182

APPENDIX A1

Glossary of terms used in biodeterioration Autotroph

An organism that uses carbon dioxid e for its carbon requirement; often used to imply lithoautotrophy , as obligate autotrophs appear to be either chemolithotrophs or photolithotrophs.

Biodeteriogen

An organism or microorganism that causes undesirable changes in the constituent materials of (esp. cultural) objects.

Biofilm

A film of microorganisms, usuall y embedded in extracellular polymers that adhere to surfaces submerged in, or subjected to, aquatic environments.

Chelate A chemical agent that combines with metal ions

Chelation

A chemical process involving the formation of heterocyclic ring compounds containing at least one metal cation or hydrogen ion.

Chemoautotroph

An organism whose energy is derived from endogenous, light-independent chemical reactions (a mode of metabolism termed chemotrophy); a chemotroph that obtains energy by the oxidation of inorgani c substrate(s ) (i.e. , a lithotroph , called chemolithotroph); one that obtains energy by metabolizin g organic substrate(s) (i.e., an organotroph, called a chemoorganotroph).

Chlorophytes

A group of algae characterized by a combination of features (e.g., containing chlorophylls a and b, ß-carotene, and xanthophyll s such as antheraxanthin, lutein , neoxanthin, violax - anthin, and zeaxanthin).

chloroplasts Organisms evolved by a double membrane; the main storage polymer is starch, which is formed within the chloroplast . Motile cells, when formed, bear flagella.

endolithic Growing within rocks, as some algae, lichens, and bacteria do in limestone.

epilithic Growing on a rock surface.

eukaryotes

A group of organisms ; literally those having "nuclei" in their cell s (i.e. , animals , plants , and fungi) in contrast to the prokaryotic cell s of bacteria and blue-green algae; possessing a nucleus bounded by a membranous nuclear envelope and having many cytoplasmic organelles.

183

foliose Leafy, having a body differentiated into stem and leaves; of lichen: having a flattened leaf like thallus more or less firmly attached to the substratum.

fruticose Shrublike; of a lichen: having a thallus that is threadlike (terete) or straplike (more or less flattened), and either erect and shrubby or pendulous , the thallus being attached to the substratum by a holdfast or unattached.

gymnosperms

A group (classified as a division Gymnospermophyta or a class Gymnospermae, or regarded as polyphyletic) containing those seed plants in which the ovules are not enclosed in carpels , the pollen typically germinating on the surface of the ovule.

heterotroph

An organism that uses organic compounds for most or all of its carbon requirements; often used to refer specifically to chemoorganoheterotrophs, although chemolithotrophs and phototrophs may also be heterotrophic.

hyphae Branched or unbranched filaments, many of which together constitute the vegetative form of the organism and (in some species) form the sterile portion of a fruiting body.

metabolite Any of various compounds produced by metabolism.

photoautotroph An autotroph that requires sunlight to provide necessary energy for biosynthesis.

photoheterotroph A heterotroph that requires sunlight to provide necessary energy for biosynthesis

photolithotroph An organism that uses inorganic substrate s (e.g., water, sulfur, sulfide, H2

photoorganotroph

, or thiosulphate) as an electron donor in photosynthesis. An organism that uses an organic substrate as an electron donor in phototrophic metabolism.

prokaryote

A type of microorganism in which the chromosomes are not separated from the cytoplasm by a specialized membrane that is typically devoid of sterol s and in which mitochondria and chloroplast s are absent.

redox reaction

A chemical reaction in which electrons are transferred between atoms, ions, or molecules.

rhizoid

A root like structure consisting of a compact bundle of hyphae; arising mainly from the lower surface of the (usually foliose) thallus and serving to anchor the thallus to the substratum; in some species, may also facilitate the uptake of water and nutrients by thallus.

184

taxonomy

The science of classification as applied to living organisms, including the study of the means of specie s formation

thallus

Plant body not differentiated into leaves, stems, and roots, but consisting of a mycelium or a colony, a filament of cells , a mycelium, or a large branching multicellular structure; the plant body of the algae, fungi, and liverworts.

Microbiologically Influenced Deterioration (MID)

Miro-organisms degrading/ deteriorating non-metals

185

APPENDIX A2

OVERVIEWOF SEVERAL TECHNIQUES USED FOR MATERIALS CHARACTERIZATION AND SURFACE ANALYSIS

Scanning electron microscopy, environmental scanning electron microscopy and energy

dispersion X-ray analysis:

SEM allows the observation and surface characterization of inorganic and organic materials

at a higher resolution and depth of field than that of the conventional optical microscope. In

the SEM analysis, a thermo-ionic emission of an electron beam is directed over the sample to

produce characteristic signals that provide information on crystalline array, chemical

composition, magnetic structure and even on the electric potential of the specimen under

observation. SEM offers a high resolution (approximately 100 Å) and high field depth giving

a 3D effect to the images. Contrary to SEM, ESEM does not operate at high vacuum and

provides fast, accurate images of biological deposits (biofilms), showing their spatial

relationship and surface chemistry without needing extensive manipulation of the samples.

This instrument has a unique secondary electron detector capable of forming high resolution

images at pressures in the range of 0.1–20 torr. At these relatively high pressures, non-

conductive biological specimens are directly observed without metallic coating and any

special preparation. If water vapor is used in the chamber, wet samples can be observed

directly and EDX data can be collected at the same time as sample morphology and

topography are photographed.

In the EDX technique, a solid-state detector is placed close to the sample to emit an electric

pulse when the interaction with the X-rays is produced. The intensity of the electric pulse is

proportional to the energy of the X-ray beam. If the number of pulses and the intensity of the

X-rays are measured, the chemical composition of the sample can be determined. The

wavelength of the radiation is analyzed by a dispersion device which diffracts the

characteristic radiation of the sample by means of a suitable crystal.

X-ray diffraction analysis

XRD is a convenient and useful tool for the identification of crystalline compounds as well as

for assessing the structure of complex natural compounds. The theoretical basis of the method

is the fact that X-ray diffraction patterns are unique for each crystalline structure. When the

X-ray beam passes trough the material, a large number of the particles can be expected to be

aligned in such directions that they can fulfill the Bragg relationship for reflection for each

possible inter-planar spacing.

186

Petrographic analysis

The polarizingmicroscope (also called petrographicmicroscope) is often used to examine thin

sections of constructional materials and it is very useful in the study of rock transformation

due to weathering effects.

Mossbauer spectroscopy

Usually the energy of a beam of γ rays emitted by an atomic nucleus which changes from an

excited level to the ground state is different from the transition energy. Under certain

experimental conditions a fraction of the γ rays beam has a similar energy to the transition

energy (Mossbauer effect). This effect depends on the firmness of the link between the

emitting nucleus and its chemical environment. In the corrosion field themost interesting

nuclei which present the Mossbauer effect are those of Fe, Ni and Zn. It is also used in

corrosion studies where it provides the complete, in situ measurement, non-destructive, three-

dimensional identification of corrosion products and is the only technique able to accurately

measure the fraction of each oxide in a corrosion product layer. Thus, it can be used to study

the electric, magnetic and structural characteristics of metals, alloys, soils andminerals. MS

can be considered as a ‘‘fingerprint’’ technique of varying degrees of sophistication in

mineralogical and geo-chemical studies.

Grazing incident diffraction

GID is a valuable technique that provides very precise information on surface and interface of

atomic arrangement in crystalline structures. By varying the angle of incidence, the

penetration of the X-ray beam into the material can be controlled allowing the analysis of

surfaces and buried interfaces. GID is a powerful tool for surface structural characterization of

properties such as atomic structure and roughness, relaxation and reconstruction. Non-

destructive analysis is a representative application of the GID. The incidence angle was varied

in order to change the penetration depth between few nanometers up to several micrometers.

GID has become well established for the investigation of the structure of films, surfaces and

interfaces.

X-ray photoelectron spectroscopy and reflection electron energy-loss spectroscopy

XPS and REELS are powerful surface-sensitive analytical techniques that are well suited for

the analysis of materials. XPS analysis of the external surface provides valuable information

about the elemental concentrations at the surface and the chemical state. The surface

composition, at the limits of the penetration depth of this

187

technique (w5 nm), can be derived from the intensity of the different photoemission peaks

detected in the general spectra corrected by their respective sensitivity factors.

Raman spectroscopy

The Raman spectroscopy has become increasingly available in the field of cultural heritage

studies. During the last decade, it has gradually become an established technique for the non-

destructive identification and study of ancient materials.

DNA-based molecular techniques

DNA-based molecular techniques allow the direct detection and identification of single

microorganisms in environmental samples. These techniques have been applied to detect

microorganisms in many natural environments since the beginning of the 90s and are also

beginning to be used in biocorrosion studies (corrosion of metallic structures due to

microorganisms). Molecular techniques have been used to characterize the microorganisms

present in biofilms damaging cultural heritage buildings.

These techniques are based on the analysis of ribosomal DNA sequences that are present in

the genome of all living organisms. These sequences can be used as biomarkers for the

accurate identification of microorganisms considering that different microbial species will

have different ribosomal gene sequences (Woese, 1987). Ribosomal genes contain both highly

conserved and highly variable regions where ‘‘signature sequences’’ have been identified for

each domain of life (Bacteria, Eucarya and Archaea) as well as for microbial groups, families,

genera and species. By knowing their ribosomal sequences, the microorganisms there can be

classified and identified.

188

APPENDIX A3

CLASSIFICATION OF ALGAE (V.J. Chapman, 1941)

In the older version of classifications the algae proper were simply divided into four principal

groups, Chlorophyceae or green algae, Cyanophyceae or blue-green algae, Phaeophyceae or

brown algae and Rhodophyceae or red algae. Now, that more is known about the simpler

organisms which were not to be regarded as algae, it has been realized that there is no real

justification for such a distinction, and so the number of algal groups has been increased. This

is because it has become evident that the Flagellata and other simple unicellular organisms

must properly be regarded as algae, even though they are of a very primitive kind. At present

it is most convenient to divide the algae into ten classes, one of which, the Nematophyceae, is

perhaps somewhat speculative. One of the principal bases of this classification is the

difference in pigmentation, and a recent study of this problem shows that it is fully justified.

(1) Cyanophyceae The plants in this group show very little evidence of differentiation,

containing only a very simple form of nuclear material, no proper chromatophore and no

motile cells with cilia or flagellae. The products of photosynthesis are sugars and glycogen.

The colour of the cells is commonly blue-green and hence their name, the colour being due to

the varying proportions of the pigments phycocyanin and phycoerythrin. There is no known

sexual reproduction, propagation taking place by simple division or else by vegetative means.

(2) Chlorophyceae This group used to comprise four great subdivisions, the Isokontae (equal

cilia), Stephanokontae (ringed cilia), Akontae (no cilia) and Heterokontae (unlike cilia). It is

now more in keeping with our present knowledge to place the last section into a separate

class, and this is the procedure adopted in most recent books. The plants of the Chlorophyceae

exhibit a great range of structure from simple unicells to plants with a relatively complex

organization, whilst the chromatophores also vary considerably in shape and size. The final

product of photosynthesis is starch together with oil, and a starch sheath can often be

demonstrated around the pyrenoids. In the bulk of the members of this class the motile cells

are very similar and commonly possesseither two or four flagellae, but in the Oedogoniales

(Stephanokontae) there is a ring of flagellae whilst in the Conjugales (Akontae) there are no

organs of propulsion. Sexual reproduction is of common occurrence and ranges from isogamy

to anisogamy and oogamy. The colour of the cells is usually a grass green because the

pigments are the same as those present in the higher plants and, furthermore, they are present

in much the same proportions.

189

(3) Xanthophyceae (Heterokontae) The plants in this group are usually of a simple nature,

but their lines of development frequently show an interesting parallel or homoplasy with those

observed in the preceding group. The chloroplast is yellow-green owing to an excess of

xanthophyll, one of the four normal constituents of chlorophyll. Oil replaces starch as the

normal storage material, the lack of starch being correlated with the absence or paucity of

pyrenoids. The motile cells possess two unequal flagellae (occasionally only one) arising from

the anterior end. Sexual reproduction is rare and when present is isogamous. The cell wall is

frequently composed of two equal or unequal halves overlapping one another.

(4) Chrysophyceae These form another very primitive group in which the brown or orange

colour of the chloroplasts is determined by the presence of accessory pigments such as

phycochrysin.Most of the forms have no cell wall and hence are “flagellates” in the old sense

of that term, although there are some members which do possess a cell wall and hence are

"algal" in the old sense of the term. Fat and leucosin (a protein-like substance) are the usual

forms of food storage, whilst another marked feature is the silicified cysts which generally

have a small aperture that is closed by a special plug. The motile cells possess one, two or,

more rarely, three equal flagellae attached at the front end, but in one subsection the paired

flagellae are unequal in length. The most advanced habit known is that of a branched filament,

e.g. Phaeothamnion, whilst the palmelloid types attain to a much higher state of

differentiation, e.g. Hydrurus, than in either the Chlorophyceae or the Xanthophyceae. Sexual

reproduction is not certain, and such records as there are point simply to isogamy.

(5) Bacillariophyceae (Diatoms) One of the characteristics of these plants is their cell walls

which are composed partly of silica and partly of pectic material. The wall is always in two

halves and frequently ornamented with delicate markings, which are so fine that microscope

manufacturers make use of them in order to determine the resolving power of their lenses.

The chromatophores are yellow or golden brown containing, in addition to the usual

pigments, accessory brown colouring materials whose nature is only just being established.

One set of forms is radially symmetrical, the other bilaterally so. The presence of flagellate

stages is highly probable in the former whilst there is a special type of sexual fusion in the

latter group.

(6) Cryptophyceae There are usually two large parietal chloroplasts with diverse colours,

though frequently of a brown shade, whilst the product of photosynthesis is starch or a closely

related compound. The motile cells have two unequal flagellae and often possess a complex

vacuolar system. Nearly all the members have a "flagellate” organization and there is no

190

example of the filamentous habit. One type, however, has been described with a tendency

towards the coccoid (non-motile unicell with a cell wall) habit, and so this must be regarded

as the least "algal "-like class. Isogamy has been recorded for one species.

(7) Dinophyceae Most of the members of this class are motile unicells, but there has been an

evolutionary tendency towards a sedentary existence and the development of short algal

filaments, e.g. Dinothrix. Many are surrounded by an elaborate cellulose wall bearing

sculptured plates and inside there are discoid chromatophores, dark yellow or brown in colour

and containing a number of special pigments. The products of photosynthesis are starch and

fat. The motile cells normally possess two furrows, one transverse and one longitudinal,

although they may be absent in some of the more primitive members. The transverse

flagellum lies in the former, and the latter is the starting point for the other flagellum which

points backwards. Sexual reproduction, if it occurs, is isogamous, and it has not been clearly

established in the few cases reported. Characteristic resting cysts are also produced by many

of the forms.

(8) Phaeophyceae This group comprises the common brown algae of the seashore and it is

worth noting that the majority are wholly confined to the sea. The brown colour is due to the

presence of a pigment, fucoxanthin, which masks those other chlorophyll constituents which

are present. The products of photosynthesis are alcohols, fats, polysaccharides and traces of

simple sugars so that there is evidence of some diversity of metabolism. The simplest forms

are filamentous, and there are all stages of development and increasing differentiation up to

the large seaweeds of the Pacific and Arctic shores with their great size and complex internal

and external differentiation. The motile reproductive cells, which possess two flagellae, one

directed forwards and the other backwards, are commonly produced in special organs or

sporangia that are either uni or plurilocular. Sexual reproduction ranges from isogamy to

oogamy, but in the latter case the ovum is normally liberated before fertilization. The life

cycles may be extremely diverse and are perhaps better regarded as race cycles.

(9) Rhodophyceae The members of this class form the red seaweeds, and although most of

them are marine nevertheless a few are fresh-water. Their colour, red or bluish, is caused by

the presence of the pigments phycoerythrin and phycocyanin, whilst the product of

photosynthesis is a material known as "floridean starch". Reproductive stages with locomotor

appendages are not known, even the male reproductive body being without any organ of

locomotion. The simplest members are filamentous, and again all stages of differentiation up

to a complex body can be found, although they do not develop to quite the same degree of

191

complexity as the Phaeophyceae. Very obvious protoplasmic connexions can be distinguished

between the cells of nearly all forms except in the small group known as the Proto-florideae.

Sexual reproduction is oogamous, the ovum being retained upon the parent plant, and

although the subsequent development of the zygote is varied to a certain extent, it usually

gives rise to filaments which bear special reproductive bodies or carpospores, and these latter

are responsible for the production of the tetrasporic diploid plant. Most of the members

exhibit a regular alternation of generations.

(10) Nematophyceae This is a fossil group of which one genus has been known for a long

time (Nematophyton) whilst the other has only recently been described (Nematothallus).

There is still considerable doubt as to their true affinities, but it would seem that a place can

best be found for them as a very highly developed type of alga. Their internal morphology

would ally them closely with the more advanced members of either the Chlorophyceae or the

Phaeophyceae. The only reproduction so far recorded is that of spores which were developed

in tetrads, and therefore may have been akin to the Rhodophycean or Phaeophycean

tetraspores.

Based on the location at which algae colonizes, another classification that is generally

recognized is: (i) marine algae – widely available near sea coasts /sea; (ii) terrestrial algae -

widely available on moist surface in inland regions; (iii) fresh water algae – widely available

on surface of water bodies.

192

APPENDIX B1

Characterization of Three Species of Algae

1. Enteromorpha clathrata (Roth) Plants less than 15cm long, soft, with repeatedly branched

thallus Fig. B2.1. Frequently uniseriate, branchlets in narrow. The cells are more or less

quadrangular in longitudinal and even in transversal rows.

2 Ulva fasciata (Delile) Plants 1-15 cm. tall, the base of the blade cuneate, above expanding

irregularly lobed, generally irregularly or sometimes pinnately divided into ligulate or linear

lobes which may become several decimeters long; in section the cells of the midline region

much taller than those of the margin, the thallus much thicker, 100 µ or somewhat more; the

margins entire to irregularly ruffled and crenate with a somewhat paler central portion Fig.

B2.2.

3 Chaetomorpha antennina (Bory) Kutzing

Chaetomorpha antennina forms stiff tufts of unbranched filaments 2 to 15 cm high composed

of large cells. Its overall color is grass green, but close examination will reveal alternating

green and white bands. The tips become pale green or yellowish when bleached by bright

sunlight Fig. B2.3. This seaweed grows intertidally on rocky coastlines exposed to large

breaking wave.

Fig. B2.1 Enteromorpha clathrata Species

193

Fig. B2.2 Ulva fasciata Species

Fig. B2.3 Chaetomorpha antennina Species

194

APPENDIX B2

Mix proportion calculations of M20 and M25 concretes

The mix design was done as per IS 10262- 1982 and the calculation for the two grades of

concrete M20 and M25 is as follows:

Design of M20 grade concrete:

From IS 456-2000, for sever exposure condition of plain concrete the recommended mix is

M20. The cement content used is 310 kg/m3.

Characteristic compressive strength of the concrete at 28 days = 20 N/mm2

Maximum size of aggregate is 10 mm

Degree of quality control is Good

Specific gravity of cement is 3.15

Specific gravity of coarse aggregate is 2.65

Specific gravity of fine aggregate is 2.70

For 10 mm coarse aggregate 3% entrapped air is used

Target mean strength of concrete = 20 + 4.6 x 1.65 = 27.59 N/mm2

For 27.59 N/mm2 from IS 456-2000 water-cement ratio recommended is 0.5.

From IS: 10262 for 20 mm nominal size of aggregates, maximum water content is

186 Kg/ m

ratioCement Water waterofWeight

3

Weight of cement (C) = = 0.5186 = 372 kg

From the total aggregate volume (1m3) deduct 3% for air entrapment. Hence net volume of

solids is 0.97 m3

10001

PCA

PFAW

PC

CAFAc

x

+++

.

TA =

Where TA – Total Aggregate

C is cement content; FA is Fine Aggregate; CA is Coarse Aggregate

PC is specific gravity of cement; PFA is specific gravity of fine aggregate

PCA

10001

65.265.0

70.235.0186

15.3372 xTATA

+++

is specific gravity of coarse aggregate

0.97 =

TA = 1752.37 kg/m

Cement = 372 kg

3

FA = 0.35 x 1752.37 = 613.33 ≈ 613 kg

CA = 0.65 x 1752.37 = 1139.04 ≈ 1139 kg

195

Design of M20 grade concrete:

From IS 456-2000, for sever exposure condition of plain concrete the recommended mix is

M25. The cement content used is 310 kg/m3.

Characteristics compressive strength of the concrete in 28 days = 25 N/mm2

Maximum size of aggregate is 10 mm

Degree of quality control is Good

Specific gravity of cement is 3.15

Specific gravity of coarse aggregate is 2.65

Specific gravity of fine aggregate is 2.70

For 10 mm coarse aggregate 3% entrapped air is used

Target mean strength of concrete = 25 + 4.6 x 1.65 = 32.59 N/mm2

For 32.59 N/mm2 from IS 456-2000 water-cement ratio recommended is 0.43.

From IS: 10262 for 20 mm nominal size of aggregates Maximum Water Content is

186 Kg/ m

ratioCement Water waterofWeight

3

Weight of cement (C) = = 0.43186 = 432.6 ≈ 433 kg

From the total aggregate volume (1m3) deduct 3% for air entrapment. Hence net volume of

solids is 0.97 m3

10001

PCA

PFAW

PC

CAFAc

x

+++

.

TA =

Where TA – Total Aggregate

C is cement conctent; FA is Fine Aggregate; CA is Coarse Aggregate

PC is specific gravity of cement; PFA is specific gravity of fine aggregate

PCA

10001

65.265.0

70.235.0186

15.3433 xTATA

+++

is specific gravity of coarse aggregate

0.97 =

TA = 1701.42 kg/m3

FA = 0.35 x 1701.42 = 595.497 ≈ 596 kg

CA = 0.65 x 1701.42 = 1105.92 ≈ 1106 kg

Cement = 433 kg

196

Table B3.1 Chemical composition of cement

Compound % Silicon-di-oxide (SiO2 20-21 ) Aluminum oxide (Al2O3 5.2-5.6 ) Ferric oxide (Fe2O3 4.4-4.8 ) Calcium oxide (CaO) 6.2-6.3 Magnesium oxide (MgO) 0.5-0.7 Sulfur-tri-oxide (SO3 2.4-2.8 ) Loss on ignition 1.5-2.5

Table B3.2 Grading of coarse and fine aggregate

Coarse Aggregate (mm)

% retained Fine Aggregate % retained

20 0 4.75 0 16 25 2.36 12

12.5 52 0.600 49 10 72 0.300 85

4.75 100 0.150 97 < 0.150 100

Table B3.3 Details of mix proportions

Type of cement

W/C ratio Cement (kg/m3

Water (kg/m) 3

Fine Aggregate

(kg/m)

3

Coarse Aggregate

(kg/m) 3

Target mean

strength (N/mm

) 2)

OPC 0.50 372 186 613 1139 27.59 OPC 0.43 433 186 596 1106 32.59

197

APPENDIX B3

Table B4.1 Temperature, RH and pH prevailed during the study period

Month & Year Temperature RH pH September’07 32.1 71.2 8.0 October’07 31.0 74.4 8.4 November’07 30.4 76.3 8.5 December’07 29.1 75.8 8.2 January'08 26.6 74.8 8.1 February'08 30.8 68.6 7.8 March'08 30.7 72.2 8.0 April'08 32.2 72.7 8.0 May'08 32.4 68.6 8.1 June'08 33.5 70.0 8.0 July'08 32.5 66.0 7.8 August'08 32.5 69.1 7.9 Spetember'08 31.8 69.7 8.3 October'08 31.2 73.9 8.5 November'08 29.2 75.8 8.4 December'08 28.5 76.0 8.3 January'09 28.2 68.6 8.4 February'09 29.9 70.6 8.1 March'09 32.1 70.6 8.2 April'09 32.5 72.1 7.8 May'09 33.2 73.1 7.8 Jun'09 33.6 64.9 7.8 July'09 32.6 62.5 7.6 August’09 33.1 70.1 8.1

Note:

(i) Maximum temperature is 33.6o

(ii) Minimum temperature is 26.6

C o

(iii) Maximum RH is 62.5%

C

(iv) Minimum RH is 76.0%

(v) pH range of actual sea water is 7.6 – 8.5

198

APPENDIX B4

Description of Phycochemical Procedures

1 Extraction

The dried, chopped, milled and weighted algal material was then soaked in methanol (MeOH)

in a large glass jar and was kept in the solvent for one month at room temperature. The extract

of the material thus obtained was then filtered to remove all solid algal particles. It was then

evaporated on a rotary evaporator under reduced pressure. This yielded a dark green, thick

residue, which was then weighed.

2 Saponification

An aliquot of the extract obtained was saponified with 10% KOH in 50% methanol and

refluxed at 100oC for 6 hours. The mixture was then concentrated under reduced pressure and

then H2O and diethyl ester (Et2O) were added. It was then shaken vigorously and the Et2O

layer was separated. The Et2O layer was evaporated and used for fatty acid analysis.

3 Esterification

All the fatty acid fractions obtained were subjected to methylation and 1.5 -2.0 mL ethereal

diazomethane was added to the fatty acid mixture. The reaction mixture was left in the fuming

chamber at room temperature, over night until dissolved. The aliquots were then directly

injected to Gas Chromatograph – Mass Spectrophotometer (GC-MS).

4. Identification

The GC-mass spectra were analytically analyzed for the identification of fatty acids.

199

APPENDIX C1

MASS SPECTROSCOPY OF METHYL ESTERS OF FATTY ACIDS OF ‘CHLOROPHYCEAE’ AT VARIOUS RETENTION TIMES

MS of ‘Chaetomorpha antennina’

Contd…

200

Contd…

201

Contd…

202

Contd…

203

204

MS of ‘Ulva fasciata’

Contd…

205

Contd…

206

Contd…

207

Contd…

208

MS of ‘Enteromorpha clathrata’

209

APPENDIX C2

OXIDATIVE CLEAVAGE OF FATTY ACIDS EXCRETED

DURING METABOLIC ACTIVITY

(A) ‘Enteromorpha clathrata’

7, 10, 13 – hexadecatrienoate CH3. CH2-CH=CH. CH2-CH=CH.CH2.CH=CH. (CH2)5.COOCH

3

Malonic Acid Malonic Acid Pimelic acid

9, 12, 15- hexadecatrienoate H2C=HC-H2C-HC=CH-CH2-CH=CH. (CH2)7.COOCH

3

Malonic Acid Malonic Acid Azelaic acid

9- heptadecenoate CH3.(CH2)6-HC=CH-(CH2)7-COOCH

3

Azelaic acid

5,7 – hexadecadiynoate CH3-(CH2)7-C≡C-C≡C-(CH2)3-COOCH

Oxalic acid Gultaric acid

3

Aerial Oxidation

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

COOH CH2

COOH

COOH CH2

COOH

COOH (CH2)5

COOH

H2O + ½ O2

H2O + ½ O2

COOH CH2

COOH

COOH (CH2)7

COOH

H2O + ½ O2

COOH (CH2)7

COOH

H2O + ½ O2

H2O + ½ O2

COOH (CH2)3

COOH

COOH

COOH

Aerial Oxidation

Aerial Oxidation

Aerial Oxidation

H2O + ½ O2

COOH CH2

COOH

210

(B) ‘Ulva Fasciata’ 8-heptadecenoate CH3-(CH2)7-CH=CH-(CH2)6-COOCH

CH

3 Suberic acid

9-heptadecenoate

3.(CH2)6.CH=CH-(CH2)7-COOCH

Azelaic acid

3

7- methyl-hexadec-6-enoate CH3-(CH2)8-C=CH-(CH2)4-COOCH

3

Adipic acid 2-trans-octadecenoate (2-18:1)

CH3-(CH2)14 H H COOCH3

Oxalic acid

H2O + ½ O2

Aerial Oxidation

COOH (CH2)6

COOH

H2O + ½ O2

Aerial Oxidation

COOH (CH2)7

COOH

C=C

H2O + ½ O2

Aerial Oxidation

COOH

COOH

H3C

H2O + ½ O2

Aerial Oxidation

COOH (CH2)4

COOH

211

6,9,12-hexadecatrienoate (6,9,12-16:3) CH3-(CH2)-CH=CH-CH2-CH=CH-CH2-CH=CH-(CH2)4-COOCH

CH

3

Malonic acid Malonic acid Adipic acid 9- heptadecenoate

3-(CH2)6-HC=HC-(CH2)7-COOCH

Azelaic acid

3

15-methyl-hexadec-9-enoate CH3-CH-(CH2)-CH=CH-(CH2)-COOCH Azelaic acid 7- methyl-hexadec-6-enoate

3

CH3- (H21C)8-CH = -(CH2)4-COOCH

3

Adipic acid

H2O + ½ O2

Aerial Oxidation H2O

+ ½ O2

H2O + ½ O2

COOH CH2

COOH

COOH CH2

COOH

COOH (CH2)4

COOH

CH3

COOH (CH2)7

COOH

COOH (CH2)7

COOH

H2O + ½ O2

Aerial Oxidation

H3C

H2O + ½ O2

Aerial Oxidation

Aerial Oxidation

COOH (CH2)4

COOH

H2O + ½ O2

212

8- heptadecenoate CH3-(CH2)7-HC=HC-(CH2)6-COOCH

3

Suberic acid 6,9,12,15- hexadecatetraenoate CH2=HC-H2C-HC=HC-CH2-HC=HC-CH2-HC=HC-(CH2)4-COOCH

3

Malonic acid Malonic acid Malonic acid Adipic acid 15-methyl-hexadec-9-enoate H3C-HC-(CH2)4-HC=HC-(CH2)7-COOCH

3

Azelaic acid 4,5-tetradecadienoate (4,5-14:2) CH3-(CH2)7-HC=C=HC-(CH2)2-COOCH Succinic acid

3

H2O + ½ O2

COOH (CH2)2

COOH

H2O + ½ O2

Aerial Oxidation

COOH (CH2)6

COOH

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

COOH (CH2)4

COOH

COOH CH2

COOH

COOH CH2

COOH

COOH CH2

COOH

Aerial Oxidation

H3C

Aerial Oxidation

H2O + ½ O2

COOH (CH2)7

COOH

Aerial Oxidation

213

2-trans-octadecenoate CH3. (CH2)14 H

H COOCH

C = C

Oxalic acid 7-methyl-hexadex-6-enoate

3

CH3. (CH2)8-HC = (CH2)4-COOCH

3

Adipic acid 9,12,15-hexadecatrienoate (16:3(n-1)) H2C=HC-H2C-HC=HC-H2C-HC=HC-(CH2)7-COOCH

3

Malonic acid Malonic acid Azelaic acid 4,5-tetradecadienoate (4,5-14:2) CH3.(CH2)7.HC=C=HC-(CH2)2-COOCH Succinic acid

3

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

COOH CH2

COOH

COOH CH2

COOH

COOH (CH2)7

COOH

H2O + ½ O2

COOH (CH2)2

COOH

Aerial Oxidation

Aerial Oxidation

H2O + ½ O2

Aerial Oxidation

COOH

COOH

Aerial Oxidation

COOH (CH2)4

COOH

H3C H2O + ½ O2

214

9-tetradecenoate (9-14:1) CH3-(CH2)3-HC=HC-(CH2)7-COOCH Azelaic acid

(C) ‘Chaetomorpha antennina’ 2-trans-octadecenoate

3

H COOCH

(CH

3 C = C

2)14

H

Oxalic acid 4,7,10,13- hexadecatetraenoate CH3-CH2-CH=HC-H2C-HC=HC-H2C-CH=CH-CH2-HC=HC-(CH2)2-COOCH3

Malonic acid Malonic acid Malonic acid Succinic acid 7,10,13 – hexadecatrienoate CH3-CH2-HC=HC-H2C-HC=HC-H2C-HC=HC-(CH2)5-COOCH Malonic acid Malonic acid Pimelic acid

3

H2O + ½ O2

COOH (CH2)7

COOH

Aerial Oxidation

H2O + ½ O2

Aerial Oxidation

COOH COOH

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

COOH CH2

COOH

COOH CH2

COOH

COOH CH2

COOH

COOH (CH2)2

COOH

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

COOH CH2

COOH

COOH CH2

COOH

COOH (CH2)5

COOH

Aerial Oxidation

Aerial Oxidation

215

2-trans-octadecenoate H COOCH3 C = C CH3(CH2)14

(CH

H Oxalic acid 15-methyl-hexadec-9-enoate

3)-CH-(CH2)4-HC = HC-(CH2)7-COOCH

3

Azelaic acid 9, 12, 15- hexadecatrienoate (16:3) H2C=HC-H2C-HC=CH-CH2-CH=CH. (CH2)7.COOCH

3

Malonic Acid Malonic Acid Azelaic acid

5,9 – hexadecadienoate CH3-(CH2)5-HC = HC-H2C-CH2-HC = HC-(CH2)3-COOCH

3

Succinic acid Gultaric acid

Aerial Oxidation H2O

+ ½ O2

COOH COOH

H2O + ½ O2

Aerial Oxidation

COOH (CH2)7

COOH

H2O + ½ O2

H2O + ½ O2

COOH CH2

COOH

COOH (CH2)7

COOH

Aerial Oxidation

H2O + ½ O2

COOH CH2

COOH

H2O + ½ O2

H2O + ½ O2

COOH (CH2)2

COOH

COOH (CH2)3

COOH

Aerial Oxidation

216

4,7,10,13- hexadecatetraenoate CH3-CH2-CH=HC-H2C-HC=HC-H2C-CH=CH-CH2-HC=HC-(CH2)2-COOCH3

Malonic acid Malonic acid Malonic acid Succinic acid 9-heptadecenoate (9-17:1) CH3.(CH2)6.CH=CH-(CH2)7-COOCH

Azelaic acid 15-methyl-hexadec-9-enoate

3

(CH3)-CH-(CH2)4-HC = HC-(CH2)7-COOCH

3

Azelaic acid 5,9 – heptadecadienoate CH3-(CH2)6-HC = HC-H2C-H2C-HC = HC-(CH2)3-COOCH

3

Succinic acid Gultaric acid

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

H2O + ½ O2

COOH CH2

COOH

COOH CH2

COOH

COOH CH2

COOH

COOH (CH2)2

COOH

Aerial Oxidation

H2O + ½ O2

Aerial Oxidation

COOH (CH2)7

COOH

H2O + ½ O2

Aerial Oxidation

COOH (CH2)7

COOH

COOH (CH2)2

COOH

COOH (CH2)3

COOH

H2O + ½ O2

H2O + ½ O2

Aerial Oxidation

217

9- Octadecenoate CH3-(CH2)7-HC = HC-(CH2)7-COOCH

3

Azelaic acid 3,7,11,15- Tetramethylhexadec-trans-2-enoate H3C COOCH

H

3 C = C

3C-CH-(CH2)3-HC-(H2C)3-HC. (CH2)3

Oxalic acid

H

H2O + ½ O2

COOH (CH2)7

COOH

H3C H3C H3C

Aerial Oxidation

Aerial Oxidation

H2O + ½ O2

COOH COOH

218

LIST OF PUBLICATIONS FROM THIS RESEARCH WORK

(A) International Journal/(s)

1. Jayakumar, S. and Saravanane, R. “Detrimental effects on coastal concrete structures by Ulva Fasciata” Accepted for publication in Thomas Telford Journal of Construction Materials

2. Jayakumar, S. and Saravanane, R. (2009) “Biodeterioration of coastal concrete

structures by Macro Algae- Chaetomorpha antennina” Journal of Materials Research, Cubo Multimidia, Brazil, 12(4)

3. Jayakumar, S., Saravanane, R. and Sundararajan, T. “Detrimental effects on coastal

concrete structures by Chaetomorpha antennina” Accepted for publication in ASCE's Journal of Materials in Civil Engineering

(B) International Conference/(s)

1. Jayakumar, S. and Saravanane, R. (2007) “Environmental effects and clean development mechanism on deterioration of structural components” Proceeding of the International Conference on Cleaner Technologies and Environmental Managemnt, Pondicherry Engineering College, Pondicherry, India, January 4-6, 717-720

2. Jayakumar, S. and Saravanane, R. (2008) “Algal detrimental effects on off shore

structural components” 14th

International Biodeterioration and Biodegradation Symposium IBBS-14, Messina, Italy, 6-11 Ottobre, 216

3. Jayakumar, S. and Saravanane, R. (2008) “Biodeterioration of concrete surface by marine chlorophyceae – Chaetomorpha antennina” International Conference on Recent Trends in Materials and Mechanical Engineering, Dr. Mahalingam College of Engineering and Technology, Pollaci, Tamilnadu, India, December 18-20, 84

4. Jayakumar, S., Saravanane, R., Sundararajan, T. and Venkatachalapathy, V.S.K.

(2009) “Biodegradation of concrete surface by marine chlorophycae – Chaetomorpha antennina” Accepted for publication in International Conference on Advances in Mechanical and Building Sciences in the 3rd Millennium, Vellore Institute of Technology, Vellore, Tamilnadu, India, December 14-16

5. Jayakumar, S., Saravanane, R., Sundararajan, T. and Venkatachalapathy, V.S.K. “Detrimental effects on coastal concrete structures by Chaetomorpha antennina” Accepted for publication in 3rd International Perspective on c

urrent & future state of water resources & the environment, Indian Institute of Technology, Chennai, India, January 5-7, 2010

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