**Author details**

Lizhao Liu and Jijun Zhao\*

\*Address all correspondence to<EMAIL_ADDRESS>
Key Laboratory of Materials Modification by Laser, Ion and Electron Beams (Dalian Univer‐ sity of Technology), Ministry of Education, Dalian 116024, China

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**Section 2**

**Electrical and Biomedical Applications of Carbon**

**Nanotubes**

**Electrical and Biomedical Applications of Carbon Nanotubes**

**Chapter 13**

**Carbon Nanotubes for Use in Medicine: Potentials and**

Structurally, Carbon nanotubes (CNTs) can be viewed as wrapped from graphene sheets. Single-walled carbon nanotubes (SWNTs) have one layer of graphene sheet, whereas, the multiwalled carbon nanotubes (MWNTs) contain multi layers of graphene sheets. The wellordered molecular structure brings CNTs many remarkable physical properties, such as, ex‐ cellent mechanic strength, ultrahigh surface area, high aspect ratio, distinct optical properties [1], and excellent electrical conductivity [2]. In last decade, CNTs are intensively explored for in-vitro and in-vivo delivery of therapeutics, which was inspired by an impor‐ tant finding that CNTs can penetrate cells by themselves without apparent cytotoxic effect to the cells [3]. The high aspect ratio makes CNTs outstanding from other types of round nanoparticles in that the needle-like CNTs allow loading large quantities of payloads along the longitude of tubes without affecting their cell penetration capability. With the adequate loading capacity, the CNTs can carry multifunctional therapeutics, including drugs, genes and targeting molecules, into one cell to exert multi-valence effects. In the other side, owing to the ultrahigh surface area along with the strong mechanical properties and electrically conductive nature, CNTs are excellent material for nanoscaffolds and three dimensional nanocomposites. In recent year, CNT-based devices have been successfully utilized in tissue engineering and stem cell based therapeutic applications, including myocardial therapy, bone formation, muscle and neuronal regeneration. Furthermore, owing to the distinct opti‐ cal properties of CNTs, such as, high absorption in the near-infrared (NIR) range, photolu‐ minescence, and strong Raman shift [4], CNTs are excellent agents for biology detection and imaging. Combined with high surface area of CNTs for attaching molecular recognition molecules, CNT-based, targeted nanodevices have been developed for selective imaging

> © 2013 Shao et al.; licensee InTech. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

© 2013 Shao et al.; licensee InTech. This is a paper distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

distribution, and reproduction in any medium, provided the original work is properly cited.

**Limitations**

**1. Introduction**

Wei Shao, Paul Arghya, Mai Yiyong, Laetitia Rodes and Satya Prakash

http://dx.doi.org/10.5772/51785

Additional information is available at the end of the chapter


*3.4. Microbial Community Analysis*

Once the experiment was completed, a sample was taken for the analysis of microbial communities. For this purpose, a 1 × 1 cm portion of carbon felt was cut using sterilised scissors. Genomic DNA from the electrodic sample was extracted with the PowerSoil® DNA Isolation Kit (MoBio Laboratories Inc., Carlsbad, CA, USA), following the manufacturer's instructions. The entire DNA extract was used for the pyrosequencing of the 16S-rRNA gene-based massive library, targeting the eubacterial region V1–V3 16S-rRNA, and performed at MR DNA (www.mrdnalab.com, Shallowater, TX, USA), utilising MiSeq equipment (Illumina, San Diego, CT, USA). The primer set used was 27Fmod (50 -AGRGTTTGATCMTGGCTCAG-30 ) /519 R modBio (50 -GTNTTACNGCGGCKGCTG-30 ). Diluted DNA extracts were used as a template for polymerase chain reaction (PCR) reactions. The obtained DNA reads were compiled in FASTq files for further bioinformatics processing. The trimming of the 16S-rRNA bar-coded sequences into libraries was carried out using QIIME software, version 1.8.018 [49]. Quality filtering of the reads was performed at Q25 quality prior to grouping into operational taxonomic units (OTUs) at a 97% sequence homology cut-off. The following steps were performed using QIIME, a denoising procedure using a denoiser algorithm [50]. Final OTUs were taxonomically classified using BLASTn against a database derived from the Ribosomal Database Project II (RDPII, http://rdp.cme.msu.edu) and the National Centre for Biotechnology Information (NCBI, www.ncbi.nlm.nih.gov).

## **4. Conclusions**

The proliferation and biocatalytic activity of *Geobacter* in air-exposed anodes depends on the anaerobic microenvironment provided by the aerobic microorganisms present in outer layers of the biofilm, which consume DO. The porous structure of the carbon felt causes the channelling of DO in internal zones. This study confirms *Geobacter*'s tolerance to sustained oxygen concentrations below 90 µM. During periods in which the biofilm is maintained without substrate, oxygen diffuses through the biofilm height. If these periods without substrate are longer than six days, the inner electroactive layers seem to be irreversibly affected, and the exoelectrogenic activity does not resume again after feeding.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/2073-4344/10/11/1341/s1, Figure S1: Dissolved oxygen profile. Rejected because the data acquisition system was accidentally interrupted at the end of the experiment; Figure S2: Another dissolved oxygen profile. Rejected because the data acquisition system was accidentally interrupted at the end of the experiment.

**Author Contributions:** R.M.A. contributed to the conceptual design of the experiment, carrying out the experimental procedures, data analysis, results interpretation and drafting the manuscript. G.P.: performed the microbiological analysis and provided assistance in the laboratory. M.I.S.-M. also provided assistance in the laboratory and contributed to results' interpretation. A.M.: contributed to results interpretation, being responsible for project administration and funding acquisition. A.E.: contributed to the conceptualization, results interpretation, project supervision and writing/reviewing the manuscript. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was possible thanks to the financial support by 'Consejería de Educación de la Junta de Castilla y León' (ref: LE320P18), a project co-financed by FEDER funds. R. M. Alonso thanks the University of León for his predoctoral contract.

**Conflicts of Interest:** The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

## **References**

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).


*3.1.3. Related datum features with maximum material requirement*

The maximum material requirement for datum features results in the following three inde‐ pendent requirements:A requirement for the surface non-violation of the maximum material virtual condition (MMVC). The maximum material virtual condition (MMVC) of the related datum feature shall not be violated by the extracted (integral) datum feature from which the datum is derived. A requirement for MMS when there is no geometrical tolerance or when there is a geometrical tolerance not followed by the symbol M. When the related datum feature has no geometrical tolerance or has a geometrical tolerance of form not followed by the symbol M, the size of the maximum material virtual condition (MMVC) of the related datum feature is the maximum material size (MMS). A requirement for MMS when there is a geometrical tolerance of form followed by the symbol M. When the datum feature has a geometrical tolerance of form and this tolerance is followed by the symbol M (see Figure 8), the size of the maximum material virtual condition (MMVC) of the related datum feature is the maximum material size (MMS) plus (for external features of size) or minus (for internal features of size) the geometrical tolerance.

The symbol M is placed on drawings after the datum letter(s) in the tolerance indicator, when MMR applies to the datum feature. If the datum is obtained from a feature of size, the use of symbol M after the datum letter is only possible indication. The corresponding sequence of letters identifying the common datum is indicated within parentheses, when maximum or least material requirement applies to all elements of the collection of surfaces of a common datum. The sequence of letters identifying the common datum is not indicated within parentheses, when maximum or least material requirement applies only to one element of the Application of New Generation Geometrical Product Specifications—Position Tolerancing http://dx.doi.org/10.5772/65412 157

**Figure 8.** Example of LMR for two concentric cylindrical features (internal and external) both controlled by size and location (position) to the same datum systems A and B.

collection of surfaces of a common datum. In this case, the requirement applies only to the feature identified by the letter placed just before the modifier, and it specifies for the surface(s) in the following rules.

## *3 1.4. Related datum features with least material requirement*

An example with the assembly comprising a plate with two holes 25 mm apart is illustrated in Figure 7b. There is a requirement about holes to be perpendicular to the contact surface of the plate. The following interpretation is according to rules and definitions given in ISO

Proceedings of the International Conference on Interdisciplinary Studies (ICIS 2016) - Interdisciplinarity and Creativity

**a.** The extracted feature of the toleranced pins shall not violate the maximum material virtual

**b.** The extracted feature of the toleranced pins shall have everywhere a local diameter larger than LMS = 9.8 mm. The RPR requirement allows the size tolerance to increase.

The location of the two MMVCs is theoretically correct —at a distance of 25 mm relative to each other and perpendicular to the datum A. The RPR allows the dimensional tolerance increasing when the geometrical deviation does not take full advantage of the maximum

In this case, the RPR indicated on drawings by the symbol R placed after the symbol L itself placed after the geometrical tolerance of the derived feature of the feature of size in the

The maximum material requirement for datum features results in the following three inde‐ pendent requirements:A requirement for the surface non-violation of the maximum material virtual condition (MMVC). The maximum material virtual condition (MMVC) of the related datum feature shall not be violated by the extracted (integral) datum feature from which the datum is derived. A requirement for MMS when there is no geometrical tolerance or when there is a geometrical tolerance not followed by the symbol M. When the related datum feature has no geometrical tolerance or has a geometrical tolerance of form not followed by the symbol M, the size of the maximum material virtual condition (MMVC) of the related datum feature is the maximum material size (MMS). A requirement for MMS when there is a geometrical tolerance of form followed by the symbol M. When the datum feature has a geometrical tolerance of form and this tolerance is followed by the symbol M (see Figure 8), the size of the maximum material virtual condition (MMVC) of the related datum feature is the maximum material size (MMS) plus (for external features of size) or minus (for internal features of size)

The symbol M is placed on drawings after the datum letter(s) in the tolerance indicator, when MMR applies to the datum feature. If the datum is obtained from a feature of size, the use of symbol M after the datum letter is only possible indication. The corresponding sequence of letters identifying the common datum is indicated within parentheses, when maximum or least material requirement applies to all elements of the collection of surfaces of a common datum. The sequence of letters identifying the common datum is not indicated within parentheses, when maximum or least material requirement applies only to one element of the

tolerance indicator to alter the least material requirement for the surface(s).

*3.1.3. Related datum features with maximum material requirement*

condition (MMVC), which has the diameter of 10.3 mm.

2692:2014:

156

in the Knowledge Society

material virtual condition (MMVC).

the geometrical tolerance.

*3.1.2. Least material requirement and RPR*

When the toleranced feature applies to the least material requirement (LMR), the indication on the drawing is the symbol L placed after the geometrical tolerance of the corresponding feature of the size in the indicator of the tolerance.

The symbol L is applied to the tolerancing of the features on both sides of the wall to fully control the minimum wall thickness. LMR can be implemented in two different ways, as follows.

In Figure 8, it shows the location tolerance for the two different sides of the wall, which can refer to the same datum axis or datum system. The symbol L, in this case, applies to the two toleranced features.

The intended function of the part illustrated in Figure 8 is the ability to resist internal pressure and prevent breakout.

The interpretation is based on the following rules and definitions given in ISO 2692:2014:

	- **c.** The diameter of the internal feature of the least material virtual condition is LMVC = 35.2 mm, and the LMVC shall be fully contained in the material.
	- **d.** Everywhere a local size of the extracted feature of the internal feature shall be larger than MMS = 35.0 mm and smaller than LMS = 35.1 mm.
	- **e.** The least material virtual condition (LMVC) of both the external and the internal features shall be in a theoretically exact orientation and the location relative to the datum system at a position of 44.44 mm.

It is permissible to have reference between location tolerances of the derived features for both sides of the wall as the datum. The tolerance for the toleranced feature and the datum letter is followed by the symbol L in this case.

**Figure 9.** Examples of application of the new concepts to indicate the positional deviation.

If the datum is obtained from a feature of size, the use of the symbol L after the datum letter is only possible. The corresponding sequence of letters identifying the common datum is indicated within parentheses, when maximum or least material requirement applies to all elements of the collection of surfaces of a common datum. The sequence of letters identifying the common datum is not indicated within parentheses, when maximum or least material requirement applies only to one element of the collection of surfaces of a common datum. In this case, the requirement applies only to the feature identified by the letter placed just before the modifier.

Figure 9 shows practical examples of application of the new concepts to indicate the positional deviation in the drawings. In Figure 9 a, dimensions are indicated by E, if required a high mounting requirement is used the symbols for MMC and LMC (see Figure 9b) and O M inform to envelop requirements with connection to form and location tolerances (see Figure 9c). Some instructions about selecting tolerance of position control modifiers are given in Figure 10.

**Figure 10.** Instruction about selecting tolerance of position control modifiers.

ISO 5458 [2] describes the principle of positional tolerancing for the location of regular and irregular features. However, for clarity, only regular-shaped features such as holes, bolts, studs or pins, parallel-sided slots, keys and keyways have been shown.

In Table 1, some specification modifiers [2] are given.

**Table 1.** Specification modifiers for linear size

**c.** The diameter of the internal feature of the least material virtual condition is LMVC = 35.2

Proceedings of the International Conference on Interdisciplinary Studies (ICIS 2016) - Interdisciplinarity and Creativity

**d.** Everywhere a local size of the extracted feature of the internal feature shall be larger than

**e.** The least material virtual condition (LMVC) of both the external and the internal features shall be in a theoretically exact orientation and the location relative to the datum system

It is permissible to have reference between location tolerances of the derived features for both sides of the wall as the datum. The tolerance for the toleranced feature and the datum letter is

mm, and the LMVC shall be fully contained in the material.

**Figure 9.** Examples of application of the new concepts to indicate the positional deviation.

If the datum is obtained from a feature of size, the use of the symbol L after the datum letter is only possible. The corresponding sequence of letters identifying the common datum is indicated within parentheses, when maximum or least material requirement applies to all

MMS = 35.0 mm and smaller than LMS = 35.1 mm.

at a position of 44.44 mm.

in the Knowledge Society

158

followed by the symbol L in this case.

By default, based on the independency principle defined in ISO 8015 [4], a geometrical specification of form, orientation, location or run-out, without modifier, applied to *n* geomet‐

rical features (*n* being greater than one) is equivalent to *n* independent geometrical specifica‐ tions: each geometrical feature shall be considered individually and each specification shall be considered individually (independent between them). The resulting independent tolerance zones correspond to an implicit indication of the separate zone (SZ) modifier: the "all around" modifier does not create itself a united feature or a pattern.

When positional tolerancing is applied to several geometrical features and all the nonredundant degrees of freedoms of the tolerance zones are unlocked, either the separate zone (SZ) modifier or the combined zone (CZ) modifier shall always be indicated in the second compartment of the tolerance indicator. To create one homogeneous pattern, the CZ modifier shall be indicated in the tolerance section of the tolerance indicator. To create a new level of pattern defined as a homogeneous pattern of more than one homogeneous pattern, an additional CZ modifier shall be indicated after the sequence defining the previous level of pattern. One CZ in the tolerance section defines a single pattern. The sequence CZ in the tolerance in the tolerance section defines a pattern of patterns of patterns (pattern of Level 3).

**Figure 11.** Example of identification of a pattern for repetition.

Figure 11 shows four specifications: The first specification (CZ SZ without datum) manages four independent patterns. For each pattern, the specification considers the following: As a toleranced feature, the collection of two extracted axes As a tolerance zone, the combined zone composed of two cylindrical zones of diameter 0.2 mm constrained in location between them at 17 mm. The second specification (CZ CZ without datum) manages four dependent patterns, resulting in only one specification considering the following: As a toleranced feature, the collection of eight extracted axes As a tolerance zone, the combined zone composed of eight cylindrical zones of diameter 0.4 mm constrained in location between them at 17 mm in a direction and 30 mm in a perpendicular direction. The third specification (CZ SZ with datum B) manages four independent patterns constrained in location from B. For each pattern, the specification considers the following: As a toleranced feature, the collection of two extracted axes As a tolerance zone, the combined zone composed of two cylindrical zones of diameter 0.3 mm constrained in location between them at 17 mm and constrained from datum B at 20 mm. The fourth specification (CZ CZ with datum systems B and A) manages four dependent patterns, constrained in orientation from datum B (perpendicularly) and in location from datum A, resulting in only one specification considering the following: As a toleranced feature, the collection of eight extracted axes As a tolerance zone, the combined zone composed of eight cylindrical zones of diameter 0.8 mm constrained in location between them at 17 mm in one direction and 30 mm in a perpendicular direction and constrained from the datum systems B and A, respectively, at 20 and 24 mm.

To avoid ambiguity, when a positional specification applies to several features and not all of the non-redundant degrees of freedom for the tolerance zones are locked, either an SZ modifier or a CZ modifier is always indicated.

All types of geometrical characteristic symbol can be used to establish a geometrical specifi‐ cation in a pattern. However, to create a pattern specification, a CZ modifier can be indicated in the second compartment of the tolerance indicator.


*4.1.3 Diagnostic fluid markers in ascites*

The colour of ascetic fluid is a very essential marker in the diagnosis of the cause of ascites. The colour ranges from clear fluid to yellowish, reddish and opaque with flakes of fibrin and debris depending on the aetiology. It is therefore important to observe the above guidelines in paracentesis in order to avoid false discolouration of the fluid, thus affecting correct diagnosis. Various colours of ascetic fluid signify a different aetiology as shown below [37]:

## *4.1.4 Clinical examination (author's personal experience)*

Clinical examination is considered a useful tool in the identification of the cause of ascites. An elevated temperature would signify an underlying infectious or inflammatory condition such as bacterial tuberculosis. An elevated capillary refill time would signify a decreased circulatory volume as a result of cancerous or infectious

**Figure 1.** *Removal of ascites through the linea alba in an Alsatian breed of dog.*

condition. Auscultation of the heat reveals various cardiovascular diseases such as muffled heart sound which is consistent with pericardial effusion and cardiac tamponade. Heart murmurs or irregular heartbeats are suggestive of right-sided heart failure. An elevated heartbeat or tachypnoea may result from dyspnea due to cranial displacement of the diaphragm into the thoracic cavity. Cardiovascular abnormalities are confirmed through the use of electrocardiograph and echocardiography.


**5.2 CCUS: EOR potential in the Eastern Llanos basin**

The review of progress in the different projects of the hydrocarbons sector in the Eastern Llanos basin means that currently there is sufficient infrastructure to move forward the capture and injection of CO2 for improved recovery, subtracting the construction of CO2 liquefaction facilities with sufficient

processing capacity, as well as facilities for the storage of liquid CO2. The coal power plants in the Oriental Cordillera and the natural gas deposits in the basin are potential CO2 sources.

After researching the many formations that make up the Llanos Basin, it is concluded that the Carbonera formation is the most favorable for CCUS because it has sandy interbeddings that are not being used (levels C-7, C-5, C-3, and C-1) and that they can be favorable as a storage formation due to their storage capability in a future CO2 capture project, all these are conditions to elaborated surveys of characterization of sand formation. Additionally, Carbonera is at a higher depth than the last potentially usable water source (Guayabo aquifer), isolated by clay layers that form natural hydraulic seals (levels C-2, C-4, C-6, and C-7) and covered by the regional seal of the León Formation (**Figure 6**).

The four clayey members of Carbonera add up to about 637 feet, which with low permeability provides the natural separation of the disposal area with the other units, guaranteeing the non-affectation to the "surface" aquifers in the Guayabo and Necesidad formations (**Figures 7** and **8**) [12].

In the Carbonera formation, porosities are comparatively invariant (18–23%) as well as permeable (100–3500 mD), suggesting that the units that are projected as receptors have stable petrophysical characteristics that allow the entry of a volume of injected CO2 without causing damage to the formation.

Porosities were measured through density and neutron curves taken in the wells. Regarding permeability, despite not having data in the aforementioned cores, a theoretical curve was obtained and then corrected and tuned with cores and logs from other blocks [13].

The Carbonera formation is not the only one with CO2 storage expectations, there are also the Mirador, Barco, Guadalupe, and Une formations that have even better values for a storage formation, and therefore, the best hydrocarbon reservoirs found in the Llanos basin are in these formations; however, as CO2 capture prospects, these formations could be used for secondary recovery (EOR/EGR) and obtain better results in production.

## **6. Conclusions**

The geological depository of CO2 should be considered as one of the most viable options to minimize the effects and emissions of polluting gases into the atmosphere related to fossil fuels.

At the time of reviewing the existing documentation on the method of capture and storage or sequestration of CO2, it is clear that in Colombia there is no current regulation on CCUS–EOR technologies, given that they are still in the experimental phase and the projects that are starting are pilots, or there are very few who have come to implement the technique.

The maturity of the Llanos basin and its tectonic evolution, as well as the stratigraphic and petrophysical characteristics of the large oil and gas fields with hydrodynamic capture, and good regional seals, lead to the conclusion that in the basin there are formations with good possibilities to store or sequester CO2; being the odd sandy intervals of the Carbonera formation and more specifically C-1 and C-7, the most suitable for the application of this method.

The technologies implemented within the hydrocarbons sector favor in a certain way the development of the CCUS method in Colombia, this is a considerable advantage compared to countries that do not have a presence of oil activity.

*Carbon Capture, Use and Storage (CCUS) as Enhanced Oil Recovery (EOR): Llanos Orientales… DOI: http://dx.doi.org/10.5772/intechopen.105949*


*3.4. Biological Assay*

The antimicrobial activities of compounds **1**–**12** against *Staphylococcus aureus*, *Escherichia coli*, and *Bacillus subtilis* were evaluated using a previous method [16], and levofloxacin was used as a positive control. The cytotoxicities of compounds **1**–**12** against DU145, SW1990, Huh7, and PANC-1 human cancer cell lines were determined using the CCK-8 method [17], with cisplatin used as a positive control. The immunosuppressive activities of compounds **1**–**12** against ConA-induced T cell proliferation were performed as previously described [18], with cyclosporin A as a positive control.


**5. Pathophysiology**

No definitive causative has been defined as regards the pathogenesis of CIN in the literature. The most accepted theory for the development of CIN following contrast administration relies upon the vasoconstriction of the vessels in the renal medulla leading to reduced oxygen delivery [40]. Reduced oxygen delivery and prolonged vasoconstriction leads to enhanced production of oxygen-free radicals like hydrogen peroxide and superoxide leading to increased damage [41]. Other suggested causes of this condition are elevated blood viscosity, reperfusion injury, direct toxic damage to the cells, and the release of dopamine, angiotensin II, and vasopressin. These factors induce further vasoconstriction and damage the renal function [42]. The use of supportive therapy in critically ill patients, for instance, the mechanical ventilation and inotropic drugs, and the variable health risks this group of patients have as anemia and sepsis have been shown to increase additional kidney damage resulting from vasoconstriction and hypoxia [43].

#### **5.1 Regional hypoxia as a cause of contrast-induced nephropathy (CIN)**

Renal perfusion is very high in the cortex. The medullary portions are maintained at the border of hypoxia such that pO2 levels can be as low as 20 mmHg [44]. This is the price for maintaining the countercurrent mechanism used for the control of urine excretion. A vulnerable kidney region is the deeper part of the outer medulla, an area far from the vasa recta which supply the renal medulla with blood. It is there where the thick ascending limbs of the loop of Henle encounter hypoxic damage [45]. The cause for the vulnerability of the outer medullary portion of the nephron is the relatively higher oxygen needs due to salt reabsorption. The addition of contrast media to the medium augments the hypoxic injury imposed upon this region by increasing the renal vascular resistance, as seen in the rat model [46]. A second factor that has been postulated to mediate CIN is

**125**

*Contrast-Induced Nephropathy*

limb is the Na-K+

consumption.

*DOI: http://dx.doi.org/10.5772/intechopen.90457*

an elevated oxygen demand due to an enhanced workload in the tubular cells. First, there is a temporary increase in GFR that follows giving contrast media [47], and second, osmotic diuresis may reduce the para-cellular absorption of the proximal tubular cells, causing a greater load of NaCl that has to be absorbed in the distal tubules. Liss et al. [48] have shown that contrast media can enhance medullary blood flow to the kidney, although pO2 decreases, which is supported also by a study of Heyman et al. [49]. These two studies suggest that an increased oxygen demand has taken place following contrast media injection. Local kidney hypoxia can be augmented by the systemic effects of some contrast media, such as transiently diminished cardiac output [50] and abnormal pulmonary ventilationperfusion relationship [51]. Also, the amount of oxygen delivered to the peripheral tissues might be impaired, since contrast media can increase oxygen affinity of hemoglobin [52]. If renal outer medullary hypoxia causes CIN, the blockade of the transporters in this portion of the nephron must have helpful effects on its prevention. The major part of the transport that occurs in the medullary thick ascending

furosemide. Blocking this transport would significantly decrease the local oxygen

Renal blood flow and intrarenal microcirculation are markedly altered by contrast medium (CM) [53]. The extent and distribution of renal hemodynamic changes depends on the species investigated as well as on the type, volume, and rate of contrast administration. Moreover, renal hemodynamic effects of CM gravely depend on the hydration state and on additional predisposing factors that may have an effect on the renal circulation and the tone of the renal vasculature, such as chronic renal disease, diabetes mellitus, senility, and inflammation. In an early study in dogs subjected to a high-osmolar CM, total renal blood flow was transiently enhanced for 5–15 min, followed by a decline by 25% below baseline, reaching beyond a couple of hours [54]. In healthy humans, renal blood flow fell 8% over 30 min after the intravenous administration of conventional doses of CM [53]. On the contrary, in patients with chronic kidney disease undergoing coronary angiography, a transient short augmentation of renal blood flow was followed by a

Investigations performed in vitro on cell lines are used to assess the renal tubular cell function or damage. A porcine cell line of renal proximal tubules, LLC-PK1, was used by Hardiek et al. [55] to investigate the occurrence of CIN. An effect on apoptosis was not elucidated, even though proliferation was impaired. Diminished proliferation will have a hazardous effect on renal function with a delay of hours to days, which may help in explaining the course of CIN. Independent of the contrast media used, tubular cell damage can occur. A more specific distortion of proximal tubular function seems to be a perturbation of mitochondrial enzymatic activity and mitochondrial membrane potential [56]. The degree of mitochondrial enzymatic activity impairment depends mainly on two features of the contrast media: the ionic nature as well as the molecular structure. Remarkably, low-osmolar contrast media had the least observed effects, followed by the iso-osmolar contrast media. Ionic compounds showed the most deleterious effects [56]. In the distal tubule, contrast media may trigger apoptotic effects in the cells, as depicted in the

**5.2 Radiocontrast-mediated changes in renal blood supply**

dramatic 40% decline, lasting for more than 3 h [54].

Madin-Darby canine kidney (MDCK) cell line model [57].

**5.3 Cytotoxic effects on renal tubular cells**

#### *Contrast-Induced Nephropathy DOI: http://dx.doi.org/10.5772/intechopen.90457*

*New Insight into Cerebrovascular Diseases - An Updated Comprehensive Review*

Most side effects attributable to contrast media are linked to its hypertonicity. Currently, four main types of contrast media are used in practice today, including nonionic low-osmolar, ionic low-osmolar, nonionic iso-osmolar, and ionic high-osmolar contrast media. In a large study which compared the nonionic lowosmolality agent iohexol to the ionic high-osmolality agent meglumine/sodium diatrizoate in patients with preexisting renal impairment undergoing angiography, patients with renal impairment who received diatrizoate were 3.3 times more liable to develop CIN in comparison with those receiving iohexol [38]. NEPHRIC trial is a randomized, prospective study that made a comparison between the nonionic iso-osmolar CM iodixanol with the nonionic low-osmolar CM iohexol in 129 patients with renal impairment and diabetes undergoing coronary or aortofemoral angiography. The incidence of CIN was 3% in the iodixanol group and 26% in the iohexol group (p = 0.002) [39]. In one other randomized study, the incidence of CIN provoked by iodixanol and iohexol was compared in 124 patients with basal creatinine level >1.7 mg/dl. The incidence of CIN was 3.7% in iodixanol group and 10% in iohexol group (p > 0.05) [17]. In addition, CM are classified as ionic and nonionic. A randomized trial of 1196 patients performing coronary angiography showed that nonionic CM lowered the incidence of CIN in patients with preexisting renal impairment [38]. In high-risk patients, it is better to avoid the use the high-

No definitive causative has been defined as regards the pathogenesis of CIN in the literature. The most accepted theory for the development of CIN following contrast administration relies upon the vasoconstriction of the vessels in the renal medulla leading to reduced oxygen delivery [40]. Reduced oxygen delivery and prolonged vasoconstriction leads to enhanced production of oxygen-free radicals like hydrogen peroxide and superoxide leading to increased damage [41]. Other suggested causes of this condition are elevated blood viscosity, reperfusion injury, direct toxic damage to the cells, and the release of dopamine, angiotensin II, and vasopressin. These factors induce further vasoconstriction and damage the renal function [42]. The use of supportive therapy in critically ill patients, for instance, the mechanical ventilation and inotropic drugs, and the variable health risks this group of patients have as anemia and sepsis have been shown to increase additional

kidney damage resulting from vasoconstriction and hypoxia [43].

**5.1 Regional hypoxia as a cause of contrast-induced nephropathy (CIN)**

Renal perfusion is very high in the cortex. The medullary portions are maintained at the border of hypoxia such that pO2 levels can be as low as 20 mmHg [44]. This is the price for maintaining the countercurrent mechanism used for the control of urine excretion. A vulnerable kidney region is the deeper part of the outer medulla, an area far from the vasa recta which supply the renal medulla with blood. It is there where the thick ascending limbs of the loop of Henle encounter hypoxic damage [45]. The cause for the vulnerability of the outer medullary portion of the nephron is the relatively higher oxygen needs due to salt reabsorption. The addition of contrast media to the medium augments the hypoxic injury imposed upon this region by increasing the renal vascular resistance, as seen in the rat model [46]. A second factor that has been postulated to mediate CIN is

*4.14.2 High-osmolar and ionic CM*

osmolar and ionic CM to lower the risk of CIN.

**5. Pathophysiology**

**124**

an elevated oxygen demand due to an enhanced workload in the tubular cells. First, there is a temporary increase in GFR that follows giving contrast media [47], and second, osmotic diuresis may reduce the para-cellular absorption of the proximal tubular cells, causing a greater load of NaCl that has to be absorbed in the distal tubules. Liss et al. [48] have shown that contrast media can enhance medullary blood flow to the kidney, although pO2 decreases, which is supported also by a study of Heyman et al. [49]. These two studies suggest that an increased oxygen demand has taken place following contrast media injection. Local kidney hypoxia can be augmented by the systemic effects of some contrast media, such as transiently diminished cardiac output [50] and abnormal pulmonary ventilationperfusion relationship [51]. Also, the amount of oxygen delivered to the peripheral tissues might be impaired, since contrast media can increase oxygen affinity of hemoglobin [52]. If renal outer medullary hypoxia causes CIN, the blockade of the transporters in this portion of the nephron must have helpful effects on its prevention. The major part of the transport that occurs in the medullary thick ascending limb is the Na-K+ -2Cl− cotransporter, which, as mentioned above, is blocked by furosemide. Blocking this transport would significantly decrease the local oxygen consumption.


**Abstract**

The building sector accounted for the largest share of both global final energy use and energy-related CO2 emissions. Despite the efforts made during the last decade to reduce energy consumption and greenhouse gas emissions, the demand for energy is increasing steadily. Thus, development of novel strategies to reduce energy costs and save the environment through a new building regulation has critical importance. Several new technologies are emerging to help achieve the aim of reducing energy usage in building sectors, eliminating greenhouse gas emissions, and recycling waste. Some of these technologies are: (1) the development of geopolymer binder that may be used as an alternative to ordinary Portland cement, (2) the adoption of three-dimensional (3D) printing technology in the civil engineering, and (3) the integration of phase change materials (PCM) in cementitious materials to increase energy efficiency of buildings. In this chapter we review some research about phase change materials-based geopolymer cement, and the adoption of the additive manufacturing technology in geopolymer applications, as well as, point to further areas of study required for wide-scale industry adoption.

**Keywords:** Aluminosilicate materials, Alkaline activator, Geopolymer, Phase change material, 3D printing


Anthracnose of Chilli: Status, Diagnosis, and Management

*Raj Kiran, Jameel Akhtar, Pardeep Kumar and Meena Shekhar*

### **Abstract**

Chilli (*Capsicum annuum* L.) is one of the most economically important vegetable crops in the world. Among different biotic constraints, anthracnose disease is the major limiting factor affecting yield and production of chilli crop. Different symptoms associated with disease are fruit rot, leaf spots, dieback on stem, seedling blight, or damping off. Many species of genus *Colletotrichum* are found associated with the disease worldwide. In India, primarily three important species, namely, *Colletotrichum truncatum*, *C. acutatum*, and *C. gleosporoides*, are responsible for the chilli anthracnose. Accurate identification of pathogen is needed for choosing the proper management strategy for controlling this disease. Both conventional and molecular methods are adapted along with different management strategies, recommended for this disease namely cultural, chemical, and other eco-friendly methods.

**Keywords:** *Colletotrichum*, biocontrol agents, diagnosis, molecular methods

## **1. Introduction**

The genus *Capsicum* includes many cultivated species, of which *Capsicum annuum* L. is one of the most widely cultivated one; besides this, other domesticated species are *C. baccatum*, *C. chinensis*, *C. frutescens*, and *C. pubescens* [1]. *C. annuum* comprises of both sweet (bell pepper) and pungent (chilli) fruits of numerous shapes and sizes. It is a good source of Vitamin A and C, potassium, and folic acid [2]. Fresh green chilli has more vitamin C than a citrus fruit, whereas red chilli has more vitamin A than in carrots [3, 4]. Besides its wide use as vegetable, spice, and condiments, it is also used in medicines and beverages. Capsaicinoid and caretenoids are the active ingredients of the chilli; the capsaicinoids are nonvolatile alkaloids that make chilli pungent [5], and caretenoids have nutritional value that also provides color to the chilli fruit [6]. In tropical and subtropical countries, chilli is considered the most important constituent of different cuisines. As the native home of chillies are considered to be tropical America, where it is still found growing in the wild state [7]. Its introduction to India is credited to voyage of Columbus who brought seeds from Spain, introducing it to Europe, which subsequently spread to Africa and Asia [8].

India is the world's largest producer of dried chillies and in 2018 India produced 1.8 million tons, out of 4.1 million tons produced worldwide [9]. There are two important commercial qualities that makes Indian chilli world famous are color and pungency levels. Chilli crop is attacked with different pests and pathogens in field and during post-harvest, contamination with mycotoxins are major constraints in chilli production. Worldwide, *Capsicum* is vulnerable to various pests, weeds, fungal, bacterial, and viral pathogens; among the fungal diseases, anthracnose/ die-back/fruit-rot of chillies is an important disease causing serious losses in field, transit, transport, and storage [10, 11].


**5. Final remarks and future directions**

*Neurostimulation and Neuromodulation in Contemporary Therapeutic Practice*

*Preoperative (a) and postoperative (b, at 7 years) magnetic resonance images of a 49-year-old female patient with Chiari malformation type I, operated of* filum terminale *sectioning, with impressive improvement of the tonsillar descent from 17 to 13 mm with respect to McRae's line (measured in the cuts with maximal descent).*

and more clinicians are starting to accept that maybe suboccipital craniectomy is not the only surgical solution to this condition. For this reason, we can present here three "surgical" testimonies in favor of the theory of caudal traction, as follows:

1.In fact, it is suboccipital craniectomy itself that opened these new perspectives when in the hands of some fearless teams [23, 24]; it started to be used for treating patients with syringomyelia without tonsillar descent, with encouraging results, but they did not realize their meaning not even when they discovered that in these children, there were image features suggesting a caudal elongation of the brainstem with displacement of the obex and increased diameter of the *foramen magnum*, as occurs also in Chiari malformation type II [24]. In **Figure 6** we show a similar example from our series of patients, where sectioning the *filum terminale* determined a marked improvement of a cervicothoracic syringomyelia in a case of Chiari malformation type 0.

2.Another ingenious team discovered that if they performed idiopathic scoliosis correction by a technique of posterior vertebral column resection with spine shortening and instrumentation after applying compressive forces, the cerebrospinal fluid flow at the level of the *foramen magnum* improved in patients with Chiari malformation type I and syringomyelia, and in many of them, even the latter diminished in size [25]. Obviously, scoliosis surgery by no means could have accomplished any *circumferential decompression* of the occipital foramen but a release of the tension in the brainstem and tonsils (or, as we

3.Yet the most important testimony came from the hands of a group which operated 318 patients presenting both tethered cord syndromes defined according to very exigent criteria and Chiari malformation type I or low-lying cerebellar tonsils of 0–4 mm descent (that we consider as being also Chiari malformation type I, together with more and more authors [1]), but the technique used was not suboccipital craniectomy, but sectioning of the *filum terminale* by means of an L4 laminectomy. Their results were excellent, both concerning the clinical picture and various morphometric criteria of the posterior fossa contents,

stated above, a welcome *longitudinal decompression*) (**Figure 7**).

**256**

**Figure 8.**

In the actual state of knowledge, it is imperative to recognize that the development of the hindbrain and the spinal cord is a complex process regulated by genetic, molecular, mechanical, endocrine, and nervous homeostatic mechanisms that compensate one for another—within certain limits—in case of imbalances and disturbances. Nevertheless, it is exactly this complexity, coupled with the elevated functional requirements that the cranio-cervical junction has to meet, that makes their union so sensitive to various pathogenetic factors and determines malformations among which the one known as Chiari malformation type I is the most common. According to all the arguments presented in this chapter, the final common pathway of these etiopathogenetic aggressions seems to be caudal traction, a complex biological phenomenon that by no means should be reduced to a simple mechanical force of axial pull. There is still much left to discover about the physiologic mechanisms that govern the coupling between the growth of the vertebral column and that of the spinal cord during somatic development, where maybe future research will define the roles played by the pineal gland, the subcommissural organ, and the *filum terminale*, just to cite a few of the possible actors eligible for this casting.


*2.2. Experimental Part*

There are two major categories, according to which the dielectric measurements regarding any High Voltage (HV) equipment are classified: the destructive and non-destructive tests. In the context of the destructive tests, AC, positive and negative LI BDV are measured for the two NF samples along with a base oil sample. In regard to the non-destructive ones, PDIV, as well as the variation of the apparent charge for various voltage levels, have been measured.

AC BDV is the fundamental parameter for the characterization of any insulation system, as long the transformer nominally operates under AC voltage. For the determination of this property, each one of the samples under investigation is stressed under AC voltage/60 Hz using a Baur DTA 100 C generator, as illustrated in Figure 2a, according to the conditions of IEC 60,156 standard. The generator can produce up to 100 kV increasing the voltage at a rate of 2 kV/s until breakdown occurs between two Rogowski electrodes, which ensure a uniform electric field distribution, with the gap distance set at 2.5 mm. At the beginning of the experimental procedure, the electrodes are polished and cleaned. 10 sets of 6 breakdown events (60 in total) are performed, with the applied interval being 2 min between two successive breakdowns and 5 min between two sets. The results are fit to Weibull and normal distributions in order the AC BDV in low probability levels to be estimated [7,23,34].

**Figure 2.** AC BDV and PD apparatuses: (**a**) BAUR DTA test cell with Rogowski electrodes. (**b**) Schematic diagram of the PD setup.

Regarding the measurement of LI BDV, this is another property of major importance as it is an indication for the strength of the insulation in emergency conditions, such as lightning strikes. A two-stage impulse generator is used, producing up to 400 kV, with positive and negative polarity, according to IEC 60,897 standard. A point–sphere geometry is used, ensuring a highly divergent field geometry, with the gap distance set at 25 mm. The radius of curvature is 50 μm, while the diameter of the sphere is 12.7 mm. Five voltage applications are conducted for each voltage level with each sample being replaced with a new one after every BD event, ensuring a good reliability for the processing of the results. The applied lightning impulse voltage 1.2/50 μs, is monitored on oscilloscope Tektronix DPO4104; 1 GHz/5 GS s-1.

In terms of the conduction of PD measurements, a HV test transformer (HIGH VOLT GmbH Transformer PEOI 40/100,100 kV) is used with a TETTEX Instruments PD Detector DDX-9101 to measure the apparent charge to a nominal capacitor (HIPOTRONICS capacitor PSF 100-1). The schematic diagram of the configuration is displayed in Figure 2b. The test cell consists of two plate electrodes with the gap between them set at 0.75 mm. Nomex® Dupont™ insulating paper with 0.75 mm thickness impregnated to the NFs and base oil samples, is put in the gap. The AC voltage value for which the measured apparent charge exceeds the value of 10 pC is considered as the PDIV for each sample under investigation. The insulating papers impregnated to iNF, sNF, and base oil samples are stressed under AC voltage while the transformer is increasing the voltage at a rate of 0.1 kV/s until the measured apparent charge exceeds the threshold value. Each experiment is repeated 4 times for ensuring the reliability of the obtained results.


**3.1 The diagnosis of CO intoxication**

The diagnosis of CO intoxication is based on the clinical history of exposure or elevated carboxyhemoglobin level (> 10%) (Handa and Tai 2005; Chang, Lee et al. 2009). There is currently no definition of clinical staging in CO intoxication in the literature, although the pathophysiology follows that of hypoxic–ischemic encephalopathy (Gutierrez, Rovira et al.).

Neuroimaging Studies in Carbon Monoxide Intoxication 357

prevalence differs among studies (Silver, Cross et al. 1996; O'Donnell, Buxton et al. 2000). One study showed 63% of abnormal lesions in the GP with 26% in the rest of the basal ganglia (O'Donnell, Buxton et al. 2000). Another study with 73 patients revealed only one patient (1.4%) with basal ganglia lesions scanned two weeks after CO poisoning (Parkinson,

Low density GP lesions, commonly seen in computed tomography (CT), are considered as characteristic findings in patients with CO intoxication (Kanaya, Imaizumi et al. 1992; Gotoh, Kuyama et al. 1993; Uchino, Hasuo et al. 1994; Chu, Jung et al. 2004; Kinoshita, Sugihara et al. 2005; Hopkins, Fearing et al. 2006). Low density lesions of the putamen and caudate nucleus, in contrast, have only been reported in one case (Ferrier, Wallace et al. 1994). The nature of GP lesions has been studied further by diffusion-weighted imaging (DWI) and apparent diffusion coefficient (ADC) mapping (Chu, Jung et al. 2004; Kinoshita, Sugihara et al. 2005). One case report interpreted low ADC values and high intensity GP lesions on DWI as restriction of water diffusion (i.e. cytotoxic edema) (Kinoshita, Sugihara et al. 2005). Vasogenic edema can also be visualized on ADC and DWI as increased signal intensity lesions (Chalela, Wolf et al.

Fig. 1. Magnetic resonance imaging study in the acute stage of carbon monoxide intoxication. Six days after CO intoxication, a 42-year-old woman with a globus pallidus interna lesion with hyperintensity in diffusion weighted imaging (1A), hypointensity in apparent diffusion coefficient (1B), hypointensity in T1 weighted image (WI) (1C), hyperintensity in T2WI (1D),

Imaging studies showing cavity-changes by T1 or T2WI often suggest necrosis of the GP (Mendelsohn and Hertzanu 1983; Pulst, Walshe et al. 1983; Ko, Ahn et al. 2004). Autopsies of patients with CO intoxication have confirmed the histology of necrosis and/or neuronal degeneration of the GP (Jones, Lagasse et al. 1994). The pathogenesis of necrosis is believed to be due to edema-induced ischemia or hemorrhage transformation (Chang, Han et al.

**4.1.1 Imaging features suggesting edematous change in the acute phase**

2001). The high signal on DWI is due to the T2 shine-through effect.

and hyperintensity in fluid-attenuated inversion recovery (1E).

**4.1.2 Imaging features suggesting necrosis**

Hopkins et al. 2002).

### **3.2 Symptoms in the acute phase**

Tightness across the forehead, headache, throbbing in the temples, nausea, vomiting, dimness of vision, dizziness, general weakness, syncope, convulsion, and coma are commonly found in patients with CO exposure within one day (Choi 2001). Cortical blindness with initially normal visual evoked potentials has also been reported in a case (Katafuchi, Nishimi et al. 1985). The pathogenesis contributing to the clinical manifestations includes change of blood flow (Penney 1990; Lo, Chen et al. 2007), hypoxia (Lo, Chen et al. 2007), and neurochemistry abnormalities (Penney 1990).


**4.3. Classification of Glucose-6-Phosphate Dehydrogenase variants**

More than 400 variants of G6PD have been distinguished based on their biochemical characteristics, enzyme kinetics, physicochemical characteristics, and other parameters (Luzzatto & Battistuzzi, 1985, Chen*, et al.*, 1991, Greene, 1993). G6PD B+ is the most commonly found enzyme type and it is used as a standard for normal enzyme activity and electrophoretic mobility. For identification of other variants, G6PD B+ is used. The rate at which NADP+ is reduced by glucose-6-phosphate with G6PD B+ as the catalyst is the standard for activity. Based on this, enzyme activity relative to G6PD B+ variants are classified as fast, normal, and slow in terms of electrophoretic mobility and as Classes I—V (Luzzatto, 1989, Beutler, 1990, Greene, 1993, Segel, 2000, Betke K, Brewer GJ, Kirkman HN, Luzzatto L,Motulsky AC, Ramot B, and Siniscalco M 1967). There are 5 classes for these variants. Class I includes chronic nonspherocytic hemolytic anemia with a severe enzyme deficiency (e.g., G6PD Minnesota, G6PD Tokyo, G6PD Campinas). Class II variants have severe enzyme deficiency without chronic nonspherocytic hemolytic anemia (e.g., G6PD mediterrian, G6PD Canton, G6PD Union, G6PD Kaiping). Class III variants includes medium or mild enzyme deficiency, with the activity at 10-60% of G6PD B+ (e.g., G6PD Aˉ). Class IV variants have a weak or no enzyme deficiency. The activity is 60-100 % of G6PD B+ (e.g., G6PD A+). Class V variants have increased enzyme activity (e.g., G6PD Hektoen) (Beutler, 1994, Segel, 2000).


**2. Chemical enhancers**

Chemical percutaneous enhancers have long been used to increase the range of drugs that can be effectively delivered through the skin (López-Castellano & Merino, 2010). To date, a plethora of chemicals have been evaluated as enhancers, but their inclusion in topical or transdermal formulations is limited due to fact that the underlying mechanisms of action of these agents remain unclear. Although different chemicals are employed by the industry as percutaneous enhancers, some of which have several desirable properties, to date none has proved to be ideal. An ideal chemical penetration enhancer should have the following attributes (Barry, 1983; López- Castellano & Merino, 2010): a) It should be non-toxic, nonirritating and non-allergenic, b) It should work rapidly, and its activity and duration of effect should be both predictable and reproducible, c) It should exert no pharmacological activity within the body, d) It should work unidirectionally, e) When removed, the skin's barrier properties should return both rapidly and fully, f) It should be compatible with both excipients and drugs, and g) It should be cosmetically acceptable and, ideally, odourless and colourless.

#### **2.1 Percutaneous penetration routes of drugs**

There are three major potential routes of percutaneous penetration: appendageal, transcellular (through the stratum corneum), and intercellular (through the stratum corneum) (Figure 1). There is a weight of evidence that suggests that passage through the intact stratum corneum constititutes the predominant route by which most molecules penetrate the skin, as the appendageal route is characterized by a limited available fractional

Fig. 1. Processes of percutaneous absorption

Chemical percutaneous enhancers have long been used to increase the range of drugs that can be effectively delivered through the skin (López-Castellano & Merino, 2010). To date, a plethora of chemicals have been evaluated as enhancers, but their inclusion in topical or transdermal formulations is limited due to fact that the underlying mechanisms of action of these agents remain unclear. Although different chemicals are employed by the industry as percutaneous enhancers, some of which have several desirable properties, to date none has proved to be ideal. An ideal chemical penetration enhancer should have the following attributes (Barry, 1983; López- Castellano & Merino, 2010): a) It should be non-toxic, nonirritating and non-allergenic, b) It should work rapidly, and its activity and duration of effect should be both predictable and reproducible, c) It should exert no pharmacological activity within the body, d) It should work unidirectionally, e) When removed, the skin's barrier properties should return both rapidly and fully, f) It should be compatible with both excipients and drugs, and g) It should be cosmetically acceptable and, ideally, odourless and colourless.

There are three major potential routes of percutaneous penetration: appendageal, transcellular (through the stratum corneum), and intercellular (through the stratum corneum) (Figure 1). There is a weight of evidence that suggests that passage through the intact stratum corneum constititutes the predominant route by which most molecules penetrate the skin, as the appendageal route is characterized by a limited available fractional

**2. Chemical enhancers**

**2.1 Percutaneous penetration routes of drugs**

Fig. 1. Processes of percutaneous absorption

area of 0.1%. In this way, diffusion through the skin is controlled by the particular characteristics of the stratum corneum. In order to obtain a sufficient drug flux and, in turn, the therapeutical objectives in question, an alternative is to use chemical percutaneous enhancers. These substances alter some of the properties of the stratum corneum. (López-Castellano & Merino, 2010)

#### **2.2 Direct effects on the skin due to the use of transdermal penetration enhancers**

The lipid–protein-partititioning theory sets out the mechanisms by which enhancers alter skin lipids, proteins and/or partitioning behaviour (Barry, 1991): i) They act on the stratum corneum intracellular keratin by denaturing it or modifying its conformation, causing subsequent swelling and increased hydration; ii) They affect the desmosomes that maintain cohesion among corneocytes; iii) They modify the intercellular lipid domains to reduce the barrier-like resistance of the bilayer lipids. Disruption to the lipid bilayers can be homogeneous when the enhancer is distributed evenly within the complex bilayer lipids, but the accelerant is more likely to be heterogeneously concentrated within the domains of the bilayer lipids and iv) They alter the solvent nature of the stratum corneum, thus aiding the partitioning of the drug or a co-solvent into the tissue.(López-Castellano & Merino, 2010)

#### **2.3 Indirect effects on the skin due to the use of transdermal penetration enhancers**

Chemical enhancers can produce: *a)* Modification of the thermodynamic activity of the vehicle. The permeation of a good solvent from the formulation, such as ethanol, can increase the thermodynamic activity of a drug; *b)* It has been suggested that, by permeating through the membrane, a solvent can 'drag' the permeant with it, though this concept is somewhat controversial and requires confirmation; *c)* Solubilising the permeant within the donor, especially when solubility is very low, as in the case of aqueous donor solutions, can reduce depletion effects and prolong drug permeation.(López-Castellano & Merino, 2010)

#### **2.4 Classification of percutaneous chemical enhancers**

The classification of percutaneous enhancers is frequently based on the chemical class to which the compounds belong. Table 1 shows the principal classes of percutaneous enhancers.

Table 1. Principal classes of percutaneous enhancers.

Chemical and Physical Enhancers for Transdermal Drug Delivery 401

Absorption of ultrasonic energy leads to tissue heating, and this has been used with therapeutic intent in many conditions. More recently it has been realized that benefit may also be obtained from the non-thermal effects that occur as ULTS travels through tissue. ULTS therapies can broadly be divided into ''high'' power and ''low'' power therapies where high power applications include high intensity focused ULTS and lithotripsy, and low power encompasses sonophoresis, sonoporation, gene therapy and bone healing. There are three distinct sets of ULTS conditions based on frequency range and applications: 1) High frequency (3–10 MHz) or diagnostic ULTS, 2) Medium frequency (0.7–3 MHz) or

The term ultrasonic refers to sound waves whose frequency is >20 KHz. The intensity (I, expressed in W/cm2), or concentration of power within a specific area in an ULTS beam, is proportional to the square of the amplitude, p, which is the maximum increase or decrease in the pressure relative to ambient conditions in the absence of the sound wave. The complete relationship is: I= p2/2ρc, where ρ is the density of the medium and c is the speed of the sound (in human soft tissue, this velocity is 1540 m/s). The intensity is progressively lost when a sound wave passes through the body or is deviated from its initial direction, a phenomenon referred to as attenuation. In homogeneous tissue, the attenuation occurs as a result of absorption, in which case the sound energy is transformed into heat and scattered. The sound waves are produced in response to an electrical impulse in the piezoelectric crystal, allowing the conversion of electrical into mechanical or vibrational energy; this transformation requires a molecular medium (solid, liquid, or gas) to be effective. The ULTS beam is composed of two fields, the ''near field,'' in the region closest to the transducer face, and the ''far field,'' corresponding to the conical diverging portion of the beam (Figure 2). The parameters controlling this configuration of the ULTS beam are principally the

Cavitation is the formation of gaseous cavities in a medium upon ULTS exposure. The primary cause of cavitation is ULTS-induced pressure variation in the medium. Cavitation involves both the rapid growth and collapse of a bubble (inertial cavitation), or the slow oscillatory motion of a bubble in an ULTS field (stable cavitation). Collapse of cavitation bubbles releases a shock wave that can cause structural alteration in the surrounding tissue (Clarke et al., 2004) ULTS can generate violent microstreams, which increase the bioavailability of the drugs (Tachibana & Tachibana, 1999). Tissues contain air pockets that are trapped in the fibrous structures that act as nuclei for cavitation upon ultrasound exposure. The cavitational effects vary inversely with ULTS frequency and directly with ULTS intensity. Cavitation might be important when low-frequency ULTS is used, gassy fluids are exposed or when small gas-filled spaces are exposed. Cavitation occurs due to the nucleation of small gaseous cavities during the negative pressure cycles of ULTS, followed by the growth of these bubbles throughout subsequent pressure cycles (Tang et al., 2001).

therapeutic ULTS, and 3) Low frequency (18 to 100 KHz) or power ULTS.

**3. Sonophoresis**

**3.1 The ultrasound**

frequency and the size of transducer.

**3.2 Mechanisms of action 3.2.1 Cavitation effects**

#### **2.5 Determination of permeation enhancement**

The great majority of studies of the effects of enhancers on skin permeability have been carried out by means of *in vitro* diffusion experiments in which various kinds of diffusion cells have been used. The most well-known of these cells are the Franz diffusion systems. These cells have two receptor compartments - donor and receptor (donor positioned above receptor) – between which the skin is placed. In general, the skin is pretreated with a solution of the chemical enhancer to be evaluated. The transdermal flux (J) of drugs can be estimated from the slope of the linear region (steady-state portion) of the accumulated amount of drug in the receptor compartment versus time plot. Permeation enhancing activity, expressed as enhancement ratio of flux (ERflux), is determined as the ratio between the flux value obtained with the chemical enhancer and that obtained with the control. A number of variables can strongly influence the permeation enhancement of drugs. The most important are the skin used in the experiments, temperature, humidity, enhancer concentration, vehicle employed and degree of saturation of the drug in the donor and receptor compartments. (López-Castellano & Merino, 2010)

#### **2.6 Uses in topical/transdermal formulations**

Some examples of drugs delivered throughout the skin using chemical enhancer are shown in Table 2.

Table 2. Examples of drugs delivered throughout the skin using chemical penetration enhancers.