Chapter 1

Introduction
1.1. Background

Optical Methodologies are burgeoning for biomedical and environmental applications for
its non-invasive and high-resolution analysis of the object under investigation. Different
optical methods have been developed based on the application of optical laws. Such laws
include mechanisms of optical radiation, reflection, the interaction of light with a substance,
etc. The optical methods of analysis include Molecular-absorbing spectral analysis, Atomic
spectroscopy, Refractometric analysis, and Fluorometric analysis. The molecular-absorbing
spectral analysis deals with the measurement of the luminous flux decay due to selective
absorption of light by the analyzed substance whereas atomic absorption spectroscopy uses
the spectra of radiation or absorption of the substance after the substance is transformed into
an atomic state by the external high-energy impact. The refractometric analysis technique
has been analyzed on the light refraction index versus nature and concentration of the
substance. The fluorometric analysis has been formulated based on the emitted light intensity
of the substance versus its composition. The prominent examples of optical spectroscopy,
which include methods like Raman spectroscopy and photoemission spectroscopy.
Photosensitive measurement technologies have been widely accepted in industrial, medical,
and environmental fields because of their non-invasive and high-resolution characteristics.
In the spectroscopy-based methods, the spectral information has been accounted for the
parameters under test. For biomedical diagnosis, the use of optical spectroscopic methods is
primarily in vitro preclinical diagnosis. The other uses include non-invasive diagnosis from
the skin surface and minimally-invasive diagnosis. The upgrading of nano-materials
improves the sensitivity of the molecular probes and photonic systems, hence multiple
medical parameters have been rapidly diagnosed and analyzed. The modern-day challenges
include the design and realization of affordable techniques without compromising the
sensitivity as well as the overall performance of the device. The spectroscopic investigation
accomplishes an important role both in diagnosing different diseases including anemia,
jaundice, oxidative stress and cancer, and in the fields of environmental solutions like

1
detection of ions, soil analysis, etc. Interaction of light with matter always fascinates human
beings since ancient times. Despite the understanding of color generation upon the
interaction of light with the matter, the term 'spectroscopy' was originated in the 17th century
while the dispersion of the white light was observed according to its wavelength by the
prism. Spectroscopy in medical science has been popularised in the early 20th century.
However, its magnificent presence in the diagnosis of the disease is still evident since the
earliest times. In ancient times the medical diagnoses were based on audial and visual
interpretation. According to the ancient Greek and Roman physicians, all diseases are caused
by the imbalance among four humors [1,2] namely blood, yellow bile, black bile, and
phlegm. Uroscopy [3], a visual inspection of urine was practiced to the diagnosis of disease
or disorder. The invention of the microscope was a revolution to understand visually the
tissue structure and the activities of the organisms that cause diseases [4].

Optical spectroscopy seems likely to be an effective screening tool for diagnosing

different physiological conditions like hemoglobin level, jaundice at their early stages. In
this thesis work, different optical methodologies based on the combinations of photonic
technologies have been applied. Marinating sustainable growth in agriculture, development
of the sensors and affordable devices have been essentially required to monitor
environmental balance. A wide spectrum of ions has been extensively investigated to address
the emerging need. In this thesis work, devised have been developed based on the fabricated
nano-sensor for quantitative detection of iron and fluorides in biological and environmental
samples.

1.2. Introduction

The recent advancement of light sources, optical detectors [5], and fiber probes improve
the quantitative measurement of the interactions between the molecules and light, which
yield information about the biochemical, structural, and pathological conditions of the
tissues [6]. The basic changes are in the way or the technique of measuring parameters with
medical relevance [7]. The interaction of light with biological medium covers the general
field of biomedical optics which has its role in developing different modern clinical
diagnostic techniques. In the following section, relevant spectroscopic techniques with

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potential applications in biomedical diagnosis and environmental monitoring have been
discussed.

1.2.1. Optical Methodologies in Biomedical Application: Application of optical

methodologies (spectroscopy) in the biomedical field ranges from pulse oximetry to non-
invasive optical biopsy. In this section, the newest ideas and strategies based on different
spectroscopic techniques for the early diagnosis of different diseases have been incorporated.
Absorption spectroscopy has been employed as an important tool to determine both the
presence and the quantity of a particular matter present in a sample under test. The amount
of light transmitted due to its interaction with the sample is the function of the wavelength.
UV/Vis absorption spectroscopy has been used in the clinical laboratory for many years.
Absorption spectroscopy is well applicable in the setup where the molecule under test has
been dissolved in a transparent solvent [8]. It has been well described in Beer-Lambert Law
that the quantitative estimation of solute concentration is possible as it changes linearly with
the absorbance. The transmission strength and the wavelength of the light depend upon the
chemical nature of the solute and its environment. Absorption spectroscopy is, therefore, a
useful method to interpret ligand-binding reactions and conformational shifts in biological
matters [9]. Being non-invasive, the devices developed on absorption spectroscopy have
been successfully used in point of care service. Pulse oximetry is one most popular tools
used for the measurement of oxygen saturation (SpO2) in point of care diagnostics. A blood-
oxygen meter is another example which measures the amount of oxygen that the blood
carrying. An arterial blood gas (ABG) test [10], and pulse oximeter both can measure the
percentage of arterial oxygen at point-of-care settings. It has been reported that the
acceptable SpO2 level for healthy lungs falls between 95 and 100 percent. Based on the Beer-
Lambert law a pulse oximeter utilizes two light-emitting diodes (LEDs) of wavelength
660nm and 940nm and a photodiode. It has been observed that in the presence of oxygen
molecules the affinity of hemoglobin towards light absorption varies. The optical density of
hemoglobin has been found more in the infrared region and comparatively less in the red
light region. The calculated ratio of the intensities at 660nm to the 940nm has been
transformed to SpO2 via a lookup table [11]. Estimation of glycated hemoglobin (HbA1c) is
also determined using absorbance spectroscopy, without involving the use of external dyes
or reagents. The optic fiber-based instrument has been used for recording absorption spectra

3
in the spectral range of 200-850nm of glycated hemoglobin. The optical densities of the
glycated hemoglobin have been verified with the standard ion-exchange HPLC method and
found the aforementioned technique may be employed as a supplementary technique to other
techniques that already exist [9]. Instruments based on atomic absorption spectroscopy
(AAS), inductively coupled plasma-atomic emission spectroscopy (ICP-OES) and flame
atomic absorption spectroscopy (FAAS) are popular to determine the concentration of the
electrolytes in biological fluids like blood serum and urine [12], and pharmaceuticals [13-
15]. The atom is the smallest unit composed of one nucleus and one or more electrons. In
the incidence of external excitation, the electron jumps from a lower energy state to a higher
state and absorbs a photon of a characteristic wavelength. In AAS, the sample has been kept
in a graphite tube and then heated electrically, whereas in FAAS a burner has been used. The
purpose of using either the burner or the graphite tube is to atomize the sample under test.
The beam of electromagnetic radiation emitted from a hollow cathode lamp has been passed
through the atomized sample. The radiation has been absorbed by the atoms in the sample
has been found linearly proportional to the atom numbers.

Fluorescence spectrometry is a powerful technique for accurate profiling of specific

biomolecules. It is primarily concerned with electronic and vibrational states of the
fluorophores. Fluorescence spectroscopy has been evidenced as a reliable technique for
investigating various illnesses including premalignant and malignant lesions [16,17].
Fluorophores used for medical diagnostic studies are likely intrinsic and extrinsic [18,19].
Endogenous fluorophores [20] are the source of autofluorescence, [21] used for live-cell
characterization and cell sorting. Examples of clinically used fluorophores are collagen,
nicotinamide adenine dinucleotide (NADH), tryptophan, elastin, porphyrins, and flavin
adenine dinucleotide (FAD), etc. Among all, both NADH and FAD are natural fluorophores
that have been considered as potential biomarkers for both metabolic and mitochondrial
activities [19,22]. Collagen and elastin are two fluorophores mainly exhibits changes in the
spectra within the tissues because of their intrinsic property [23,24]. Exogenous fluorophores
have been used for the samples having no fluorescent properties. The addition of extrinsic
fluorophores change the spectral properties of the sample, which in turn help to determine
the disorder in the sample under test. In this context exogenous near-infrared fluorophores
(NIRF) have been fabricated with small organic fluorophores and nanoparticles. In cancer

4
diagnosis NIRF imaging has been identified as a promising tool because of its imaging
modality [25,26]. Dansyl, fluorescein, and rho-damine are the few examples of exogenous
fluorophores. The malignant tumors have been identified using steady-state UV fluorescence
spectroscopy of benign and normal tissues [23] and the technique thus recognized widely as
the disease detection tool. Stimulating the tissue samples under test at 300nm, the emission
has been observed in the range of 320nm to 550nm and found to be dissimilar for cancerous
tissues from that of normal and benign tissues. Frequent use of such fluorophores has been
limited as UV light is toxic for the living systems. Additionally, it is hard to discriminate the
fluorescence signals obtained from these tags and cell autofluorescence. The concept of
synthetic fluorophore [27] has been introduced in 1942 and DAPI (4’,6-diamidino-2-
phenylindole) was the first fabricated fluorophore. In this context, time-resolved
fluorescence is a strong technique to check the heterogeneous nature of chromophores that
are responsible for the tissue fluorescence. In the device, the fluorescence relaxation decay
obtained from photo-excited tissues has been considered to be the measuring parameter.
Different exponential decay profiles with slow and fast components obtained from the
fluorescence kinetics confirm the malignant and non-malignant condition of the tissues
respectively and therefore has been considered a novel methodology to acquire the
fundamental evidence of cancer.

Laser-induced fluorescence (LIF) spectroscopy, a non-invasive real-time technique [28]

that has been used to study the structure of molecules. LIF has been recognized as a species-
selective spatially resolved detection technique. The laser of specific wavelength has been
used to excite the molecules of the sample under test to the higher energy level. The
molecules emit light once they start de-excitation. The LIF spectral profiles of carcinogens
have been found significantly different from normal tissues [28,29].

Raman spectroscopy is another powerful analytical method to determine the chemical

structure of living tissues [30]. Any disorder in the tissue leads to significant variations in
the Raman spectra. The changes, therefore, can be used for diagnostics, prognostics, or as a
tool for evaluating new therapies. Besides chemical and molecular analysis, Raman
spectroscopy is relatively new to address the biomedical problems by advancing technology,
particularly lasers and detectors. The application of Raman spectroscopy in medical

5
diagnosis [31] includes invasive detection of cancer [32] (brain, breast, lung, skin, prostate,
colorectal); minimally invasive bio-fluid analysis for the detection of asthma, inflammatory
response, coagulant and anticoagulant factors in human blood, malaria, etc.

1.2.2. Optical Methodologies to Monitor Environmental Issues: Application of Optical

methodologies (spectroscopy) in medical science has not been limited to medical diagnosis
only. Spectroscopy has a significant role in environmental monitoring applications including
the screening and estimation of ions like iron and fluoride from groundwater.

Though nanoparticles are indirectly related to spectroscopic studies, thus the working of
the nanoparticles can be confirmed utilizing spectroscopic methods only. Hence, applying
the spectroscopic methods, the ions present in environmental and biological samples can be
screened in the presence of specific nanosensors. However, in some cases, the effective
application of the nanoparticles could be enhanced through the application of external light
or magnetic field [33]. The colloidal particles with small size in the order of a nanometer
and large surface area have been used as sensors to estimate essential ions present in the
environment.

Thus, the optical methodology might be a useful tool to investigate the relevant structure
and functionality of the biological as well as environmental systems. The ambition of the
thesis is to improvise different optical methodologies to develop devices with extended
sensitivity, specificity, and spatial resolution for the betterment of human life.

1.3. Scope and Objective

The state-of-the-art of the existing technologies for screening anemia in terms of

hemoglobin count and the average cell morphology varies from low-tech to high-tech
depending on the target resource setting [34]. The measurement hemoglobin count is usually
executed by blood sampling [35] and tested in a standard laboratory with sophisticated
instruments. Such instruments are developed on either electrochemical impedance
spectroscopy [36,37] or Coulter principle (e.g. automated hematology analyzer) [38]. The
aforesaid instruments are effective but involve complex and expensive experimental setup,
operational technical skill, and time-consuming measurements. In this context, a semi-
quantitative gravimetric copper sulfate method is useful for hemoglobin count

6
determination. HemoCue [39,40] is an example that implements the gravimetric copper
sulfate method that comes in a portable format and associated recurring cost. In addition,
many non-invasive methods have been developed for the measurement of hemoglobin count
[41-43] including Pulse CO-Oximeter, Pronto-7TM monitor (version 2.1.9, Masimo
Corporation, Irvine, USA), NBM-200MP monitor (Orsense, Ness Ziona, Israel) and
spectrophotometer [44]. However, the characteristics of these instruments depend on
different physiological factors like arterial pulse, body movements, skin tones, etc. A survey
reveals that aforesaid instruments are an accurate, economical, and portable device for
anemia diagnosis and suitable in low resource territories. In recent times some algorithms
have been deployed in smartphones with the associated add-on for blood analyses and
determination of hemoglobin concentration [45]. One such example is the use of disposable
plastic cuvette-based add-on developed by Prof. Ozcan and his group from the University of
California, Los Angeles [46] for the hemoglobin detection using processed whole blood. In
absorption spectroscopy a sample with a very high extinction coefficient needs significant
dilution, to avoid a systematic error in the measurement. In the experimental set-up used by
the referred group does not use any reference wavelength for the differential or ratio-metric
calculation for hemoglobin concentration. The application of differential or ratio-metric
calculation offers better accuracy compared to that of the absolute result in optical
measurement. In the present context, a reliable, minimally invasive portable device strategy
on untreated whole blood has been addressed in the project for hemoglobin concentration
estimation at point-of-care.

Recent fact sheets have been updated by the World Health Organization (WHO) on global
statistics of different types of hepatitis. According to the report, 400 million and above
people were detected of deadly liver infection and every year more than 1.3 million people
decease due to either critical or chronic consequences of liver malfunction. The child death
due to liver breakdown is highly alarming according to the global statistics. It has been stated
in the report from UNICEF (2012) that in most underdeveloped/developing countries 21
children per minute die of neonatal jaundice which is preventive in nature. Jaundice is a
condition of high bilirubin levels in the blood which is seen as the yellowish appearance of
the skin and conjunctiva [35,47]. The presence of excess bilirubin indicates liver disease like
hepatitis or liver cancer [48]. Jaundice due to hepatitis E in neonatal and maternal subject

7
can be prevented and cured if diagnosed earlier. The raising of bilirubin concentration and
its oxidative products in adults cause several serious diseases [49,50]. As consequences of
elevated bilirubin levels in the neonates, Kernicterus [51,52] damage the white matter of the
brain [53] which causes athetoid cerebral palsy and hearing loss. Making the diagnosis to be
non-contact and non-invasive for neonates is one of the biggest challenges as contamination
is very feasible. A non-invasive and non-contact device has been addressed in the projects
to screen neonatal hyperbilirubinemia.

Monitoring of any biological parameters using an instrument, with different physiological

conditions, end users have to be sure that the equipment provides accurate measurement. In
this background, appropriate monitoring of the bilirubin count in newborns associated with
clinical risk factors including those who receive phototherapy [54] is significantly important
for the appropriate management of hyperbilirubinemia [55] more accurately identifies. The
performance of the developed non-contact transcutaneous screening device has been
investigated with the reference of the conventional total serum bilirubin test in the Asian
population of different gestational ages under prescribed settings along with the effect of
different risk factors and phototherapy.

Among all the available electrolytes, sodium and potassium are found to be the most
common analytes because of their substantial presence in the human body as well as in
nature. Despite the significant clinical demands to measure the concentration of both the
electrolytes, fewer analytical principals are available to meet the need. The ion-selective
electrodes is a widely accepted method for the measurement of electrolyte concentrations
using serum or other biological fluids as a sample for the test. Beside ISE, some other
techniques like flame photometry and AAS are also used for the detection of the electrolyte
concentrations [56-59]. Earlier, flame photometry has been adopted for the simultaneous
detection of Na+, Li+, and K+ in laboratory samples [60]. Even though the data accuracy of
existing standard instruments based on the aforesaid methodologies, several factors like high
cost, immobility, and associated recurring cost of reagents and appropriate electrode [61,62],
limit their accessibility for rapid detection of the electrolyte and availability for patient care
at any place any time, which is an essential need of today. Biological fluids like urine, serum,
plasma, and whole blood are used for the determination of electrolyte concentration. The

8
selection of the sample type relies upon the design of the measuring instrument itself.
Besides the conventional methodologies, spark emission spectroscopy is another method to
sense the electrolyte concentration as reported in the literature [63]. A strategy for the
detection of the essential electrolytes like Na+, K+, and Li+ at the same time with no rigorous
sample preparation has also been demonstrated in the project.

The invention of the ISE by Frant and Ross [64], was a revolution and the technique has
been widely accepted as one of the conventional methods [65,66] for the detection of
different ion [67] concentration that present in groundwater. In this context, it has been well
reported in several studies that the fluoride forms an Al(III)-fluoride complex [68,69] in the
presence of Al(III). Thus, the determination of the fluoride ions, in the presence of Al(III)
ion in drinking water severely compromises the detection efficiency in the ISE technique.
The use of potassium alum (Al(III)-based compound) to the water has been widely accepted
for the drinking water treatment and the possibility of Al(III) ion leaching is obvious. Some
other fluoride detection strategies based on NIR method [70], chemo-sensor [71],
colorimetric methods [72,73], molecular recognition linked transduction mechanism and
organic receptors having amide, urea, thiourea, guanidinium or pyrrole functionalities
capable of hydrogen bonding with the fluoride anion in their binding sites are in use
[69,74,75]. However, interference from other anions and cations essentially complicates the
detection of fluoride in drinking water, demands the improvement of developing an
economic detection method with comparable sensitivity. In this context, a nano-sensor has
been fabricated in the project and using the sensor an affordable device has been developed
for determining the fluoride concentration in the drinking water.

In contemporary time, different spectrophotometry techniques like AAS [76], ICP-OES

[77,78], electrochemistry [78,79] are the well-accepted mechanism for the detection of iron
concentration in a biological sample. But, high setup cost and complexity in the sample
preparation limited their use in point-of-care settings. In addition to the existing expensive
instrument, some other inexpensive and portable instruments like photometer and color
comparator are used for the same purpose. Change in color of the reagents in the presence
of the ion concentration of the homogeneous sample solution plays a key role. Different
chemicals thus are essential requirements to run such instruments. In reality, the living

9
samples are heterogeneous in nature. Thus photometer and color comparator both are not a
good choice for the iron concentration determination in the biological samples. In the
project, a nanosensor has been fabricated on Triton X-100, a neutral micelle duly sensitized
by porphyrin [80]. The nanosensor has been devised to determine the iron concentration in
water, in protein and also in different parts of the growing plant.

1.4. Summary of the Work Done

1.4.1. Optical Methodology for the Minimally Invasive Diagnosis of Anemia.

1.4.1.1. Digital Camera Based Spectrometry for the Development of Point-of-Care

Anemia Detection on Ultra-low Volume Whole Blood Sample [81]: Here, we have
described a method for initial screening of hemoglobin level at the place of patient care. In
this study, an affordable method has been developed for rapid and reliable screening of the
hemoglobin level from whole blood. A simple scattering-based method has also been
introduced in this study to predict the average structure of red blood cells. A digital camera
has been used as a detector in the developed device for the screening of hemoglobin
concentration from 10µL whole blood in a low resource setting. Hemoglobins of a wide
range of concentrations have been collected from a significant number (300) of human
subjects for calibration and validation with respect to a standard automated hematology
analyzer. The developed device comes with the fabricated microcuvette that needs only
10µL unprocessed whole blood sample for the detection of hemoglobin level.

1.4.2. Development of Optical Spectroscopy-based Methodology for the Non-contact

Non-invasive Diagnostic Strategy for Hyperbilirubinemia.

1.4.2.1. A Novel Whole Spectrum-based Non-invasive Screening Device for Neonatal

Hyperbilirubinemia [82]: A new non-contact and non-invasive methodology has been
studied for the determination of bilirubin in neonatal subjects. Careful screening of bilirubin
levels in a newborn is mandatory as per the American Academy of Pediatrics (2004), to
reduce incidents of kernicterus and acute bilirubin encephalopathy. Although an invasive
capillary collection of blood and the subsequent biochemical test is considered a
conventional for jaundice detection in neonates, transcutaneous bilirubin measurement using
various non-invasive instruments are also used sporadically across the globe. The major aim

10
of this study is to develop a non-invasive spectrometry-based technique for measurement of
neonatal bilirubin level as an alternative of total serum bilirubin (TSB) test without
limitations of other available bilirubin meters. The instrument comprises a light source and
a spectroscopic detector. A light beam from source incident on the neonatal nail plate
through optical fibers. The retroreflected light is acquired using the detector. An
indigenously developed software is used to acquire and analyze the optical signal and to
calculate the bilirubin value. The instrument was calibrated and validated about TSB on 1033
subjects. The result (r = 0.95 and P < 0.001) indicates a strong correlation between the
bilirubin value obtained from our method and TSB. Time variant analysis of the subjects
undergoing phototherapy provided a good correlation (r = 0.98). The repeatability test result
shows the mean coefficient of variation is less than 5.0%. The indigenously developed non-
invasive technique successfully detects the bilirubin level in a newborn under various
physiological conditions with high accuracy and precision.

1.4.2.2. Large scale Validation of a new Non-Invasive and Non-Contact

Bilirubinometer in Neonates with risk factors [83]: The performance of a developed non-
invasive and non-contact transcutaneous bilirubin measurement device named AJO-Neo has
been established as an alternate to the conventional TSB test in different gestational ages
suffering from hyperbilirubinemia associated with risk factors, and/or undergoing
phototherapy. The safety and efficacy of the device were assessed in 1968 neonates with
gestational ages ranging from 28 to 41 weeks and suffering from incidences of
hyperbilirubinemia. The evaluated linear regression coefficient, r=0.79 indicates a good
correlation between AJO-Neo and the conventional bilirubin test. The small bias (0.27
mg/dL) and limits of agreements (-3.44 to 3.99 mg/dL) are found to be in the range of clinical
acceptance. The device was also exhibited precisely in the measurement of bilirubin levels
in all subgroups of the study. The receiver operator curve (ROC), which takes account of
both sensitivity and specificity of a device, has been exposed to the high efficacy of the
device (area under the curve, AUC=0.83) in the detection of bilirubin. Besides, monitoring
the bilirubin level during phototherapy, the device indicated promising results with good
agreement with TSB. Hence, AJO-Neo is found reliable in measuring bilirubin levels in
preterm, and term neonates irrespective of gestational or postnatal age, sex, risk factors,
feeding behavior, or skin color.

11
1.4.3. Development of Optical Spectroscopy-based Methodology for Minimally-
invasive Diagnosis Essential Salt in Human Body.

1.4.3.1. NaLiK, a Self-developed Device for Rapid, Reliable and Simultaneous

Assessment of Sodium, Lithium, and Potassium for Management of Fluid Balance and
Bipolar Disorder in Human Subjects [84]: Here a minimally invasive optical approach for
detection of electrolytes like sodium, potassium, magnesium, calcium, etc have been studied.
Sodium and potassium are two essential electrolytes that have been recognized for
maintaining fluid balance and homeostasis in the human body. In understanding the
physiological disorder due to diseases, sodium and potassium have been found useful
biomarkers. The major objective of this study has been aimed to develop a strategy for rapid,
reliable, and simultaneous assessment of three electrolytes, sodium (Na+), lithium (Li+), and
potassium (K+) in the conventional sample taken from human subjects. The fundamental of
the device has been designed employing the law of optical emission spectroscopy (OES).
The photons have been acquired during the ionic transition and analyzed for the rapid and
simultaneous detection of the specific electrolytes. An indigenously developed software has
been used to control every instrumentation module and carries out all mathematical analysis
to produce the result. Satisfactory results have been obtained from the linear regression
analysis for the three electrolytes (adjusted R2 values 0.954, 0.932, and 0.984 for Na+, Li+,
and K+ respectively) with respect to the standard ISE method while the tests have been
conducted on the same sample. The detection system has been designed on a sample volume
of about 100L serum and provides a good correlation with a commercially available ion-
selective electrode device. The indigenously developed low-cost device has a feature to offer
a rapid and simultaneous assessment of the essential electrolytes. To our understanding, the
device would be ideal for the diagnosis of Chronic Kidney Diseases (CKD) and Bipolar
Disorder in economically challenging countries.

1.4.4. Development of a Spectroscopy-based Sensor for Detection of Fluoride Ion in

Drinking Water.

1.4.4.1. Selective and Fast Responsive Sensitized Micelle for Detection of Fluoride Level
in Drinking Water [85]: In the present work we have developed a novel FeFlu sensor and
device for fluoride detection. Consumption of excess fluorides through drinking water and

12
its adverse effects on human health is a global concern. Given that people from at least 25
countries are suffering from fluorosis, global management of its issue is of immense
importance. Here we have developed a sensor called FeFlu, based on an anionic micelle
(SDS)-hematoporphyrin (Hp) complex, sensitized by Fe(III) chloride, for the detection of
fluoride in water. The SDS-Hp complex with iron is almost non-fluorescent, but, the
prototype (FeFlu device) based on it displays a strong turn-on fluorescence upon recognition
of fluoride in water. Besides, the detection of fluorides using FeFlu is found to suffer
insignificant interference from environmentally relevant anions and cations. In the present
study, we have also fabricated a fluorescence-based prototype FeFlu device based on a
photophysical investigation of several sensitized micelle-Hp complexes and validated the
results with fluoride ion-selective electrode, which is widely utilized for the detection of
fluorides in drinking water easily and affordably.

1.4.5. Development of a Low-Cost Digital Camera based Iron Sensor for

Environmental and Biomedical Application.

1.4.5.1. Development of A Nano-sensor (FeNSOR) Based Device for Estimation of Iron

ions in Biological and Environmental Samples [86]: Here we have developed a sensor
and a device to detect iron from biological and environmental samples. Iron is one important
key nutrient, essential for the development of biological systems. Inaccurate presence of iron
in groundwater and biological systems becomes the source of different health problems. In
the study, a nano-sensor called FeNSOR has been fabricated from porphyrin sensitized
neutral micelle (TX-100). Based on the developed nano-sensor, a device has been designed
for the detection of iron ion concentration in low resource settings. The device functions on
the fundamental law of fluorescence spectrometry and therefore the placement of the light
source and the detector has been kept at 90° geometry. A Pi-camera has been used in the
device as a detector and a 365nm LED for the light source. The efficiency of the fabricated
nano-sensor and the functionality of the developed device found excellent in the assessment
of iron concentration both in the environment and biological sources in the micro-molar
range. The sensor has been found negligibly interfered with other ions.

13
1.5. Plan of the Thesis

The plan of the thesis is as follows:

Chapter 1: The chapter briefly introduces different types of optical methodologies used in
various biomedical and environmental applications. The summary and future scope of the
work done has also been included in this chapter.

Chapter 2: The chapter provides an overview of spectroscopic strategies, both the

dynamical and steady-state, the structural aspects of biologically important systems and
probes used in the research.

Chapter 3: Details of instrumentation, data analysis, and experimental procedures have

been discussed in this chapter.

Chapter 4: The chapter demonstrates the working of a digital camera as the optical detector
for the development of an anemia screening device with excessive-low volume untreated
blood samples for point-of-care use.

Chapter 5: The chapter gives out the idea about a non-contact transcutaneous screening
device for neonatal hyperbilirubinemia using the absorption spectrum of blood. The
performance of AJO-Neo in an Asian population of preterm, near-term, and term neonates
under prescribed conditions with risk factors and phototherapy are also validated with the
reference of the conventional biochemical method (TSB).

Chapter 6: The chapter deals with the application of optical emission spectroscopy as a tool
for the rapid detection of essential electrolytes like sodium, lithium, and potassium at the
same time from very low volume blood serum of human subjects.

Chapter 7: The chapter offers the synthesis and characterization procedures of an iron-based
fluoride sensor (FeFlu) and the development of a digital camera-based device to monitor
fluoride ions in drinking water.

Chapter 8: The chapter deals with the synthesis of a micelle based nanosensor (FeNSOR)
and the implementation of the nanosensor to form a device. The sensor efficacy for the

14
estimation of iron ions from groundwater, human serum albumin (HSA), and the plant
environment have also been discussed.

15
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