Intermediate results obtained from WQI method.
\r\n\tThus, we call for research and review papers on the chemistry and physics of dyes, pigments, and their intermediates, including chemical constituents, spectroscopic aspects, surface, solution, crystal formation, photochemical, and ecological or biological properties. The book will be of interest to a wide variety of researchers worldwide whose work involves various fields of activity such as dyes and pigment synthesis, imaging, sensor, energy, medicine, polymers, food product, toxicological properties, etc.
",isbn:"978-1-83768-114-3",printIsbn:"978-1-83768-113-6",pdfIsbn:"978-1-83768-115-0",doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!0,isSalesforceBook:!1,isNomenclature:!1,hash:"fcd069956c2e931195925b19a74ce9a3",bookSignature:"Dr. Brajesh Kumar",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/12081.jpg",keywords:"Heterocycles Pigments, Azo, Nitro, Indigo, Alizarin, Chromophores, Chromophores, Photochemical, Sulphonation, Diazotisation, UV-Vis Spectroscopy, Metal-Ligand",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:null,numberOfDimensionsCitations:null,numberOfTotalCitations:null,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"May 19th 2022",dateEndSecondStepPublish:"June 16th 2022",dateEndThirdStepPublish:"August 15th 2022",dateEndFourthStepPublish:"November 3rd 2022",dateEndFifthStepPublish:"January 2nd 2023",dateConfirmationOfParticipation:null,remainingDaysToSecondStep:"9 days",secondStepPassed:!0,areRegistrationsClosed:!1,currentStepOfPublishingProcess:3,editedByType:null,kuFlag:!1,biosketch:"Dr. Brajesh Kumar has worked as a faculty member in various universities in India, Ecuador, and South Korea. He has published numerous SCI/SCIE/Scopus research articles and is an active reviewer of more than 50 Journals. Dr. Kumar is a member of the American Chemical Society, the Indian Society of Chemists and Biologists, and the Indian Science Congress Association and holder of two registered patents. He is included in the top 2% of the scientist list prepared by experts at Stanford University,",coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"176093",title:"Dr.",name:"Brajesh",middleName:null,surname:"Kumar",slug:"brajesh-kumar",fullName:"Brajesh Kumar",profilePictureURL:"https://mts.intechopen.com/storage/users/176093/images/system/176093.JPG",biography:"Dr. Brajesh Kumar is currently working as an Assistant Professor and Head in the Post Graduate Department of Chemistry, TATA College, Chaibasa, India. He received a Ph.D. in Chemistry from the University of Delhi, India. 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Water can be classified according to its source into [1, 2]:
Surface water (water located at the surface of the soil) - Surface water sources are represented by running waters, water seas, oceans, rivers, lakes, icebergs;
Groundwater water below the surface in the saturated area and in direct contact with soil or subsoil. Groundwater sources are represented by groundwater aquifers, deep aquifers, springs; and,
Atmospheric water.
According to the field of use, the water is classified as:
Drinking water: for domestic consumption and for agriculture; and,
Industrial water: auxiliary in manufacturing processes, raw material for various industries, power generator, coolant agent, heating agent, etc.
Wastewater is water that has changed its original properties through use, in other words has been contaminated by human beings [2].
Water quality is represented by a set of physical, chemical, biological and bacteriological characteristics. These characteristics are also called parameters or indicators. Physical, chemical, biological and bacteriological parameters allow water to be classified in some categories, leading to its utilization for a specific use.
Water quality requirements depend on the purposes for which the water will be used. Thus, drinking water must not contain chemicals or micro-organisms which can affect the human health. Water used in agriculture must not contain large amounts of sodium ions, high concentrations of nitrates or high concentrations of other contaminants. Requirements for water use in industry are less rigorous than drinking water [3].
Water quality also depends on the type of water source and changes with geological, meteorological and land use conditions. The World Health Organization (WHO) has established regulations and standards for water safety in support of public health [3]. The European Union has, also, established a legal framework for water protection [4]. Water quality criteria in all countries have been established in accordance with the WHO guidelines [3]. In European countries, the framework directives of the European Union are closely followed [4].
The EU Framework Directive requires that operational monitoring should be specific and based on monitoring relevant biological, hydro-morphological and physic-chemical parameters. These world environmental monitoring systems provide for water quality measurements in three categories of parameters:
Basic parameters: Temperature, pH, Conductivity, Dissolved Oxygen (DO), Total Dissolved Solids (TDS), Coli bacilli, Biochemical Oxygen Consumption (CBO5), Turbidity, Nitrites and Phosphates concentration;
Indicators of persistent pollution: Cadmium, Mercury, Organo-halogen compounds and Mineral oils; and,
Optional parameters: Total Organic Carbon (TOC), Anionic detergents, Heavy metals, Arsenic, Boron, Sodium, Cyanides, Total oils, Streptococci [3].
Water quality monitoring indicators established by the European Union rules are grouped according to their common characteristics as it follows [4, 5]:
Indicators that give information about oxygen condition: Dissolved Oxygen, Biochemical Oxygen Consumption (CBO5), Chemical Oxygen Consumption with chromium, Chemical Oxygen Consumption with manganese;
Indicators that give information about the presence of nutrients that contribute to eutrophication: Ammonium (NH4+), Nitrates (NO2−), Nitrites (NO3−), Total nitrogen (N), Orthophosphates (PO43−), Total phosphorus (P);
Salinity indicators: Chlorides (Cl−), Sulphates (SO42−), Ca2+, Mg2+, Na+;
Heavy metals (e.g. Fe, Cu, Cd, Pb, Hg); and,
Other relevant indicators: Phenols, Anionic surfactants, Absorbed organic halides [4, 5].
To understand the overall health of an ecosystem and the condition of water, a number of water quality parameters or indicators must be analyzed and monitored. In 1998, Sene and Farquharson [6] stated that monitoring of the surface water quality is necessary to assess spatial and temporal regional variations. The process of monitoring the quality of ambient water has led to the development of water standards and the periodic assessments of the environment.
The monitoring program and the parameters to be measured for the study of water quality should be chosen specifically for each locality and each type of water. Although many parameters of water are important for human health or the health of an ecosystem, the analysis of all parameters is not feasible. The standards recommend the analysis of specific parameters for both drinking water and non-drinking water [6, 7].
Chemical and physical parameters are important in the rapid determination of water quality while biological parameters provide a detailed and complex analysis of the environment [8].
Oxygen is necessary for aquatic life. A series of aerobic chemical processes take place through dissolved oxygen: the oxidation processes of organic matter, oxidation of mineral substances, and bio-chemical decomposition of the dead bodies in water [9]. With the decrease of oxygen, the self-purification capacity of natural water is reduced, favoring the persistence of pollution with its undesirable consequences. Other indicators of oxygen regime are Biochemical Oxygen Consumption (CBO5) and Chemical Oxygen Consumption (CCO).
The phosphate content in natural water is relatively low. High amounts of phosphorus in water can come from excessive use of nitrogen and phosphorus fertilizers. Higher concentrations of phosphorus in surface water can result in eutrophication.
Heavy metals are those metals that have a high density (i.e. 5 g/cm3) [10]. In low concentrations, heavy metal ions are essential for the development of metabolic processes in plants and animals. These metals (e.g., cadmium, chromium, cobalt, lead, nickel, mercury, selenium) can come from natural or anthropogenic processes. If certain concentrations are exceeded, then they become toxic substances for the living organisms.
Water quality and its changes due to various forms of pollution may influence the composition of aquatic biocenoses. Biological analysis consists of an inventory of phytoplankton, zooplankton, benthic organisms or periphyton from water samples.
The microbial flora found in the water can be classified into two categories: water-specific microbial flora and microbial impurity flora. Water-specific microbial flora consists of microorganisms that commonly inhabit water and soil: cocci bacilli, different fungi and bacterial species which play a role in the natural degradation processes of organic substances. Microbial impurity flora consists of species of microorganisms of human or animal origin. This category can include pathogenic saprophytes. These microbes are generally accompanied by high concentrations of organic matter which provide their nutritional support [11].
In bacteriological analysis of water, the total number of germs and the determination of the bacillus coli have been adopted as bacteriological indicators.
Water quality is determined by the biological, chemical and physical parameters of the water. Most often, it is not enough to measure these water quality indicators. In order to draw some solid conclusions, it is necessary to apply adequate statistical method to the measurements. These statistical methods can provide useful information that can lead to actionable advice regarding water management. There are a large number of statistical methods for examining water quality.
The main differences between these methods are the statistical techniques used and the significance of the values determined for each parameter. Statistical indices developed using water quality parameters can be linear, non-linear, segmented linear or segmented non-linear [12]. In order to have a global vision of the changes of the water quality in space and in time, various indices have been developed [13].
The water quality index (WQI) is represented by a number that expresses the general water quality in a particular location, over time, based on several water quality parameters. The aim of this index is to transform a large number of complex water quality measurements into information that is easy for water managers and the public to understand and to use. Are a multitude of methods for calculating water quality indices (WQI). In the following, we present the weighted average method. This method was proposed by Horton in 1965 and developed by Brown et al. in the year 1970 [14].
For the calculation of the WQI, the following expression was used [15, 16, 17, 18, 19]:
where:
where:
Water quality is Excellent if the WQI index score is between 0 and 25; Good for values of 26–50; Poor for WQI = 51–75; and, Very Poor for values between 76 and 100. If the value of the WQI index exceeds the value of 100, then the water is unsuitable for drinking and cannot be transformed into drinking water by any process [19, 21].
To study the relationship between two parameters of water samples, several correlation coefficients can be used. The statistics used most often are Pearson and Spearman coefficients. Linear correlation can be determined using the Pearson correlation coefficient while non-linear correlation can be determined using the Spearman coefficient. The Pearson correlation coefficient is a statistical technique that measures and describes the degree of linear association between two normally distributed continuous quantitative variables [21]. Let
where
Examples of the geometric significance of the Pearson coefficient.
The minimum value of the Pearson coefficient (r = 0) is not an indicator of independence of the two characteristics (variables), but only of their non-correlation. The coefficient of determination (r2) is the square of the Pearson coefficient. The coefficient of determination indicates the percentage of the total variation of the dependent variable (y) which is explained by the independent variable (x).
Spearman method is a non-parametric method used when the relationship between two variables is not linear (monotonic correlation) [23, 24, 25]. The Spearman coefficient addresses some limitations of the Pearson coefficient. It is denoted either with ρ or with rS and represents an alternative to the Pearson coefficient. To calculate the coefficient, the data must have an order or rank. Coefficient can be calculated using the formula:
where:
where:
Eq. (6) is usually used when all n ranks are distinct integers or do not have tied ranks. When there are tied ranks, Eq. (6) is replaced by the following form:
where
Spearman coefficient values are in the range [−1, 1]. The interpretation of these values is similar with that of the Pearson coefficient [21].
For a correct interpretation, the correlation coefficient must be accompanied by a significance test. The correlation coefficient has statistical significance if the value level of confidence factor p < 0.05. This significance coefficient p means the probability of making erroneous statements. If p < 0.05, we could reject the null hypothesis H0 and the computed results has certain statistical significance [24]. If the p result of the test is less than the significance threshold α (α = 0.05), hypothesis H1 is accepted: there is monotonic correlation. If p is greater than 0.05, then the H0 hypothesis is valid, which considers that there is no monotonic correlation [25].
In this section, we provide an example of how to apply these methods in order to achieve a rapid assessment of water quality. The data set chosen for statistical analysis comes from our previous work [15, 16] and consists of 13 water quality parameters that were determined from samples taken from the Danube River. Sampling points were located along the river in the neighborhood of Galati. Galati is a Danube port city in the south-eastern part of Romania. Water samples were collected from November 2016 to December 2017. We will use data from 3 locations coded with D1, D4 and D7. All locations are along the Danube’s left bank, D1 being located upstream and D7 downstream (Figure 2). The measured parameters were: potassium and calcium ions, nitrites, nitrates, total nitrogen, ammonium, chlorides, total phosphorus, sulphates, cadmium, chrome, copper, lead, iron, zinc, density, dissolved oxygen, chemical oxygen demand (CCO-Cr), biochemical oxygen demand (CBO5), electrical conductivity, the density of the conductivity, resistivity, pH, salinity, total dissolved solids [15].
Sampling points [
From our previous work [15, 16], the scatter plot diagrams and the box plot diagrams of the parameters indicated that quality class thresholds were exceeded during certain time periods. Correlations between the measured parameters could not provide a clear conclusion on the water quality condition.
For these reasons to provide clear information on the water quality condition, we calculated the Water Quality Index (WQI).
The Water Quality Index evaluation consisted of several stages. It is important to scale and weight the values of the monitored parameters according to the allowed limit values.
The water quality standards,
Table 1 presents the intermediate results obtained from the application of the Water Quality Index method. The Unit Weights (
Parameter | Standard (interval) | K from Eq. (4) | Wn from Eq. (1) | Vid |
---|---|---|---|---|
pH | 6.5–8.5 | 0.01585 | 0.001864787 | 7 |
CCO-Cr (mg O2/l) | 50 | 0.000317014 | 0 | |
CBO5 (mg O2/l) | 7 | 0.002264384 | 0 | |
TDS (ppm) | 200 | 7.92534E-05 | 0 | |
Dissolved oxygen (mg/l) | 5 | 0.003170138 | 14.6 | |
Chlorides (mg/l) | 250 | 6.34028E-05 | 0 | |
Nitrites (mg/l) | 0.06 | 0.264178133 | 0 | |
Nitrates (mg/l) | 5.6 | 0.00283048 | 0 | |
Ammonium (mg N/l) | 1.2 | 0.013208907 | 0 | |
Total nitrogen (mg/l) | 12 | 0.001320891 | 0 | |
Sulphates(mg/l) | 250 | 6.34028E-05 | 0 | |
Lead(mg/l) | 0.025 | 0.63402752 | 0 | |
Cadmium(mg/l) | 2 | 0.007925344 | 0 | |
Total Phosphorus(mg/l) | 0.75 | 0.021134251 | 0 | |
Iron(mg/l) | 1 | 0.015850688 | 0 | |
Zinc(mg/l) | 0.5 | 0.031701376 | 0 |
Intermediate results obtained from WQI method.
The obtained values for the water quality index corresponding to the three locations are presented in Figure 3.
Monthly values of WQI in the three locations during the monitoring.
According to the diagram from Figure 3, during the time interval November 2016–June 2017, the WQI values for the Danube River water were found in the range of 100 to 2310. Between July and December 2017 the values decreased in the range 0–25. In the first-time interval (November 2016 to June 2017), the water quality index shows that the water was not suitable for consumption and cannot be transformed into drinking water by any process. However, by the end of the monitoring time interval (December 2017) the water quality was good or excellent.
According to our previous work [15, 16], during November 2016–June 2017, the following indicators had exceeded the limit permitted by Romanian law: all metals, Chlorides, Nitrates, Nitrites, Ammonium, Total Phosphorus, Sulphates, Solvent Extractable Substances and Anionic Surface Agents, Chemical Oxygen Consumption with chromium (CCOCr), Biochemical Oxygen Consumption (CBO5). The high values obtained for these indicators were determined by the wastewater discharges into the Danube water. The high values of these indicators determined high values of WQI. At the end of the monitoring time interval the values of the studied indicators have improved. This improvement is found in the low values of WQI. The substantial improvement in water quality that occurred is due to the actions taken by the organizations responsible for environmental protection.
Figure 4 shows the boxplot diagram representing the main values of WQI in the 3 chosen locations. The average values of WQI are influenced by the extreme values. According to the 3rd quartile (Q3) and the median of the upper half of the data set, 75% of the values in the data set lie below Q3. The high average and median values, the values of the 3rd quartile frame depict waters as having severe pollution.
Boxplot diagram of WQI in the locations D1, D3, D7.
The value of the third quartile indicates that 75% of the determined values of WQI fall into the category of highly polluted waters. Based on Figure 3, only 25% (the 1st quartile - Q1) of the values of the WQI lie below low values that would classify the studied water into the category of unpolluted waters. The information obtained from the WQI calculation was particularly useful in order to analyze how the overall water quality has evolved over time.
An easy method to identify possible sources of pollution is to calculate the correlations between the measured parameters. Using a Pearson Correlation Matrix [15] there was a strong positive linear correlation between TDS and Salinity (r = 0.9394) and TDS and Electrical Conductivity EC (r = 0.9174). Significant correlations also existed between the nitrites concentration and pH and between the nitrates concentration and pH there was a moderate negative corelation (r = −0.65 and − 0.68 respectively).
To identify possible sources of pollution, the Pearson correlation matrix was computed between WQI and a series of measured parameters (Table 2).
WQI | ||
---|---|---|
Parameter | r | r2 |
pH | 0.614 | 0.376996 |
CCO-Cr | 0.28 | 0.0833 |
CBO5 | −0.4297 | 0.1846 |
TDS | 0.3092 | 0.0956 |
DO | −0.51102 | 0.2611 |
Chlorides | 0.944033 | 0.891198 |
Nitrites | 0.881528 | 0.777091 |
Nitrates | 0.930999 | 0.86676 |
Ammonium | 0.847447 | 0.718166 |
Total nitrogen | 0.624669 | 0.390211 |
Sulphates | 0.855873 | 0.732519 |
Lead | 0.763807 | 0.583401 |
Cadmium | 0.946214 | 0.895321 |
Total Phosphorus | −0.03909 | 0.001528 |
Iron | 0.934618 | 0.873511 |
Zinc | 0.934375 | 0.873057 |
Pearson correlation coefficients.
In the absence of dedicated statistical software, the correlation coefficients can also be determined using free tabular software tool. We could exemplify quite easily this technique, for the Pearson coefficient between WQI and CCOCr, for the first location D1 (Table 3).
( | ( | ( | Sx | Sy | r | ||
---|---|---|---|---|---|---|---|
406.854 | −65.837 | 165530.052 | 4334.5 | −26786.0468 | 2623.25 | 186.16 | 0.28858 |
288.854 | −65.837 | 83436.544 | 4334.5 | −19017.2808 | |||
−166.696 | −58.457 | 27787.608 | 3417.212 | 9744.548072 | |||
375.374 | 34.163 | 140905.524 | 1167.116 | 12823.90196 | |||
1656.854 | 52.663 | 2745164.668 | 2773.4 | 87254.9022 | |||
276.854 | 52.163 | 76648.052 | 2720.987 | 14441.5352 | |||
1055.854 | 56.163 | 1114827.344 | 3154.291 | 59299.9282 | |||
−648.946 | −65.837 | 421131.111 | 4334.5 | 42724.6578 | |||
−648.036 | −65.837 | 419950.857 | 4334.5 | 42664.74613 | |||
−649.106 | 34.163 | 421338.799 | 1167.116 | −22175.40828 | |||
−649.146 | 34.163 | 421390.729 | 1167.116 | −22176.7748 | |||
−649.596 | 24.163 | 421975.163 | 583.854 | −15696.18815 | |||
−649.116 | 34.163 | 421351.781 | 1167.116 | −22175.74991 | |||
Calculation of the Pearson correlation coefficient between WQI (x) components and chemical oxygen consumption with chromium-CCOCr (y).
Table 3 shows the values of Pearson Correlation Coefficient (r) and coefficient of determination (r2) for the water quality data set. The major influence of several parameters on the high values of WQI is due to strong positive correlation values. Therefore, excessive pollution was likely due to the presence of high concentrations of chlorides, nitrates, nitrites, ammonium, sulphates, lead, cadmium, iron, zinc.
The values of coefficient of determination (r2) indicate that 89% of the variance of WQI is explained by the chlorides and cadmium concentrations while 87% is due to effect of iron and zinc. Nitrates concentration in the Danube River water explains 86% variation of the WQI. The high levels of covariance explained by the three groupings suggest significant co-linearity among the nutrient groups.
The strength and direction of monotonic association between water quality variables can be highlighted by the Spearman correlation. Table 4 shows the Spearman coefficients between the WQI and the water quality indicators that were measured.
WQI | ||
---|---|---|
Parameter | rS | p (2-tailed) |
pH | −0.7459 | 0.00342 |
CCO-Cr | 0.24304 | 0.42366 |
CBO5 | −0.3499 | 0.24126 |
TDS | −0.1658 | 0.58839. |
DO | −0.674 | 0.01153 |
Chlorides | 0.95461 | 0 |
Nitrites | 0.93095 | 0 |
Nitrates | 0.92562 | 0 |
Ammonium | 0.86046, | 0.00016 |
Total nitrogen | 0.8138 | 0.00071 |
Sulphates | 0.83252 | 0.00041 |
Lead | 0.75674 | 0.00275 |
Cadmium | 0.85436 | 0.0002 |
Total Phosphorus | 0.59311 | 0.03263 |
Iron | 0.82855 | 0.00047 |
Zinc | 0.8856 | 0 |
Spearman rank correlation coefficients.
Table 4 shows the high values obtained for WQI are associated with the high values obtained for chlorides, nitrates, nitrites, ammonium, total nitrogen, sulphates, lead, cadmium, iron, zinc. The association between these variables would be considered statistically significant.
Table 5 exemplifies such a calculation for the correlation coefficient between WQI and CCO-Cr.
x | y | rx | ry | rs | ||||||
---|---|---|---|---|---|---|---|---|---|---|
1060 | 10 | 11 | 2.5 | 4 | −4.5 | −18 | 16 | 20.25 | 0.24 | |
942 | 10 | 9 | 2.5 | 2 | −4.5 | −9 | 4 | 20.25 | ||
486.45 | 17.38 | 7 | 5 | 0 | −2 | 0 | 0 | 4 | ||
1028.5 | 110 | 10 | 8.5 | 3 | 1.5 | 4.5 | 9 | 2.25 | ||
2310 | 128.5 | 13 | 12 | 6 | 5 | 30 | 36 | 25 | ||
930 | 128 | 8 | 11 | 1 | 4 | 4 | 1 | 16 | ||
1709 | 132 | 12 | 13 | 5 | 6 | 30 | 25 | 36 | ||
4.2 | 10 | 5 | 2.5 | −2 | −4.5 | 9 | 4 | 20.25 | ||
5.11 | 10 | 6 | 2.5 | −1 | −4.5 | 4.5 | 1 | 20.25 | ||
4.04 | 110 | 4 | 8.5 | −3 | 1.5 | −4.5 | 9 | 2.25 | ||
4 | 110 | 2 | 8.5 | −5 | 1.5 | −7.5 | 25 | 2.25 | ||
3.55 | 100 | 1 | 6 | −6 | −1 | 6 | 36 | 1 | ||
4.03 | 110 | 3 | 8.5 | −4 | 1.5 | −6 | 16 | 2.25 | ||
Intermediate results for the Spearman correlation.
For chlorides, nitrates, nitrites, ammonium, sulphates, lead, cadmium, iron, zinc, total nitrogen, the values of p coefficient are less than 0.001 (i.e., highly significant with confidence greater than 99.99%). For pH and DO, p < 0.01 means the statistical links are significant and the confidence is 99%.
Once the correlations between pollutants and WQI are identified, the sources of pollution can be established or the related process.
This chapter highlighted the importance of using statistical methods to display the water quality condition, using WQI evaluation and Pearson and Spearman correlations.
In order to exemplify the statistical methods, we have used a series of data from our previous work, consisting of 13 parameters measured for water samples taken from the Danube River, from Galati City area, Romania. Statistical correlations were made between quality parameters and Water Quality Index; thus, it was possible to identify which are the pollutants that determined an advanced degree of water pollution. The excessive pollution which occurred during the time interval November 2016–June 2017 is due to the presence of high concentrations of chlorides, nitrates, nitrites, ammonium, sulphates, lead, cadmium, iron and zinc. In recent times there are many statistical software for water quality analysis. If we do not have, for various reasons such programs, the statistical approach can be done classically. Water Quality Index (WQI) provides information on the overall quality of the water, while the correlation coefficients may indicate the parameters that influenced the changes in water quality.
This study was funded by the main author’s personal resources.
As a promising interdisciplinary field, nanotechnology acts as a bridge for various disciplines, such as material sciences, engineering, physics and chemistry, and is dedicated to the production of different materials in the nanometer scale (<100 nm), with assorted physical, chemical and mechanical properties. Although nanoscience and nanotechnology are new research interests, the application of nanomaterials for humankind was well-known since ancient times. Among the periodic elements, carbon has the intense ability of catenation and great tendency to form various hybrid orbitals (e.g., sp, sp2, and sp3) which results in the formation of various smart compounds having different physical and chemical properties according to their structure [1, 2]. Due to the properties mentioned above, carbon have tendency of forming different allotropes of different dimensions, like quantum dots (0D), carbon nanotubes (1D), fullerenes (0D), graphene (2D), graphite (3D), among which, graphene got lot of attention in the past decade. Graphene has hexagonally packed honeycomb like geometry in which a unit layer of carbon atoms are arranged in two-dimensional (2D) lattice [3, 4, 5, 6]. The hybrid orbital of carbon-carbon atoms are in sp2 hybridized form, in which the in-plane σ (C▬C) bonds are much stronger than the out-of-plane π (C▬C) bonds, which is highly accountable for the delocalized array of electrons and come up with the weak polar interaction between graphitic layers of the graphene sheets as well as with graphene and other molecules.
In scheming a potent drug delivery carrier, besides its physicochemical constancy in the biological surroundings, reactivity and toxic issues, diffusivity, immunogenicity, interactions with biological systems, drug loading and release characteristics, blood circulation half-life, drug transportation ability of the biological medium to aim the cells within tissues, etc. are the significant issues (Figure 1). Due to the electrostatic interaction and presence of different alkali and alkaline earth metal ions
Properties and applications of graphene oxide.
Number of research groups have been investigating the surface modification of GO with different kinds of biocompatible polymers as a nanodrug carrier for development of targeted drug delivery systems. The polymers are selectively preferred according to their functional groups, bioavailability and compatibility in the cell medium [10]. Therefore responding to specific stimulus, surface fabrication of GO with various polymers for this particular drug delivery application is limited [11]. For well-organized diagnosis, expressive intracellular drug release is elected over the contemporary arrival of the drug in the system.
With the innovation of the smart material graphene, the curiosity of researchers remarkably moved toward graphene and its oxygenated derivatives from the previously invented other nanomaterials of carbon family, and various scientists are working to organize the surface modification of graphene through the spacious understanding of different functionalization methods [12, 13, 14]. From the chemical point of view, production of graphene oxide that possesses many reactive oxygen-containing groups is appropriate for advance modification and relative aspects of GO sheets, or we can say that the reactivity of GO is highly desirable.
Beside this, graphene is sparingly soluble in the water, polar solvents and in the cell environment due to aromatic character [15]. Graphene oxide on the other hand has water contact angle of 30.7° [16] and proficient of composing weak hydrogen bonds and metal incorporated complex ion due to the polar oxygenated groups present on the basal plane and negatively charged carboxylic groups present on the edge site [17, 18, 19, 20]. The distinctive arrangement of graphene oxide and its strong carbon-carbon covalent bonding provides outstanding thermal and electrical conductivity with very low thermal expansion quotient. These properties of graphene are also significantly affected by alteration such as edge scattering defect [21] and isotopic doping [22] due to diffusion or localization of phonons at the defect sites. Light absorption and optical imaging are highly dependent on the total number of layers present in GO sheets, as they increase accordingly with the number of layers present in GO [23]. Optoelectronic devices based on GO derivatives are developed as tunable IR detectors, modulators and emitters by electrophysiology and charge multiplexers [24]. This capability to organize the rearrangement and partition of surface electrons can be oppressed in emergent bio-imaging applications [25, 26].
The GO sheets are highly influenced by the divalent ions and some specific polymers which play a crucial role in mechanical properties of GO sheets by inter connecting both the molecules [27, 28]. Due to the superior mechanical properties of GO, it has been reported that by incorporating the different polymers with it, the tensile strength of the respective polymer increases. Graphene supported polymethyl methacrylate (PMMA) and poly-l-lactic acid (PLLA) drastically amplifies the young’s modulus and hardness of these polymer nanocomposites for mechanical applications [29].
With reference to different physicochemical properties of nanomaterials of graphene family, it can be projected that they will demonstrate numerous mutual connections with biological moieties such as cells and tissues depending on chemical modification, thickness and dimensions of graphene sheets, etc. [30, 31]. With more supplementary data of graphene application in biomedical field, research on its cellular activity and other intracellular processes is rising. Introduction of polar and reactive oxygenated functionalities give rise to oxidative force in objective cells is supposed to be the effective mechanism for potency of graphene oxide [32, 33]. Due to their dose dependent cytotoxicity and bactericidal activity, graphene-based materials are being explored for applications in antimicrobial products. A number of studies have been performed reporting the antibacterial activities of CNTs, graphene, GO and rGO against
Different methods are used to set up the preparation of graphene sheets according to its structural and chemical behavior with various biocompatible molecules. The methods are widely categorized as colloidal suspension (size specific), arc discharge (electric charge specific), and chemical or mechanical exfoliation. For the broad and extensive production of graphene sheets, mechanical exfoliation method were not used as it is expensive but for the fabrication of electronic devices it is widely applied. Graphene oxide (GO) the oxygenated derivative and replacement of graphene is synthesized by the chemical exfoliation method, in which the sp2-hybridized C▬C hybrid orbitals breaks and the different oxygenated groups such as hydroxyl, carboxyl and epoxy are introduced [39]. The surface modifications of the graphene sheets are site specific as the bulky group carboxyl attached toward the edges of the sheets while on the other hand hydroxyl and epoxy groups tends to form bond with the basal plane of the graphene sheets [40, 41]. These oxygenated and highly reactive functional groups offers reactive handles for a range of surface-functionalization reactions covalently and non-covalently, which can be used to build up surface modified GO, its biocompatible composites. For the large-scale synthesis of graphene, the most common methods required exfoliation of graphene. The only variances among graphene made by different methodologies are the defect content and yield of their products [42]. Various methods are available for graphene synthesis, but for the large production of graphene oxide (GO) oxidative-exfoliation methods give excellent results. There are some additional treatments required to reduce typically defective graphene-like nanosheet into reduced graphene oxide (RGO) [43]. During the oxidation process of graphene, functional groups containing oxygen attached to the surface increase the distance among graphitic layers and responsible for enhancing the exfoliation by weakening the van der Waals forces [44]. After the oxidation process, several washing steps are required so that oxidizing agents and some other impurities removed from graphite oxide to enhance the exfoliation. For large-scale washing of graphite oxide, different conventional approaches such as filtration process [45], centrifugation process [46], and dialysis process [47] are mostly used. Among all these processes infiltration processes, after some time, particles of graphite oxide choke the filter pores and make it time-consuming process.
For the synthesis of graphene oxide, the most used source of graphite is flake graphite, which occurs as a natural mineral which further use to purify to remove heteroatomic contamination [48]. GO prepared by the use of flake graphite have the property to easily dispersed in water hence used on a large scale [49]. This expanded form of graphite powder has been used for the synthesis of GO sheets by the following method.
Graphene oxide is synthesized by modified Hummer’s method using graphite powder [50]. In this method, in a round bottom flask, a mixture of 1 g of NaNO3 and calculated graphite powder are mixed. In this mixture drop by drop, 46 mL of H2SO4 was added in an ice water bath with continuous stirring. After 4 h stirring without any pause at 32°C temperature, 6 g of KMnO4 was poured into a slurry mixture of NaNO3 and graphite. Later 2 h continuous stirring 92 mL of DD water was combined in it at 95°C. Again after 2 h, 200 mL of DD water was poured in it and leave for 1 h for constant stirring. Lastly, at room temperature, 20 ml of H2O2 was mixed and mixed it repeatedly for 1 h. The obtained supreme oxidized product washes by 10% HCl solution for purification by abundant quantity by ion free water. Finally, it is filtered by 0.2 μm Nylon membranes until neutralizing the final product.
From the last decades, the interest has increased in the scientific community for the use of graphene oxide (GO) in biological and biomedical field applications [51]. GO is a two dimensional material with a large surface area containing single-layer sheets of carbon atoms with sp2 hybridization and carbon sites with sp3 hybridization in which hydrophilic functional groups with oxygen are present [52]. Thus, GO has many possibilities for surface functionalization due to outstanding solubility in water [53]. There are different methods for the functionalization of the surface of graphene oxide; some of them are discussed below (Figure 2).
Surface modification of graphene oxide.
The nanocarrier thus synthesized was characterized by some advanced spectroscopic techniques, like RAMAN (Figure 3), Fourier Transform infrared (FT-IR) (Figure 4), Transmittance Electron Microscope (TEM) (Figure 5). Thermal stability and quantitative analysis were characterized by Thermo gravimetric Analyzer TGA under a nitrogen atmosphere at a heating rate of 10°C/min from 30 to 600°C (Figure 6).
Raman spectra for GO, GO-PVP, and GO-β-CD.
Fourier transform infrared (FT-IR) spectra of GO, GO-PVP and GO-βCD.
TEM images of (a) GO (b) GO-PVP, and (c) GO-β-CD.
TGA analysis of GO, PVP, and GO-PVP.
During the processes of oxidation and exfoliation of graphite, there is a large extent of carboxylic group forms on the graphene surface. These groups better modified by different methods; one of them is covalent functionalization. In covalent functionalization, graphene is coupled with reagents, such as 1-ethyl-3-(3-dimethyllaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) [54], or can also be converted to acyl chlorides using thionyl chloride (SOCl) [55]. Covalent functionalization is a multipurpose methodology for modification of graphene surface which tailoring the chemical properties as well as electronic properties of graphene [56]. The methodology of GO functionalization by covalent functionalization depends on the environments of the reaction, nature of solvent, temperature condition, different functional groups of the incoming molecules, and other factors like reaction time. When polymer attach to the GO nanosheets it creates stress on GO surface, the covalent mode of functionalization is helpful in controlling the chemical properties of GO and reduces the stress caused by polymer [57]. GO surfaces have the ability for excellent covalent functionalization which makes it a unique nanomaterial which is helpful in developing studies of biological system. According to previous studies functionalization of GO shows excellent results when perform its use in targeted drug delivery applications [58]. There are chances of this because GO surface has the affinity for the adsorption of huge amount of hydrophobic drugs easily and due to specificity of covalent functionalization it releases the drug to particular regions of organisms. This functionalization of GO is also applicable in other biological activities like anti-bacterial activity, bioimaging [59], and photo-dynamic therapy [60]. Even though some procedures of graphene surface modification by covalent functionalization have validated efficient results but some methods generate some supplemental defects on the surface of graphene which are responsible for the changes in graphene structure.
Non-covalent functionalization is a more effective method in order to make maximum use of the inherent structure and mechanical properties of graphene oxide or graphene. Non-covalent functionalization is largely preferred in place of covalent functionalization as it does not alter the structure and electronic properties of graphene and it simultaneously introduces new chemical groups on the surface. The most common examples of non-covalent functionalization on graphene surface include polymer wrapping, π-π interactions, electron donor-acceptor complexes, hydrogen bonding, and van der Waals forces. Non-covalent functionalization of graphene results in the enhancement of dispersibility, biocompatibility, and reactivity, binding capacity, or sensing properties. Non-covalent interactions also known as supramolecular interactions are found in all types of materials that experience attractive as well as repulsive forces between them. These type of interactions are found in many natural and synthetic systems [61, 62]. In comparison to covalent bonds the energies of individual non-covalent interactions are normally lower [63]. In graphene, two types of π-π interactions occur between the electron-rich and electron-poor regions, which influence its interaction with other molecules or nanomaterials. This is commonly seen in the face-to-face and edge-to-face arrangement [64]. Graphene materials, with the π-π interactions have dissociation energies less than 50 kJ mol−1. The weakest forces, that is, London-dispersion forces or van der Waals interactions are responsible for the non-covalent interaction affect all atoms in close proximity. The hydrophobic effects caused by different types of interactions are influence not only dispersibility of GO but recognition interactions [65, 66].
In forthcoming nanotheranostics will be accepted as an efficient nanomedicine due to their unique properties like imaging, target selectivity and ability to load the drug in nanocarrier. In the process of nanotheranostics evolution as a potential nanoplatform various challenges encountered for detection of clinical complications. An appropriate technology is required for the treatment and selection of effective therapeutic agents for respective diseases like metallic nanocrystals, image-contrasting agents and choosing an efficient therapeutic agents for corresponding diseases like metallic nanocrystal and concatinate them as nanomaterial. The advanced nanomaterial high selectivity to the target site is required for advance nanotheranostic for excellent delivery of drug targeted nanotheranostic should contain delivery and loading capacity. The biocompatible material should be used in the preparation of nanotheranostics the normal tissue should not damage and easily excreted by human system. Whole designing of nanotheranostics will cheap with no side effects to body.
The introduction of a new drug to the site is not only expensive, but also time consuming. It includes discovery, clinical testing, development, and approval. Improving safety/efficacy ratio of marketed drugs is more cost-effective. All this can be done by controlling the time, rate, and place of drug release in the body through a drug-delivery system. Hence, a drug-delivery system could be seen as an interface between the patient and the drug [67]. Since past decades, a growing number of drugs were discovered and were optimized for an enhanced efficiency. However, about 40% of the new drugs, especially those based on biomolecules, like peptides, nucleotides, or proteins, often present a low bioavailability and are rejected by the pharmaceutical industry [68]. For controlled release the ideal materials must control certain important issues like easy reach to the target site in the body, ability to transport the necessary volume of active compounds, and a certain level of release with a certain speed, apart from the properties needed to ensure a better and safe interaction with the human body [69].
Graphene is considered as the finest and most durable monolayer capable of free existence. The specific 2D geometry and presence of pi electrons in graphene basal plane further applied for valuable drug loading via hydrophobic interactions and π-π stacking. Furthermore, large surface area of graphene allows for high density surface fabrication via different surface modification. A number of research on the in vivo behavior and bioactivity of graphene (Figure 7) has been investigated previously.
Graphene-based composites for various biomedical applications.
Graphene is the finest and most durable monolayer material which is capable of free existence. In graphene, its 2D structure and presence of delocalized π electrons on its surface can be used for effective drug loading via hydrophobic interactions and π-π stacking. In addition to this, large surface area of graphene allows it for high density bio-functionalization via both covalent and non-covalent surface modification methods. Various studies based on the in vivo behaviour and bioactivity of graphene shows that the nanocarriers interact with the cell membranes and enter into the cells by endocytosis. For targeted drug delivery to the cell nucleus, it is essential that the drug carrier escapes endosomal compartment and release loaded drug into the cytosolic compartments [70, 71]. This process proposed a strategy to reverse cancer drug resistance in DOX resistant MCF-7/ADR cells by loading DOX on graphene oxide surface via physical mixing [72]. High pH dependent release for drug loading with of DOX was observed
Attachment of nanoparticles such as iron oxide with graphene-based nanomaterial makes them super paramagnetic in property and can be useful in drug delivery applications [78]. The resulting magnetic hybrids dispersed uniformly in aqueous solution before and after loading of DOX. Magnetic hybrids show agglomeration behavior in acidic medium and redispersion behavior observed in basic medium. This pH triggered magnetic behavior of GO-Fe3O4 nanoparticle hybrids can be help in controlled drug delivery. Similar pH-dependent drug release system was reported for 5-FU-loaded nanohybrid system composed of graphene nanosheets (GN), carbon nanotube (CNT) and Fe3O4 [79].
Gene therapy is used in many expanding area to treat genetic disorders like Parkinson’s disease, cystic fibrosis and cancer. An effective gene therapy needs efficient and safe gene vectors that also protect DNA from nuclease degradation as well as facilitate DNA uptake with high transfection efficiency [80, 81]. According to review of literature, graphene has been reported for wide applications in the field of gene delivery, gene-drug co-delivery and protein delivery with. PEI has been extensively investigated as nonviral gene vector having strong electrostatic interactions with negatively charged phosphates of RNA and DNA. Chemical modification is very easy in PEI which offers increased transfection efficiency, cell selectivity and reduced cytotoxicity however low biocompatibility and high toxicity of (Polyethyleneimine) PEI limit its use for biomedical applications [82]. Chitosan-GO complex are also used for simultaneous drug and gene delivery [83]. Chitosan-GO converts pDNA into stable nano-sized complexes. Amine-terminated PEGylated GO was effectively used to deliver high protein payloads due to non-covalent interactions with surface of PEG-GO [84]. Bone morphogenic protein-2 (BMP-2) was loaded onto Ti substrate coated with alternate layers of positively (GO-NH3+) and negatively (GO-COO-) charged GO nanosheets with high loading efficacy and conserved bioactivity. Osteogenic differentiation of MSCs was enriched on Ti coated GO surfaces carrying BMP-2 than only Ti surface with BMP-2. In vivo studies in mouse also exhibited vigorous new bone formation with Ti-GO-BMP2 implants compared to Ti or Ti-GO or Ti-BMP2 implants and making the new composite a very effective carrier for therapeutic drug delivery [85]. All above studies have highlighted potential of graphene-based materials as drug and gene delivery vehicles
Tissue engineering is an interdisciplinary field that endeavors to develop biological substitutes to resolve, retain or enhance functionality of a tissue or whole organ [86]. Recently, graphene-based materials have been used to treat wound healing, stem cell engineering, regenerative medicine and tissue engineering. Hydrogels have viscoelastic and transport properties to mimicking natural tissues [87], but their weak mechanical properties can limit their use in many tissue engineering applications. Graphene has a platform for tailoring various functionalities on flat surfaces with outstanding mechanical properties like high elasticity, strength, flexibility. Potentially, graphene has a wide range of applications in the field of hydrogels, biodegradable films, electrospun fibers and other tissue engineering scaffolds. When GO incorporated into PVA-based hydrogels it potentially increases tensile stability (132%) and compressive strength (36%) of composite hydrogel without altering their cytoaffinity [88]. According to Lu et al. graphene-based composite materials are applicable for wound healing by formulating graphene containing chitosan-PVA nanofibrous scaffolds. These three groups, chitosan-PVA-graphene electrospun fibers, chitosan-PVA fibers were also studied without graphene and control (no scaffold), to check wound healing affinity in mice and rabbit [89]. Graphene containing samples healed the wound completely in faster rate in comparison to without graphene-based samples in both mice and rabbit. Graphene-based materials also have applications in the area of musculoskeletal tissue engineering using mouse myoblast C2C12 cell lines [90]. Cellular behavior on graphene derivatives are enhanced by the Surface roughness and surface oxygen content and adsorption of serum proteins. Thus, graphene materials can be useful in reinforcing tissue engineering scaffolds due to its mechanical and electrical properties. Graphene materials have properties like large surface area which adsorb proteins/DNA and can be useful in many therapeutic applications. For instance, Mahmoudi et al. [91] recently reported protective role of GO and protein-coated GO surfaces in amyloid beta fibrillation process, which is implicated in various neurodegenerative disorders. However, along with detailed
Graphene-based nanomaterials have been widely explored in biomedical fields such as bioimaging, drug delivery, theranostics, and so on. The recent advances in bioimaging of graphene-based nanomaterials, including graphene, graphene oxide, reduced graphene oxide, graphene quantum dots, and their derivatives, the synthetic methods of graphene-based nanomaterials are included in situ synthesis and binding method. The bioimaging modalities, including optical imaging (fluorescence [FL], two-photon FL), positron emission tomography/single-photon emission computed tomography, magnetic resonance imaging, photoacoustic imaging, Raman imaging, and multimodal imaging are highlighted (Figure 8).
Different types of bioimaging graphene-based bioimaging.
GO- and rGO-based composites are extensively used in the bioimaging field as the arrangement of tissues were comprehend with the help of optical imaging and with the application of unique characteristics of photon-based visible light [92]. As compared to the other progression GO derivatives has immense superiority including high-sensitivity non-ionizing energy, relative and economical advantage, real-time imaging, multiplexing capability, short range free space optical communication [93]. Although, this advancement is affected by low tissue penetration (0–2 cm), high tissue spreading of photons in the visible spectra region and considerable conditions because of tissue auto fluorescence and light absorption by macronutrients, oxygen binding groups and even by water molecules [94]. GO-based composites were dynamically developed for future aspects of different optical imaging techniques, such as (Fluorescence Imaging) FL imaging and (Two-Photon Fluorescence Imaging) TPFI.
Fluorescence imaging is a nonpersistent technique depends on the intensity of photons emitted from the probe used for fluorescent imaging [95]. A number of research groups focused on organic fluorescent dyes to fabricate GO and rGO
Due to some background noise disturb the fluorescence imaging due to single photon fluorescence imaging Two-Photon Fluorescence Imaging came into the picture. Two-Photon Fluorescence Imaging has fascinated much concentration because of its potential applications in fundamental study and medical diagnostics [101]. With the help of TPFI more detailed examination of various in vivo activities of deep located tumorous cells. Compared with one photon excitation using simple continuous-wave lasers, two-photon nonlinear excitation usually uses a nonlinear femtosecond laser to obtain a high reflux of excitation photons. Recently, graphene-based nanomaterials aroused considerable interest in the field of TPFI. Wang and Gu et al. first reported transferrin functionalized GO-PEG with strong two-photon luminescence as a nonbleaching optical probe for three-dimensional TPFI and laser-based cancer microsurgery, using an ultrafast pulsed laser as the excitation source [102]. Gong group employed nitrogen doping GQDs (N-GQDs) with an average size of ∼3 nm as efficient two photon fluorescent probes for cellular and deep-tissue imaging [103]. Taking dimethylformamide as solvent and nitrogen sources, the nitrogen was successfully doping to GQDs by a facile one-pot solvothermal method. Obviously, the TPFI using N-GQDs as fluorescent probe is particularly suitable for in vivo investigation of biostructures in the 800–1500 μm region.
Optical imaging cannot provide quantitative results and sometimes may be interfered by FL quenching of fluorescent dyes, light absorption and scattering of tissues, and autofluorescence background. Radiolabeling method would be able to accurately track the labeled substances in vivo in a quantitative manner with excellent sensitivity (∼10–11–10–12 mol/L) and nearly no penetration depth limit. The radionuclide-based imaging mainly contains PET and SPECT. PET and SPECT images are acquired over a nominally low background signal and require little signal amplification [104]. Graphene-based nanomaterials as promising nanoplatforms are playing an important role in PET/SPECT imaging. In 2011, Liu et al. reported a method to label nGO-PEG with 125I by anchoring iodine atoms on the defects and edges of GO [105]. After that a number of studies have been developed based on this method. Cai group explored in vivo active tumor targeting using 64Cu-labeled nGO-PEG [106]. In comparison to PET, SPECT is ∼10 times less sensitive (∼10–10–10–11 mol/L); however, SPECT enables concurrent imaging of multiple radionuclides of different energies [107]. Cornelissen et al. explored the use of anti-HER2 antibody (trastuzumab)-conjugated nGO, radiolabeled with 111In-benzyl-diethylenetriaminepentaacetic acid (BnDTPA) via π-π stacking, for in vivo targeting and SPECT imaging. The radiolabeled nGO-trastuzumab conjugates demonstrated better pharmacokinetics compared with radiolabeled trastuzumab without NGO, with more rapid clearance from the circulation [108].
Noninvasive technique MRI have been extensive used for the detection of morphological feature of tissue related bioscience in comparison to other optical imaging. But somehow the lower sensitivity for the detection of different concentration and inappropriate signaling time has been assigned as the huge drawbacks of MRI [109]. While the biomolecules and ions with paramagnetic nature of metal ions having manganese (Mn) and gadolinium (Gd) as major contributions are reported as the toxic in most of the cases [110]. Whereas such metallic ions can be incorporated with GO utilizing chelation procedure in between metals and different graphene layers [111]. The Gd (gadolinium) incorporated graphene oxide and amidoamine polymer dendrimer-based composite for the delivery of anti-cancerous drugs on the targeting sites have been reported by Wei et al. [112]. The composite of nitrogen doped graphene oxide have been studied for the detection of tumor containing sites on the defective cells. Starting from GO andiron hydrate the reduced graphene oxide-based composites were synthesized by Liu et al. following autoclave-based thermal treatment methodology. The hydrophobic interaction-based functionalization of polyetheneglycol and maleic anhydride-alt-1-octadecene molecule with iron-based nanoparticles were reported by the same to restore the magnetic properties along with the enhancement of thermal stability of developed solutions [113]. Graphene oxide/iron oxide nanoparticle-based system was fabricated to study and diagnosis of pancreatic cancer by Fu et al. [114]. The graphene and iron nanoparticle-based composite was reported as the magnificent composite to help the surgeons into the preparations of cancer cells. The dual mapping is main device that can easily radiate the difference in between normal and RLN tissues, thus further the lymph nodes can be treated with laser. The penetration effects of lower energy waves are much larger and deeper, while the radio waves worked as the low scatter for biomedical systems, i.e., tissues, cells or organs, etc. the PAI cells photographic technique, which take advantage of the absorption of longer wavelength containing waves into thermal energy for thermal expansion [115].
The reduced graphene nanomaterials are irreplaceable candidates to absorb near infra-red light in comparison to graphene oxide that reflects sp2 hybridization of carbon [116]. The reduced nature of reduced GO reflects hydrophobic nature of the graphene oxide thus finally result its poor water solubility. In order to find out a unique solution, the microwave heating-based-reduced graphene sheets having lower oxygen containing functional groups were synthesized by Patel et al. [117]. The Hummers method has been utilized to reduce the major oxygen content present in the graphitic powder, the methodology of Hummers method includes the acidic oxidations. The GO sheets can also be reduced to rGO by single step thermal reduction methodology and reported rGO possess excellent stability and lesser cell toxicity [118].
The advanced characterizations technique, i.e., the RAMAN technique is excellently advanced tool for the analytical and experimental extension for the detection of related various biochemical problems. The RAMAN spectrum including D and G bands exactly mentions and elaborates the enhancements of combinations of various nanoparticles [119]. The folic acid hybrid incorporated Ag/GO composite have been developed for specifically targeting of defective cancerous cells [120]. The in situ synthesis of gold- and GO-based composites have been incorporated in HeLa 229 cells which have been found to display excellent peaks and shifts in Raman spectra. The gold nanoparticle incorporated with nitrogen doped graphene oxide was reported by Ma and coworkers. The in situ synthesis of gold nanocomposites were also assumed to have physical forces of attraction between NOPs and gold particles. The further modifications of GO and reduced graphene sheets with 2-mercaptopyridine were reported by the non-covalent linkage [121]. Gold-based GO composites were reported for the development of good substrate than the Au NPs. Similarly, polyethylene glycol functionalized gold/copper nanoparticles along with graphene were incorporated through CVD method by the group of Tan et al. [122]. The unique Raman signals of graphite-based nanoparticles were reported along with further cell labeling and SERS detection. Recently, bio-imaging applications with more modalities have gained excellent popularity in recent decade [123].
The multimodality of such imaging applications has been referred as the better technique over the individual imaging technique for the higher accuracy and for the better diagnosis [124, 125]. While the multiphase analysis of single agent lack the potential problem on the probe, i.e., tissues blood for the further removal of impurities along with several doses [126]. Liu et al. have developed rGO-IONP for triple modulation, i.e., FL, PA along with MR [127]. The rGO-based composites of iron and GO have been synthesized via hydrothermal methodology where the polyethylene glycol was incorporated with poly(maleicanhydride-alt-1-octadecene) (C18PMH-PEG), further NIR was performed for the detection of magnetic absorbance. Similarly, Chen et al. reported the graphene oxide-based composite of PEG having non-covalent interactions, i.e., π-π stacking for the detection of tumorous cells. The recent reports of Wu et al. have synthesized the BaGdF5 nanoparticles which were reported to formulate on the graphene oxide sheets in moderate presence of polyethylene glycol. In the transmission electron microscopy (TEM), the exact morphology of GO/BaGdF5-based PEG composite was shown. This showed excellent separation of layers, along with the accuracy in size of sheets, where the size was reported to exist as smaller than corresponding pure GO sheets. While the SAED (selected area electron diffraction) spectra showed the excellently good crystal nature of BaGdF nanoparticles having cubic shape [128].
The unique ability of catenation of carbon and tendency to form various hybrid orbitals results in the formation of various smart compounds with different physical and chemical properties. Its 2D hexagonally packed unique structure of in-plane sigma C▬C bonds accounts for certain physical and chemical properties in biological media has led to its varied applications in the field of drug delivery, gene delivery, tissue engineering and various imaging techniques, etc. The electrostatic interaction and presence of metallic ions in biological media tends to agglomerate and reduces the surface area of graphene sheets. Therefore, covalent and non-covalent methods of surface modifications are used to increase the efficacy of graphene sheets. Further surface fabrication of GO with various polymers allows its use in the fields of drug delivery, tissue engineering and different imaging techniques. Surface modification by way of exfoliation is used for large scale synthesis of graphene. Modified Hummer’s method is a common procedure of synthesizing GO from the natural mineral source, i.e., flake graphite. The carboxylic group is found on the surface of graphite during the process of oxidation and exfoliation is modified by covalent functionalization making its use possible in the studies of biological systems and also found applications in biological activities like anti-bacterial activity, bioimaging and photodynamic therapy. Non-covalent functionalization has added advantage of not altering the structure or electronic properties of graphene while introducing new chemical groups on its surface. This results in enhancement of its dispersibility, biocompatibility, reactivity, binding capacity and sensing properties. Graphene has the properties of high surface region, distinctive geometry and structure, flexibility, extra ordinary physico-chemical properties, counting the high fracture strength, high Young’s modulus, great thermal and electrical conductivity, highly mobile charge carriers and biocompatibility. All mentioned properties makes graphene a valuable and important material for applications in biological systems and other biomedical processes.
In contrast to pristine graphene synthesized GO has high dispersibility in physiological media leading to better contact with biologically important organic molecules. Outstanding thermal and electrical conductivity and very low thermal expansion quotient of GO allows its use in energy conversion storage devices and bio sensors. GO derivative-based optoelectronic devices have been developed as IR detectors and electrophysiological modulators and emitters. The optical properties like intense light transmittance, fluorescence, photoluminescence and high mobility of charge make graphene an important material for application in MRI and biomedical imaging. Superior mechanical properties of GO like high tensile strength and extensive stiffness has enabled its use in the field of biomedical implant and tissue engineering. Cell-graphene and biomolecule-graphene studies have opened a vast area of GOs exploration in the fields of cellular biology, genomics and development of antibiotics, etc. Despite its varied uses certain challenges still remain in the field of nanotheranostic designing in terms of bioavailability, selectivity, biocompatibility and safety. In the field of pharmacology, better targeted and relatively lower dose drug delivery with graphene complex has proved cheaper and efficient than the discovery of newer drug. Graphene-based materials as drug and gene delivery vehicles have used successfully in
Authors acknowledge the financial assistance from NMHS research grant KU/NMHS/MG/2016/002/8603/007, GBPIHED, Kosi-Katarmal, Almora, India.
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