

Al Solami, 2018) In the work, I investigated whether

the level of NDMA in the body tissues and metabolic

activation in the liver (IARC Sci Publ. 1980) causes

the rate of getting Nasopharyngeal carcinoma (NPC)

to increase.

1.1 Purpose

Nasopharyngeal carcinoma (NPC) is a common

tumor of the head and neck with originates in the

nasopharynx (Osong Public Health and Research

Perspectives. 2016), especially in southern China.

(Su-Mei Cao,

Malcolm J. Simons and Chao-Nan

Qian. 2011) There is an increasing amount of people

diagnosed with NPC in Guangdong in the last 29

years (1970-1999). (Karen Michell Othaya Kumar

and Rabiatul Basria S.M.N. Mydin. 2019) A previous

study has reported that NPC is mainly caused by N-

nitrosodimethylamine (NDMA). This study will

investigate whether the level of NDMA in the body

tissues and metabolic activation in the liver (IARC

Sci Publ. 1980) causes the rate of getting

Nasopharyngeal carcinoma (NPC) to increase.

1.2 Methods

The experiments will use Male Ydd strain mice,

which will be injected with increasing amounts of

NDMA as the experiment measures liver function

through assay. An increasing amount of pyrazole will

also be injected, for speeding up the reduction of

metabolic activation in the liver. pyrazole (used to

replace part of the NDMA  speeding up the

reduction of metabolic. By doing so, it is less time

Cen, W.

The Effect of N-nitrosodimethylamine and Metabolic Activation in the Liver on the Formation of Nasopharyngeal Carcinoma.

DOI: 10.5220/0011295200003443

In Proceedings of the 4th International Conference on Biomedical Engineering and Bioinformatics (ICBEB 2022), pages 785-790

ISBN: 978-989-758-595-1

785

consuming and can save money) (Metabolic

activation means the chemical conversion of a

relatively benign substance into a more hazardous one

by normal biochemical processes in cells and tissues.)

A timecourse experiment is then performed, for

measuring the amount of NDMA in the liver and body

tissues. Also, the time takes for NPC to form under

different dosages of pyrazole. The amount of NPC

formed is measured using an MRI.

1.3 Possible Results

There are three most possible results: (1) Mice

pretreated with a larger amount of pyrazole would

have a higher recovery of NDMA administration; (2)

pH value would decrease administration of

pyrazole; (3) NPC is to form in

pretreated mice quicker than in control mice.

1.4 Methods

Materials

This experiment will use

 male ddY strain mice (20-25g)

 pyrazole (100-300 mg/5 ml of saline/kg):

Speeding up the reduction of metabolic activation in

the liver of mice.

 Saline(5ml/kg): Clean the mice, clear

possible cancerogen that may affect the result.

 NDMA (6.25-100 nmole/200kd of

water/mouse): Main source of causing NPC.

 14-C Aminopyrine (0.063-1.0

kdmole/100kd of water/mouse)：ABT rate measures

the severity level of liver tissue.

 MRI: Detect NPC in the mice.

Animals:

Male ddY strain mice (20-25g).

ABT rate and metabolic activation of liver tissues:

[Control mice]: First, clear out all the possible

cancergen inside the control mice by injecting it with

Saline(51/kg), 60 minutes before the administration.

Then use 14-C aminopyrine (0.063-

1.0xmole/100~tlof water/mouse) for measuring the

ABT rate (severity level of liver tissue) and the

metabolic activation of the liver tissues in the control

mice.

[Mice-1]: First, clear out all the possible

cancergen inside mice-1 by injecting it with

Saline(51/kg). Second, inject mice-1 with NDMA

(6.25-100nmole/200kd of water/mouse), which

provides mice-1 with small amount of cancergen of

NPC and source for reduction of metabolic activation

in liver. Third, inject mice-1 with

pyrazole(100mg/kg), for speeding up the reduction of

metabolic activation in mice-1’s liver. (60 minutes

before the administration). Last, use 14-C

aminopyrine (0.063-1.0xmole/100~tlof water/mouse)

for measuring the ABT rate (severity level of liver

tissue) and the metabolic activation of the liver tissues

in mice-1.

[Mice-2]: First, clear out all the possible

cancergen inside mice-2 by injecting it with

Saline(51/kg). Second, inject mice-2 with NDMA

(6.25-100nmole/200kd of water/mouse), which

provides mice-2 with medium amount of cancergen

of NPC and source for reduction of metabolic

activation in liver. Third, inject mice-2 with

pyrazole(200mg/kg), for speeding up the reduction of

metabolic activation in mice-2’s liver. (60 minutes

before the administration). Last, use 14-C

aminopyrine (0.063-1.0xmole/100~tlof water/mouse)

for measuring the ABT rate (severity level of liver

tissue) and the metabolic activation of the liver tissues

in mice-2.

[Mice-3]: First, clear out all the possible

cancergen inside mice-3 by injecting it with

Saline(51/kg). Second, inject mice-3 with NDMA

(6.25-100nmole/200kd of water/mouse), which

provides mice-3 with large amount of cancergen of

NPC and source for reduction of metabolic activation

in liver. Third, inject mice-3 with

pyrazole(300mg/kg), for speeding up the reduction of

metabolic activation in mice-3’s liver. (60 minutes

before the administration). Last, use 14-C

aminopyrine (0.063-1.0xmole/100~tlof water/mouse)

for measuring the ABT rate (severity level of liver

tissue) and the metabolic activation of the liver tissues

in mice-3.

Amount of NDMA remaining in the body tissues:

Measure the amount of NDMA in the body by

testing the blood/urine.

Time takes for an NPC to form based on the

amount of pyrazole used:

[Control mice]: First, Inject the control mice with

no pyrazole, measure the time it takes to form an NPC

in control mice by using timecourse. Then, detect the

formation of NPC with an MRI. Check the amount of

NPC formed in the control mice 7days per once.

[Mice-1]: First, inject mice-1 with NDMA (6.25-

100nmole/200kd of water/mouse) second, inject

mice-1 with pyrazole(100mg/kg). （Pyrazole is used

to speed up the reduction of metabolic activation of

all the body tissue without using a large amount of

NDMA which is a way of saving time and money.）

Measure the time it takes to form NPC in mice-1 by

using timecourse. Last, detect the formation of NPC

with an MRI. Check the amount of NPC formed in

mice-1 7days per once.

ICBEB 2022 - The International Conference on Biomedical Engineering and Bioinformatics

786

[Mice-2]: First, inject mice-2 with NDMA (6.25-

100nmole/200kd of water/mouse) second, inject

mice-2 with pyrazole (200mg/kg). Measure the time

it takes to form a NPC in mice-2 by using timecourse.

Last detect the formation of NPC with an MRI. Check

the amount of NPC formed in mice-2 7days per once.

[Mice-3]: First, inject mice-3 with NDMA (6.25-

100nmole/200kd of water/mouse) second, inject

mice-3 with pyrazole (300mg/kg). Measure the time

it takes to form a NPC in mice-3 by using timecourse.

Last detect the formation of NPC with an MRI. Check

the amount of NPC formed in mice-3 7days per once.

1.5 Result

Table 1 Percentage of NDMA recovery 60 minutes after the NDMA administration based on amount of pyrazole injected to

the mice. With amounts of pyrazole injected to the mice listed.

Male ddY strain

mice (20-25g)

Amount of pyrazole injected

to the mice (mg/kg)

Percentage of NDMA recovery 60 minutes after

the NDMA administration (%)

Control mice 0mg/kg 0

Mice-1 100mg/kg 50

Mice-2 200mg/kg 71

Mice-3 300mg/kg 84

Figure 1 Percentage of NDMA recovery 60 minutes after the NDMA administration based on amount of pyrazole injected to

the mice.

Table 2 Comparison of positive or negative result of NDMA presence between control mice, Mice-1, Mice-2, Mice-3.

Control mice Mice-1 Mice-2 Mice-3

Formation of NPC - + + +

NDMA present in urine - + + +

NDMA present in blood - + + +

Control mice Mice-1 Mice-2 Mice-3

The Effect of N-nitrosodimethylamine and Metabolic Activation in the Liver on the Formation of Nasopharyngeal Carcinoma

787

Table 3 time takes for an NPC to form based on the amount of pyrazole used

7 da

s 14da

s 21da

s 28da

s 35da

s 43da

s 50da

s 57da

Control mice- - ------

Mice-1 - - ----++

Mice-2 - - - - ++++

Mice-3 - - ++++++

Possible result 1: No recovery of NDMA in control

mice.

Possible result 2: Mice pretreated with 100mg/kg

of pyrazole, NDMA recovery is below 50% 60

minutes after the NDMA administration.

Possible result 3: Mice pretreated with 200mg/kg

of pyrazole, NDMA recovery is about 71.2% 60

minutes after the NDMA administration.

Possible result4: Mice pretreated with 300mg/kg

of pyrazole, NDMA recovery is about 83.7% 60

minutes after the NDMA administration. Also, more

than 80% of NDMA was not metabolized.

Possible result 5: NPC is to formed in pretreated

mice quicker than in control mice.

1.6 Discussion

Pass research shows that NDMA is the main factor to

increase the possibility of NPC (Hakami, Prepageran,

Thubaity, Juaid, Solami, 2018) forming in the human

body. In this research, I found that the rate of

diagnosed with NPC is directly related to the amount

of NDMA in the tissue body and metabolic activation

of the liver. Possible result 1as shown in table 2,

neither NPC nor recovery of NDMA is found in

control mice as it wasn’t injected with any NDMA,

therefore would not cause a reduction of metabolic

activation in the liver.

On the other hand, possible result 2, 3, 4 and 5 as

shown in table 2, are injected with both NDMA and

pyrazole, show appearance of NPC, which prove that

NDMA causes NPC.

The percentage of recovery of NDMA based on

the amount of pyrazole injected, as shown in figure 1,

proves that once the metabolic activation of the liver

is wakened the NDMA is uneasy to be destroyed by

other organ tissues.

The appearance of NDMA in blood, urine and

formation of NPC proves that if the liver is unable to

metalize the NDMA other body tissue would take the

job. The body tissue is unable to clear NDMA out of

the body completely which then cause the rate of

diagnosed with NPC to increase.

Both table 1 and table 3 show that the higher

amount of pyrazole was injected into the mice, the

faster NPC is formed, which proves that NDMA is the

main cause of NPC.

One of the problems in this experiment that may

cause unreliable results is that I haven’t repeat the

experiment and number of mice (4) used are not

enough to make the experiment result accurate.

Because their may be human and environmental

errors (injected too much or too less amount of

pyrazole or NDMA into the mice, timecourse carried

out inaccurately, NPC in mice that’s too small to

detect by MRI, one, two, three or four of the four

mice’s gene that has higher possibility to allow NPC

forming in their body (e.g. heredity) ). Moreover, data

used for experiment prediction have a time gap for 41

years long (1980- 2021), which makes the prediction

inaccurate to the result (The effect of N-

nitrosodimethylamine and metabolic activation in the

liver on the formation of Nasopharyngeal carcinoma).

On the other hand, if the experiment was repeated I

could have collected the result as an average, which

would make the experiment more reliable. Also, if

data are collected from literature written around the

same time prediction to the result (The effect of N-

nitrosodimethylamine and metabolic activation in the

liver on the formation of Nasopharyngeal carcinoma)

would have been more specific. However, colleting

literature from 1980- 2021 about the topic NPC

shows that the scientist are aware of the problem and

the research on NPC has been improving

continuously.

2 CONCLUSION

The result of this study provides doctors with a larger

reference of the cause of NPC in future medical

diagnoses. Future studies will focus on decreasing

amount of NPC taken in by human body through food

by medicines/ health care products.

On average, about 1/100,000 people are

diagnosed with NPC per year all around the world

[less than 1% of children (0-10) are diagnosed with

NPC (Journal of Cancer Research and Practice.

2016). There are two peaks for males to be diagnosed

with NPC: between 30 to 39 and 50 to 59 years old.

Whereas the peak for females is 20-39 years old.

(Beyene, Ketema, Alebachew, Saleh and

Gebremariam. 2021) About 8.8/100,000 people are

diagnosed with NPC per year in southeast China

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(Hamid. 2020); About 32/100,000 people are

diagnosed with NPC per year in Guangzhou and

13.9/100,000 people are diagnosed with NPC per year

in Hong Kong. (Cao,

Simons and Qian. (2011); (Dr.

Anthony C.H. YING. 2008) NPC also has a high risk

of causing death. Around 0.84/100,000 people died

per year, globally. And 31413/100,000 people died

per year in China. (Xu, Zheng,

Zhang,

Zou and

Chen. 2013) Furthermore, the annual cost of cancer

treatment in China (2015) is 221.4 billion RMB (Cai,

Xue, Chen, Hu, Miao, Lan, Zheng and Meng. 2017).

And the population in parts of China being diagnose

with NPC is 3085 in 2009. Therefore, NPC is a

financial burden towards China. Research in the past

has found that salted fish and Siu Mei contain a large

amount of NDMA, (Food and Cosmetics Toxicology.

1981) People from Guangzhou and Hong Kong love

eating these two kinds of food, which explains why

people around these areas have such a high rate of

being diagnosed with NPC comparing to the world’s

probability. Inside the human body, once NDMA is

being detected, the liver would metabolize it out

within a short period, which causes the other organs

not to have enough interactions with NDMA. Causing

the level of NDMA found in peripheral blood lower

than expected (based on total dietary exposure). The

reduction of the metabolic activation in the liver

(IARC Sci Publ. 1980) causes other tissues to be used

to metabolize NDMA. NPC then starts to form

between the head and neck of the human body (Osong

Public Health and Research Perspectives. 2016), as

other tissues can’t metabolize NDMA fully and take

too long. Therefore, I prediced the reduction of liver

ability to clear NDMA will cause NDMA induced

NPC in the human body. This paper talks about the

tumor Nasopharyngeal carcinoma, which is a

common tumor in Asia of the head of the neck with

originates in the nasopharynx (Osong Public Health

and Research Perspectives. 2016); (Cao,

Simons and

Qian. 2011). In this work, I investigated: ‘The effect

of different levels of N-nitrosodimethylamine in the

body tissues and metabolic activation in liver on the

rate of formation of Nasopharyngeal carcinoma to

increase.’. (IARC Sci Publ. 1980) The experiments

will use Male Ydd strain mice, which will be injected

with increasing amounts of NDMA as the experiment

measures liver function through assay. An increasing

amount of pyrazole will also be injected, for speeding

up the reduction of metabolic activation in the liver.

(Metabolic activation means the chemical conversion

of a relatively benign substance into a more

hazardous one by normal biochemical processes in

cells and tissues.)

A timecourse experiment is then performed, for

measuring the amount of NDMA in the liver and body

tissues. Also, the time takes for NPC to form under

different dosages of pyrazole. The amount of NPC

formed is measured using an MRI. There are three

most possible results: (1) Mice pretreated with a

larger amount of pyrazole would have a higher

recovery of NDMA administration; (2) pH value

would decrease administration of pyrazole; (3) NPC

is to form in pretreated mice quicker than in control

mice.

Generally, this study explores the effect of

NDMA on the formation of NPC. The result of my

study will indicate how the rate of formation of NPC

is effected by amount of NDMA in the human body.

The possible results on NPC shows that the larger

amount NDMA intaking the worse the metabolic

activation is being damaged. FEHD as an example,

should stop business selling food that contain high

amount of NDMA, as Hong Kong is one of the place

that has high rate of population diagnosed with NPC.

And in the future study, scientists can focus on the

prevalence of NPC in Asia, and investigate the

connection between the dietary habit of Asia and the

formation of NPC.

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The Effect of N-nitrosodimethylamine and Metabolic Activation in the Liver on the Formation of Nasopharyngeal Carcinoma

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