Journal of Pharmacy and Pharmacology 10 (2022) 62-65

doi: 10.17265/2328-2150/2022.02.005
D DAVID PUBLISHING

Estimating Age of Saliva by Enzyme Activity at Different

Temperatures

Komal Dave1, Manjushri Bagul1, Smitesh Nalage1 and Suraj Jadhav2

1. Department of Forensic Science, Yashavantrao Chavan Institute of Science (Autonomous), Satara, Maharashtra, India
2. Department of Nanoscience and Technology, Yashavantrao Chavan Institute of Science (Autonomous), Satara, Maharashtra,
India

Abstract: Saliva is a product of exocrine gland, which are release their secretions (Saliva) into ducts that lead to mouth.Salivary
amylase is a produced by the extrinsic type of salivary glands. Amylase is breakdown the starch molecules into glucose and maltose.
Saliva is most important forensic evidence, which can provide major information about the relation between the victim and offender.
Based on above method for salivary amylase activity can be concluded that given saliva sample has most activation at 37 °C and 2
hours on the basis of the amylase activity results will help to identify the age of saliva sample. At lower temperature, the salivary
amylase enzyme is deactivated and higher temperature the enzyme is denatured so most time will be taken by salivary amylase to
digest the starch of lower and higher temperature.

Key words: Salivary Amylase, denaturation, age estimation, forensic evidence.

1. Introduction In oral cavity salivary, the enzyme quantity for two 40

to 50% of the full salivary glands produces protein
Saliva is produced from pairs of salivary glands. An
and is secreted by acinar cells of paratroid glands [2].
individual produces between 1.0 and 1.5 liters of
Previous result findings shows that amylase activity
saliva per day. Saliva is composed of water with few
levels and DNA concentration in saliva vary greatly
amounts of enzymes and electrolytes. Saliva is secreted
between individuals hence, there is no significant
by parotid, submandibular, sublingual salivary glands
correlation between the two. Amount of DNA in
present in the mouth. Saliva samples can be examined
saliva depends on shedding of buckle cells in the oral
from many types of medium such as body parts,
cavity. They observed inter personal range of amylase
papers, envelope, glass bottles, metal cans, cigarettes
activity found is comparable with previous result.
butts, plastic etc. Saliva is the chief evidence which
Saliva screening using amylase can help in analysis as
can provide major information about the personal link
body fluid. Salivary α amylase has been proposed as a
between the victim and offender [1]. Presence of
sensitive non invasive bio marker for stress induced
saliva can be found out by the starch Iodine and
changes in the body that reflects the activity of the
phoebes test. Starch Iodine and side a bus test do not
sympathetic nervous system [3]. The effect of stress
certify the presence of human’s saliva. This test is
induced changes on salivary Alfa amylase level in
only to certify or find out whether the amylase has
different age groups in last studies. Documents
come from human or non-human. In the presiding
frequently come across in criminal cases often time lot
studies, it was studied with that saliva alpha amylase
of people moist their finger tips with saliva while
is calcium containing metal enzyme that hydrolyzed
turning pages [4]. Taking into consideration, a
the alfalfa 14 linkage of starch to glucose and maltose.
research experiment was designed to take DNA edge
of the behavior to possibilities establish the handler of
Corresponding author: Smitesh Nalage, assistant professor,
research fields: forensic science. Email: nalagess@ycis.ac.in. the document from foregoing works on documents
 Estimating Age of Saliva by Enzyme Activity at Different Temperatures
was related to touch. The recent study has to do with
the determination of habit of using saliva for flicking
the pages. The main objective of study is to discover
whether saliva stains are likely to relent DNA
profiling from various paper substrate [5].
Amylase levels in body fluids are reported, and it is
shown that high amylase levels may occasionally
occur in body fluids other than saliva. Low amylase
levels are reported in a saliva component which has
been previously described as being rich in blood
group active material. The use of amylase activity in
the localization and identification of saliva is
discussed in detail and guidelines for the interpretation
of results are proposed. This is the significance of
amylase in forensic investigation of body fluids [6].
The unequivocal tissue identification in forensic
casework samples is a key step for crime scene
reconstruction. Just knowing the origin of a fluid can
sometimes be enough to either prove or disprove a
fact in court. Despite the importance of this test, very
few data are available in literature concerning human
saliva identification in old forensic caseworks. In this
work the stability of human α-amylase activity in aged
samples is described by using three different methods
integrated with DNA profiling techniques. This
analytical protocol was successfully applied on
26-years old samples coming from anonymous threat
letters sent to prosecutors who were working on “the
Monster of Florence”, a case of serial murders
happened around Florence (Italy) between 1968 and
1985 [7].
The specific activity (enzyme activity/protein
concentration) of amylase was determined for the
identification of saliva stains. The specific activity of
amylase in saliva stains rapidly decreased during the
first hour but, from 1 to 28 days, this decrease was
much less when the stains were kept at room
temperature [8]. Stains of various human biological
materials, breast milk, nasal secretion, meconium and
vaginal secretion showed comparatively high amylase
activity, but the saliva stains could be differentiated by
63
their high specific activity of amylase, over 2 I.U./mg.
When saliva stains were contaminated with blood or
vaginal secretions at various ratios, the specific
activity of amylase decreased with increase in the ratio
of contaminant, especially when the contaminant was
blood. However, the specific activity of amylase was
still higher than 2 I.U./mg even after 1/5 volume of
blood was added or after five volumes of the extract of
the stains of vaginal secretions was added [9, 10]
Gender differences in the function and anatomical
features of salivary glands are well known. However,
specific gender differences in the biochemical
composition and salivary flow rate (SFR) remain
uncertain. Collection methods affect the assessment of
the salivary composition and SFR, which are also
highly affected by acid stimulation [8]. Stimulation, as
measured by 3 different collection methods
sequentially [11].
2. Discussion
Saliva is complex body fluid and trace evidence in
forensic science and recovery of traces of body fluids
is the one of the most important types of forensic
science. Saliva plays a critical role in forensic
evidence. Saliva may get deposited on skin in
instances of biting, licking or sucking and can act as a
significant source of evidence [12, 13].
Drug abuse tests, such as poisoning, animal bites,
alcohol abuse, hormone testing. Saliva stains can be
presents on cloths, paper, and other inanimate object.
So that helps to DNA analysis. Salivary biomarkers
are also used in cases involving bite-marks for general
identification of people and victim identification in the
instances of mass disasters. By performing the
enzymatic activity assay this Salivary Amylase will
helpful to find out the age of the saliva stain since
death [14].
In some cases, on the basis of salivary amylase
activity can be concluded that given saliva sample
have most active at 37 °C temperature and 2 hr time.
At lower temperatures, the salivary amylase enzyme is
64 Estimating Age of Saliva by Enzyme Activity at Different Temperatures
deactivated and higher temperature the enzyme is
denatured. As a result, salivary amylase takes longer
to digest starch at higher or lower temperatures. On
the basis of the Amylase Activity Assay results will
help to identify the Age of Saliva sample [15].
A conclusion can be drawn out that majority of
subjects under study were in habit of using saliva turn
pages, and this can be an important evidence to help
criminal justice system as DNA profiles could be
developed successfully from the substrates. This can
be very good evidence in respect of identifying the
individual(s) who handled the document [16].
Substantial gender differences in biochemistry and
flow of saliva exist, and these findings are robust, as
evidenced by reasonable consistency of the data
among different saliva sampling methods [17, 18].
Saliva as a body fluid has numerous functions in the
body which not only shield human body from foreign
particles, but also acts as a potential route for the
increase of diseases. The pandemic of COVID-19
should strengthen the awareness of endodontic on the
transmission risk during dental practice. Additional
protective measures should be taken at this special
moment to reduce potential saliva derived danger to
both of the dentist and the patients [19, 20].
Another interesting use of salivary analysis is in
forensic science. Retrieving saliva, either directly
from an accused or from the injuries sustained by the
victim, can facilitate the analysis of the DNA. In order
to collect saliva sample cotton swabs are used. Even
dried saliva from bite wounds suffered by the victim
can be analysed using an amylase assay kit.
3. Conclusions
From previous findings we conclude that neither
plasma osmolality nor saliva osmolality correlated
closely with BML associated with mild dehydration
and especially in males shows potential as a
dehydrations screening biomarkers changes in whole
saliva osmolality (sosm) compares with changes in
plasma osmolality during mild dehydration as
measured by BML saliva osmolality increased from
pre (73.29 ± 15.42) to post exercise (102.10 ± 25.28)
and have greater percentage than posm. In most of the
cases particularly when comparing candy wet loss
with stimulated parotid and sub mandibular and sub
lingual saliva. The candy weight loss test is a rapid
measure of salivary hypo function which co-relate
saliva output and reports of subjective dry mouth.
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