Description of Liver Histology of Mice (Mus musculus L) after Giving

Nano Herbal Haramonting (Rhodomyrtus tomentosa)

Putri Cahaya Situmorang

and Syafruddin Ilyas

Department of Biology, Faculty of Mathematics and Natural Sciences,

Universitas Sumatera Utara, Medan, Indonesia, 20155

Keywords: Nanoherbal, Rhodomyrtus tomentosa, Liver.

Abstract: The objective of this study was to determine the description of liver histology of male mice (Mus musculus

L.) after giving Nano Herbal Haramonting (Rhodomyrtus tomentosa). This research use the Completely

Randomized Design (CRD), which consists of six groups of CMC 0.5% and Nano Herbal Haramonting

(R.tomentosa) at dosage of (100; 141,42; 200; 282,82; 400)mg/20g body weight. The livers were made into

preparations by using the paraffin method and Hematoxylin Erlich-Eosin staining (HE). The value of each

cell with Manja Roenigk's histophatolgy model. Nano-R.tomentosa administration was not significantly

different in body weight of mice (P>0,05), but It was significantly different to weigh of liver (P<0,05). The

results of histological liver showed that there is a significant difference (P<0,05) between control group and

treatment groups with damaged cells liver of male mice Mus musculus L.) after giving of Nano Herbal

Haramonting (R.tomentosa).

1 INTRODUCTION

Nanoherbal is the herb that uses nanoscale technology

(wave power) and use quality herbal raw materials.

The use of herbal remedies has been practiced for

thousands of years and a part of several countries such

as Indonesia, China and India. Nanotechnology

commonly refers to structures that are up to several

100 nm in size which can be increased up to 1000 nm.

Nanotechnology and nano science studies have

develops briskly during the past years in a large range

of product results. It gives opportunities for the

development of materials, including those for medical

applications, where conventional techniques may

reach their limits (Ratnam et al, 2006). Herbal

medicines have less adverse effects as compared with

modern medicines due to poor solubility, poor

permeability, low bioavailability, instability in

biological milieu and extensive first pass metabolism

by developing new formulation as nano herbal

medicines like nanoparticles, dendrimers, Nano

crystals, Quantum dots, Nanosperes, Nanocapsules,

herbal market get good feedback (Rinku et al, 2018)

One study using nano herbal is Nano curcumin. In

this study Nano-curcumin significantly inhibited the

growth of MCF-7 breast cancer cell line and resulted

in synergism cytotoxicity effects (Parisa et al, 2018)

Haramonting (R.tomentosa) is an ornamental,

evergreen shrub grows up to four meters. This plant

species is native to southern and southeastern Asia

(Awinita et al, 2005). The potential of R. tomentosa

as a new source of health-promoting compounds such

as dietary fibers, essential fatty acids, and phenolic

compounds. A total of 19 phenolic compounds were

tentatively characterized, including stilbenes and

ellagitannins as major components, followed by

anthocyanins, flavonols, and gallic acid. Piceatannol,

a promising health-promoting stilbene component,

was the major phenolic compound found in R.

tomentosa fruits (Lai et al, 2013).

Hepatic sistem is the major organ system involved in

the metabolism, detoxification and excretion of

various endogenous and exogenously. Liver damage

is always asssociated with necrosis, Strengthening of

inbuilt protective mechanism or exogenous

administration of antioxidant may be useful in

protecting the liver (Pramodh et al, 2008). In vitro

antioxidant activity of the different extracts of R.

tomentosa has been reported by different methods

(Geetha et al, 2010). Natural antioxidants are known

to exert beneficial effects in hepatitis induced by

antitubercular agents (Limsuwan et al, 2009).

982

Situmorang, P. and Ilyas, S.

Description of Liver Histology of Mice (Mus musculus L) after Giving Nano Herbal Haramonting (Rhodomyrtus tomentosa).

DOI: 10.5220/0010083409820986

In Proceedings of the International Conference of Science, Technology, Engineering, Environmental and Ramiﬁcation Researches (ICOSTEERR 2018) - Research in Industry 4.0, pages

982-986

ISBN: 978-989-758-449-7

Oxidative stress induced damage to hepatocytes has

been found to have a key role in antitubercular drugs

induced hepatitis (Shakun and Shmnan’ko, 1986). R.

tomentosa extract demonstrated its free radical

scavenging effects in concentration dependent

manner and the results suggest that R. tomentosa

extract can serve as a potent antioxidant (Sodhi,

1998). So in this study, we examined the effect of

Nano- R.tomentosa to liver.

2 METHODS

2.1 Materials

The material used is male mice (Mus musculus L)

strain DDW, Haramonting leaves (R.tomentosa)

obtained from plantation residents in Tapanuli North

Sumatera, Feed the mice no. PB 551, staining

Hematoxylin and Eosin. This research use the

Completely Randomized Design (CRD) using 30

male mice with an average weight of 20-25g age 12-

18 weeks. Male mice are kept in Animal Cages

Biological Laboratory, Faculty Mathematics and

Natural Science, Universitas Sumatera Utara.

Experimental animal handling is done ethically

(Ethical Clearance).

2.2 Making of Nano- R.tomentosa

R.tomentosa leaves are washed, then dried in

accordance with the requirements of water content

using high energy milling (HEM), then Simplicia as

the destructive medium is inserted into the jar

container, Inserting balls with larger diameter size

then continued by inserting small balls and the last

sample. The total volume of the balls and the samples.

entered does not exceed 2/3 of the volume of the jar.

The usual Ball to Powder Ratio (BPR) is 20:1, 10:1,

8:1. Example BPR 20:1 means 1 gram of sample then

milled with 20 grams of ball weight. Jar that has been

filled with the ball, samples are closed tightly then

mounted on the jar inside the HEM tool, then HEM is

turned on for 2 hours.

2.3 Treatment and Observation of

Hystology

The treatment consisted of 6 groups consisting

of 5 male mice ie CMC 0.5% and Nano Herbal

Haramonting (R.tomentosa) at dosage of (100;

141,42; 200; 282,82; 400)mg/20g bw in 14 days, then

killed by the method of disclasio cervicalis to take

the liver. Preparation of histologist by paraffin

method and Hematoxylin Erlich-Eosin staining (HE)

to observe the damage to liver cells due to the

administration of Nano-R.tomentosa. Mixture is

observed with magnification 400x. In each field of

view counted 20 cells at random and in the value of

each cell with Manja Roenigk's histophatolgy model

(1: Normal, 2: Parenchymatous Degeneration, 3:

Hydrophic Degeneration 4: Necrosis) then the data

processed with SPSS 22 program with Kruskal Wallis

test.

3 RESULT AND DISCUSSION

The result will be discussed in 3 subsection i.e

average weight of body, weight of liver and damage

of liver cells.

3.1 Weight of Body

Figure 1. Average of weight of male mice after giving

Nano-R.tomentosa. (

± SD).

The results of average body weight of mice

CMC 0,5% group has the highest average weight

while the lowest weight is in T1 (Figure 1). Nano-

R.tomentosa administration was not significantly

different in body weight of mice (P>0,05). Nano-

R.tomentosa does not affect the weight of male mice,

Allegedly due to outside variables that can not be

controlled such as the psychological condition of the

mice that is affected by the surrounding environment,

repeated treatment, different appetite and fights

between mice.

3.2 Weight of Liver

The result of statistic test P<0,05 showed that Nano-

R.tomentosa administration at each treatment was

significantly different in weight of liver except in

Control group with T1, T3,T4 and T5, T1 group with

T4 and T5, T2 group with T3, T3 group with T4

Description of Liver Histology of Mice (Mus musculus L) after Giving Nano Herbal Haramonting (Rhodomyrtus tomentosa)

983

(P>0.05) . T5 group has the highest average liver

weight while the lowest liver weight is in T2 (figure

2). T1 higher than T2, and also T1 higher than control

is assumed because the outside variable is difficult to

control such as weight of body, different appetite and

activity of mice.The liver is the largest organ in the

body that is involved in the body's metabolism,

neutralize toxins and toxic substances through the

detoxification process by cell kupfer in the body. So

that the weight of the liver can be increased or

decreased due to the entry of excessive toxic

substances in the body.

3.3 Damage of Liver Cells

Table 1. Average Hepatocyte Normal and degree of damage (Parenchymatous Degeneration, Hydrophic Degeneration and

necrosis) after giving Nano-R.tomentosa (

± SD).

Figure 3. Histology of Liver, a: Normal, b: Parenchymatous Degeneration, c: Hydrophic Degeneration, d: Necrosis (400x).

The results P<0,05 showed cell damage due to

nano-R.tomentosa administration is significantly

different from each treatment except Control group

with T1 (P>0.05). Description of liver damage is

different for each treatment (figure 3). Damage with

Parenchymatous degeneration occurs lowest in CMC

Treatmen

Normal Parenchymatous

Degeneration

Hydrophic

Degeneration

Necrosis

CMC

0.5%

14.40 ± 1.66 4.56 ± 1.47 4.20 ± 3.00 7.52 ± 4.05

T1 13.60 ±1.50 4.88 ± 1.42 5.16 ± 2.81 9.12 ± 4.09

T2 12.30 ±1.14 5.52 ± 1.94 12.5 ± 2.48 10.24 ± 4.01

T3 6.76 ±1.51 6.80 ± 2.00 12.5 ± 4.30 22.72 ± 8.54

T4 4.84 ±1.52 7.76 ± 2.60 11.5 ± 3.32 29.76 ± 8.01

T5 3.40 ±1.35 6.80 ± 2.71 9.48 ± 3.84 40.16 ±8.06

Figure 2. Weight of liver after giving N

ICOSTEERR 2018 - International Conference of Science, Technology, Engineering, Environmental and Ramiﬁcation Researches

984

CMC

0.5%

T1 T2 T3 T4 T5

Percentage of cells

damage

0,5% (Control) and highes in T4 (table 1) due the

mice are also given aquades so that the cells can

regenerate. cells in liver organ with parecimatous

degeneration may improve, cells undergoing necrosis

over time will be replaced with new liver cells due

cell regeneration process in liver organ (Geetha et al,

2012). Parenchymatous degeneration is the mildest

degenerate levelIn parenchymatous degeneration

cells, granules are found in the cytoplasm due the

precipitate that causes the cytoplasm to become turbid

and swelling of the cells. Hydrophic Degeneration is

low in control, and increases in T2 and T3 (table 1).

This degeneration is more severe damage, there are

vacuoles containing water and cytoplasm that do not

contain fat and glycogen. This change is generally a

result of metabolic disorders such as hypoxia or

chemical poisoning. This degeneration is also

reversible although it may be irreversible if the cause

of the injury persists. The process of Necrosis

increases from Controls to P5 and Normal cells

decreases from control to treatment level (table

1)(figure 3). At each dose level, the toxic ingredients

in the liver are getting out of process, causing

parenchymatous degeneration, hydrophic

degeneration and necrosis in liver. The target of a

toxic substance in the body is the molecular structure

of transport of bile acids, membranes, intracellular

fats, proteins and nucleic acids. As a result the target

molecule becomes a non-functioning unit and may

activate secondary pathways such as apoptosis,

necrosis, autofagocytes and mitochondrial disorders

and other immunological reactions (Kandena et al,

2011).

Figure 4. Percentage of cells damage.

The degree of damage is known that the

percentage of liver damage from control to treatment

with high doses continue to be damaged (figure 4).

R.tomentosa leaves contain secondary metabolites of

phenol, flavonoids, saponins, tannins, steroids and

triterpenoids. Phenols, flavonoids, tannins, steroids

and triterpenoids that have an antioxidant effect.

Saponins can cause haemolysis by affecting the lipid

bilayer in the protein membrane of the red blood cells

causing the formation of pores in the red blood cell

membrane (Kaplowitz, 2002)(Baumann et al, 2000).

Damage of liver cells is thought to be caused by

saponins, and tannins in nano-R.tomentosa who

accumulated and irritant or toxic. Sentrolobuler

damage in liver due tannin and saponin compound

administration in research that spans short time,

usually seen cell swelling, necrosis to cause death in

mice. So the liver loses its function by no longer able

to change the compounds that are very toxic to be less

toxic.

4 CONCLUSIONS

Cell damage due to Nano-R.tomentosa administration

is significantly different from each treatment

(P<0.05) and Giving Nano herbal Haramonting

(Rhodomyrtus tomentosa) with excessive doses can

cause a decrease in hepatic weight and damage to

liver cells, So the liver loses its function especially the

detoxification function of toxic.

ACKNOWLEDGEMENTS

We are grateful to Directorate of research and

community service, Directorate general of research

and development, Ministry of research, Technology,

and Higher Education in accordance with research

and community service funding agreement for

budgeting year 2018 (Fund of the research

postgraduate team I) to funding our research.

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