PHYSIOLOGICAL ADAPTATIONS OF MANGROVE FLORA

OF COCHIN

u Introduction

Physiology of S a l t Tolerance

Adaptations to Respirat ion

Adaptations t o Reproduction

o Materials and Methods

Analysis o f P a lan ine

Data Analysis

u Results

u D ~ s c u s s i o n

PHYSlOLOGICkL ADAPTATIONS OF MANGROVE FLORA OF

COCHIN

INTRODUCTION

The physiological processes which allow mangroves to live in the

constantly changing environment where the land meets the sea, are unique among plants.

A mangrove must contend with drying effects of the sun and wind, osmotic imbalance

caused by the high salinity of seawater in which it is immersed, and growth in salty,

oxygen deficient, and waterlogged soils. Mangroves must also have the ability to

withstand the action of tides, exposure to freshwater, destruction effects of storm surges

and the diurnal and seasonal fluctuations of temperature.

Evolutionary adjustments to varying coastal marine environments have

produced some astounding biological characteristics within mangrove plant communities.

Although these plants grow in the areas which are well saturated with water yet they

cannot directly available to them because of high concentration of salts in the soil and

water. Thus mangroves are growing on soils of physically wet but physiologically dry in

nature (Sharrna, 1997). These plants also have to cope with the problems of salinity and

excess of water (Blascol 1075). The latter creates difficulty of anchoring, aeration and

seed germination Mangrove trees have specially adapted aerial and salt-filtering ~oo t s

and salt-secretin2 leaves that enable them to occupy the saline wetlands where other plant

life cannot susvive ( Snedaker, 1993)

In response to environmental extremes, mangroves have developed

specialised means in ordel to establish. grow and reproduce.

Physiology of Salt Tolerance

Mangroves have adapted to life in a saline environment in a number of

ways. Exclusion of sodlium ions and chloride ions at the roots by a process called ultra

filtration allows Brzibwiera g~mt~orhiza, A~~icet~tr ia rnarita. Rhizophora slylosa,

Sorit~eratia sps. fi,xcoecc~ria sps arrd Aegialitis sps to remove salt from seawater as water

is absorbed for normal plant physiological processes (Anonymous,l999b). This is a

passive process which does not require the expenditure of energy by the plant.

Salt exclusi~on mechanism may be performed due to the presence of ultra

filtration mechanism in their roots (Scholander, 1968). By this ultra filtration mechanism

when the water is absorbed the salt ion remain as filtered out (Tomlinson 1994). In

Aegicerns cort~rctrla/rrn, special salt secreting glands are present in their leaves. Salt

secretion is an active process which requires the expenditure of metabolic energy to

proceed. A white film of salt crystals usually forms on the upper leaf surface of salt

secreting mangroves. Salt may also be removed by storage in the older leaves which

eventually fall from the plimt.

Salt extrusion mechanisms controlled by the salt glands are present in the

leaves of A~~icrru~ru ntaiiria and Acutrthrr.~ sp.~. also. Joshi el a/.. 1975 have pointed out

that Avicrtnritr sp. are th~e most efficient salt extruding species and can grow in high saline

conditions. but Acntr/hrc!; sps. and Aeg~certrs .ys. mostly grow in less saline habitat.

Salt resistarlce of mangroves may be controlled by the presence of salt

gland, viviparous gel-mination and succulent nature of leaves (Naskar and Mandal, 1999).

I.rrt~~tritzet~tr racettro.s~r an,d ,Sotrtrr~.~r/~a .sp.s. accumulate extra salt in its salt glands, on both

sides of the leaves. as such leaves are succulent. Salt accumulation mechanisms in

mangroves are also preisent in the species like Excoecaria agallocha, Sorlrteratia and

Xylocarprrs sr). These mangrove species deposit the sodium and chloride ions in their

stem barks and pneumatophores or knee roots and in their older leaves, as well (Joshi, et

a1 ., 1972). Due to their salt accumulation abilities, these mangrove species mostly bear

succulent leaves (Jeenings, 1968)

Adaptations to Respiration

Some mangroves have specialised root structures to deal with growth in

saline and anaerobic :;oils which are periodically covered by seawater. To enable these

root systems to breathe during times of tidal drying, specialised aerial roots known as

pneumatophores serve: to aerate those parts of mangrove plants which are deprived of an

adequate oxygen supply and in particular those root structures buried in anaerobic soil.

Thick knobbly, 'bent-knee' roots are characteristic of Ceriops t a p / . Aeration in water

logged soils is achieved by the development of a surface systems o f prop roots around the

main trunk (Natarajan., 1'398).

'The prop roots of some mangrove species such as Rhizophora and the

pneumatophores of othel-s such as A~dcrr?tlin contain many small breathing pores, called

lenticels. 'These allow oxygen to diffuse into the plant and down to the underground roots

by means of air space tissue in the cortex called aerenchyma. The lenticels are inactive

durins hi& tide

Stilt ro'3ts are observed in R/tizo(,/tora, Hrr~grrirm arid Crricy).~. The stilt

root becornin3 shallow buttresses in old trees. In Soturera~ia root system including an

extended series of catlle roots giving rise to narrow, shallowly descending lateral roots

and erect pncunlatophores. Pneumatophores at first greenish gray with a flaky back,

extending to as much as 2m. at matuarity, tapering woody via secondary thickening and

with numerous narrow second order roots developing horizontally in the substrate

(Tomlinson, 1994). Root knees are seen in Brttguiera, where the knee is initiated as the

result of primary growth. Plank roots are found in Xylocarpus p z a t u r n and Heritiera.

The following mangroves normally lack any elaborated aerial part to the

root system,: Argiceras, AAegialiris, Exc:oecaria, Katzdelia and Nypa. However, certain

structures may be developed to aerate the root system. The base of the stem in Aegialiiis

is enlarged and tluted, with a very spongy texture. Karzdelia tends to develop aerial roots

in some limiting environments

Stmctulres regarded as hydathodes have been described in mangrove

leaves. I t must be emphasized that there is no record of guttation from mangrove leaves,

but this is not surprising because positive pressures in the xylem are hardly possible with

plants rooted in sea water. Stomata are scarcely sunken, but guard cells are somewhat

thick walled, ofien with prominent or even elaborated ledges, suggesting some increased

resistance to stomata1 1:ranspiration.

Adaptations to Reproduction

Mangroves are flowering plants with male and female flower parts

situated on either the same tree or on separate trees as in Excoecaria agaNocha L.. a

milky rnangrove Flowers and fruits may appear throughout the year according to

variations in regional climate. In I<hi:ophorn slylosa aerial roots usually produce baby

trees. The baby trees can float for a long time before it finally find a good place and settle

down. A V I C ~ I I I I ~ O ninritin and 1,rrgutlcrrlnrio rncmtosn are producing seeds.

Througll viviparity embryo germination begins on the tree itself; the tree

later drops its developed embryos, called seedlings, which may have take root in the soil

beneath. Viviparity may lhave evolved as an adaptive mechanism to prepare the seedlings

for long-distance dispersal, and survival and growth within a harsh saline environment.

During this viviparous development, the propagules are nourished on the parent tree, thus

accumulating the carbohydrates and other compounds required for later autonomous

growth. The structural complexity achieved by the seedlings at this early stage of plant

development helps acclimate the seedlings to extreme physical conditions which

otherwise might preclude normal seed germination (Snedaker, 1993). According to

Gunasekar, (1995) the concentrations of chlorophyll, total sugars and starch and net

photosynthetic rate increased during the viviparous germination of Hhizophora

nirrcrorlnicr and Rhizo,~>/iorn apicr~lntn hypocotyles.

Another special adaptation is the dispersal of certain mangrove propagules

which hang from the branches of mature trees. These fall off and eventually take root in

the soil around the parent tree or are carried to distant shorelines. Depending on the

species, this propagult:~ may float for extended periods upto a year, and still remain

viable Vivipary and the long lived propagules allow these mangrove species to disperse

over wide areas

Under saline conditions more aminoacids are formed in mangroves (Joshi

et (11.. 1977). This en,hances nitrogen metabolism. The nitrogen value in halophytes

ranges from 1 54 to 2.17 (Myers, 1990). High percentage of polyphenols occur in

A~Jcet~tr;<r r?ffici~wll.s. The Nacl- salinity results in accumulation of proline in halophytes.

The halophytes show 0.4 to 46% sodium and 0.24 to 2.72 % of Potassium (Dagar et

a1.,1991).

Joshi rt al. (1975) studied the photosynthetic carbon metabolism in the

prominent Indian mangrove species. They found that aspartate and alanine are the major

product of less than 10 seconds of photosynthesis in most of the Indian mangroves. These

initially synthesized aminoacids, provide carbon for production of larger organic

materials. Studies with carboxylating enzymes showed that phosphoenol pyruvate

carboxylase was more active and incorporates more carbon, than ribulose diphosphate

carboxylase Mangroves .are Cq types of plants and possibly of aspartate formers,

however they lack kranz type of anatomy unlike other Cq plants (Dagar et a/., 1991).

Stomata of mangroves open between 8 and 10 a.m. to facilitate maximum carbon

assimilation, but in the early afternoon as the temperature increases, they open to a slight

extent and in the evening they again close and remain closed all night.

Mangroves develop a tolerance to soil salinity because they maintain a

high cellular water potential and are relatively insensitive to salt toxicity. In India, studies

have also shown that different mangrove species display tolerance for a range of salinity

(Rao, 1986) The mechanisms of salt tolerance are complex and variable and involve

factors such as ionic potentials across membranes, osmotic relationships, enzyme

activation and protein synthesis (Tomlinson, 1994). The salt exclusion mechanism of

martgroves therefore must be selective. Decreased potassium concentration at high

salinities are noticed in Ai'icei~nitr and Argicerns. This is significant to their studies of

photosynthetic responses to high salinities since potassium ions are involved in the

stomata1 mechanism and indirectly the potassium concentration could affect gas

exchanse

In the present investigation quantity of P alanine content in different

species of mangroves in Cochin area were determined to find out the influence of P

alanine in their distribution. Alanine is a non protein aminoacid which has been detected

from natural sources. Many of these are of physiological importance and interfere in

important metabolic processes. The molecular formula of alanine is C,H702N. In P

alanine NH2 assumes the position of P . The structural formula of 13 alanine is

NH2-CHZ-CH2-COOH. Seasonal variation of P alanine content in the Acanthus ilicifolirrs

with respect to salinity istratss was also determined during this investigation.

MATERIALS AND METHODS

The fresh leaf samples of all the true mangrove species identified from the

sampling stations were collected and analysed for f3 alanine during premonsoon season

1999. Prenlonsoon season showed maximum water salinity during the investigation

period. Sampling was done from the stations where the mangrove species were

represented in rnaxirnum density 411 sample collections were made from the trees of 2

10 years oltl Ac.~urr/~rrs i:lIc~fi)Ii~i.s L was found distributed in maximum area at Cochin.

So monthly analysis of ,lclrt~/hrrs il/c!foli~r.s L. leaves were conducted for an year to find

out the seasonal variation of p alanine content. Leaves were collected 1 m above the

water level In order to find out the variation of !3 alanine content, 3 replications were

taken fro111 different plants of the same species at each station. Water salinity

quantification i the plants was also carried out along with the P alanine determination

in the plants

Tree size, morphological peculiarities, leaf size and texture of the samples

collected are given below.

Rhizophoru mucrongtay Lamk :- A glabrous evergreen tree with characteristic aerial stilt

roots, appearing butressed by the mud. Bark brown, with vertical clefts. The leaves are

typically much larger simple entire, opposite, blade elliptical, mucronate up to 10 cm

broad. Leaf texture coriaceous, glabrous, but with numerous microscopic cork warts on

the lower surface visible on older leaves as fine black dots. The height of the tree was

12 m and age of the tree was approximately 15 years.

Rhizc~phoru candelaria, DC. :- A glabrous small evergreen tree, with large dark glossy

green leaves. Bark brown. Height of the tree was 20 m and the age was 20 years

approximately Size of the leaves measured as 8.65 cm.

Bruguiera gymnorhiza L. :- Trees to 20 m high with short buttresses. Bark rough, black,

fissured pattern Shoots plagiotropic by apposition, developing terminal short shoots.

Leaves elliptic, oblong.. coriacious, I0 X 6 cm, apex bluntly pointed, petiole up to 4 cm

long often glaucous with a white wax. The age of the tree was approximately 20 years.

Ilruguieru cylindricu L. :- Tree to about 18 m high with short butresses, bark greyish.

Leaves elliptic. 8 X 5 cm, with a bluntly pointed apex and cuneate base, petiole to 4 cm

long. The age of the tret: was approximately 12 years.

Kundeliu currrlel L :- Small tree, ~r-oxiny to 5 m. Butresses and pneumatophores absent.

Bark smooth. greyish. Leaves opposite. 10 X 5 cm oblong elliptic, apex obtuse, margins

entire, petiole 1 5 crn long, age of the tree was aproximatelyltj years.

Sonneratia caseoiaris L :- Trees to 10 m with continuos growth but diffuse branching,

branches horizontal. Leaves glabrous, opposite, shortly petiolate to almost sessile. Adult

leaves broadly ovate ~isually with a blunt apex some what fleshy with an entire margin,

4 X 6 cm, Leaves lanciolate. with extended red petioles, root system including an

extended series of cable roots giving rise to narrow, shallowly descending lateral roots

and erect pneumatophores. Pneumatophores extending to as much as 2 m at maturity.

Age of the tree was 1 I years aproximately.

A~~iccnniu r$fcinalis L. :- A small tree to 9 m high much branched, dense crowned.

Aerial stilt roots presenlt. Under ground roots extended, cable like and supporting as

leteral branches, erect exposed preumatophores and descending absorbing roots. The bark

is smooth. lenticellate liight coloured and not fissured. Leaf shape is ovate and with

rounded apex The tree was approximately 12 years old.

Acanthus ilicifolius L. :- A low sprawling herb to a height of 2 m. Branching infrequent

and commonly from older parts Aerial roots from lower surface of reclining stems.

Leaves decussate. us~ially with a pair of spines at the insertion of each leaf Leaves

glabrous, petiole short (Icm), blade upto 10 cm long. Gradually tapered below. The apex

usually with a sinuous, spiny margin, broadly tridentate including an apical spine. Age of

the herb was approxiriately 20 years.

l%coecuriu ugall~~-lzo~ LA :- A dioecious tree to 14 m high with abundant white latex.

E3ark grey. lenticels PI-orninent on younger twigs. Leaves spirally arranged, shoots with

infrequent ditt'use brartchiny. Leaves simple, coriaceous with a terete petiole 2 cm long,

blade ovate-elliptic, up t'o 6 cm long and 4 cm wide. Apex rounded slightly emarginate.

Margin incotlspicuously notched. with a minute gland in the notch in young leaves. Basal

glands 2, usually I on each side of blade at its insertion on the petiole. Approximately

tree was with 10 ye;m old.

Excuecoriu indicu Willd :- A tree to about 15m high with thorny bark. Regularly

crenulate, almost lanceolate leaves. The age of the tree was approximately 20 years. Leaf

size of the tree was slightly bigger than the leaves of Fxcmcaria agailocha L.

(approximately 7cm 1.

ANALYSIS OF 0 A,LANINE

& ulunine sepurution 1y TIdC :- 10 gnis of raw leaves were taken and the tissue was first

pounded in a pestle and mortar into a pulp and then extracted with 80 % ethanol in water

directly. The mixtur'e was heated to about 70' to 80' during extraction. 'The pooled

extracts were centrifuge'd and the clear supernatant was concentrated. (Jayaraman, 1988).

6-alanine were separated by thin layer chromatography using silica gel G

coated plates. The solvent used was 96% ethanol - water (7:3 vfv) The spray reagent

used was 0 3 % solut~~on of Ninhydrin in butanol containing 3 ml acetic acid. The

coloured spots &ere developed by heatlng the plates at 110 '~ for 10 minutes. Ones the

colour had developed., the plates were exposed to vopours of concentrated ammonium

hydroxide which helped in the stabilization of colours. The Rf value was calculated. The

standard Rf value of D--alanine listed is 0.47. Then identified the spot of p-alanine

comparing with the standiud.

Quantitative Estimation of p- alanine (Colorimetric method):

Scratched the identified alanine spot completely and taken in a test tube.

The sttlve~it used was hlethylcellosolve. The concentration of it was calculated by

colorimetric analysils, by applying a derivation of Beer Lambert law (Wilson, 1995;

Concentration of Optical dencity of test solution X Concentration of standard the test solution :.

Optical dencity o f standard solution.

DATA ANALYSlS

Mean anti standard deviation of salinity and p-alanine were calculated for

the 3 sampling seasons (Snedecor and Cochran, 1967) ANOVA for seasonal data were

also computed using excel programme Results of the investigation is given in the form

tables and figures

RESCJLTS

Accordin!; to the nature of salt resistant adaptations the Cochin mangroves

can be categorized into three different types. They are described below:-

Hrrig~iiercr cvlir~driccr WA. i- Salt exclus~on 1 ype I~hiroj~horcr niricrorrn~tr Lamk

( I<hizophorn ccrr~u'rliirro DC.

Salt extrus~on type

ACLIII~/~II.Y ilic1f0llii.s I , .

Av~cr~i~r i t r t?fficirrcrli.s I L-

Salt accumulation type I t.;xcorcarra npNochn L

L

Morphological adaptations for the physiological need were also observed

in different mangrove species of the Cochin area. In Karumallor stations Soti~~rraila

c ~ . s c o l ~ ~ ~ . L exhibited branched and unbranched lengthy pneumatophores. The entire

leaf surtice of .lciri,r/hrr.s iltcif~litcs I ~ . . was covered by salt and the plant had stunted

gromth 31 Putll~li ypu station

I he aminoacid 0 alanine was present in all the leaf samples analysed The

maximutii quantity of f3 alanine was obsened in k;xcorcnritr itrd;c.rr L, i r . . 246.10

i n t i e I ) . The next higher quantity was noticed in b'rrrg11ter.a cyIN~drictr

WA . Kur~tlrl~~r i<r11dt>l L and Arlnrlhr~s ilic~follrrs L. The rninimum !3 alanine content was

observed in Sotr~rrrtr/icr c n ~ ~ ~ o l ~ r ~ r . s L.: which was exclusively represented at stations such

as Nettoor. l'anarnbukad and Kammallor [Table XVII]. These stations showed a mean

salinity ol'7. 26 I and 7 9s 10~ ' l-espectively during the investigation period.

\ariation between the 13 alanine sample replications of all the mangrove

species showed insigfiifi~cant values. Leaf samples collected from the mangroves of the

same species and froni the same station showed more or less same P alanine content. The

amount 131' 11 alaliine content in the different mangrove species were dissimilar. Amino

acid content i r l iilansrove varies according to the salinity variation. alanine content of

Ac1t1 t11 I I I ~ I J / I I 1 . was lo\+ in tile salinity regions of stations 10 and 4. At station 9,

where sa l i~ i~ t \ \ \ d s the lwwest (1- 0 x 10~") Acotrtl~rr.~ ilic!fblirw L. was absent [Table XVlll

and Fi~ul-i. S /

4 ",- 4 " 100.40 --

I-! !

Fig. 7 Results o f analysis of alanine content in different mangrove species collected from the stations o f maximom density /ha. during 1999 Prenlonsoon

Table XVll

Results of a11alyse:s o'f p-alanine content (mgI100g.) in leaf samples collected during

True mangrove species and the Sample Sample Sample sites of collection -

1 I,I 1 Mean 1 Standard 1 I I 1 deviation

A13iceinricr offici~ru/i.c. . . L.

(Kumbalam)

Acotrthi~t 11r~~1filir1.s L.

(Kumbaiaiigi) ~ ~.--~

Hl~i:(y)hr~rrr n~~rcrot~u ln Lamk.

(Panambukad) - - Khlzophor~r ctr~rde/trrln DC.

(Panambuhad)

Hrtrguirlrr c:slirtdr.icr~ WA. 0.265

(Puthuvypu) - -~

Hrrrgrl~er~r mnlrrorhiza Lamk.

(Panambukad) ~ ~

Soinrrrnl~u ccr.s~~o1uri.c L.

(Panambukad) -~ -~

Klrirdellcr ccrit~/el I .

(Nettoor) - ~~

F;xcoecur-111 i ~ ~ ~ d l o c h r r L

i m r a 111 ) - - --- . -

Excoectwitr ~ialictr Wi lld.

(Karumallor) - ------ -

Table XVIlI

Station wise Seasonal Variation of Water Salinity and [3 Alanine content in

Acanthus ilicifulius observed during 1999

~~ -- .- I~

Salinity ppt. 1 a Alanine mg1100 gm

Mon- Postman- Mean soon soon soon soon

187.0 , 187.74

1 185.75 ) 185.54

2 6 % "6.80 " i ~ 8 . 0 1 187.40 1 187.5' 8 7 . 6 4

26 07 r 2 6 . I 1 187.20 187.01 187.00 187.07

186.48 186.51

- -~

2 8; .4cantl?us 111cifi)lius L. \\as 11ot found in this station -- -~

-370 3.70 1 130.53 1 130.09 1 130.10 / 130.24 I 19.78 --T19101i 180.50 j 180.64 180.59

__~ 1 - L -

STATIONS

SERIES 1 0 p-alanine mg/100g

1 SERIES 2 .+ Mean salinity ppt. 1 Fig 8 Station wise variation of water salinity and B alanine content in the leaves of

Acnntitus iZicifoNous L observed during 1999

The observation shows that salt extrusion type mangroves are most widely

distributed in Cochin area when compared to other species. The salt exclusion types like

Ka~tdrlru c<u~dt.f L. and Rhkopizorn c~zt~delc~riu DC. are found to have almost disappeared

from the Cochin area being limited to very restricted area. They require special protection

in this location otherwise they may be eliminated from the system. Similarly salt

accumulation types like fixcoecn/.irr it~dicn Willd. and So~r~~erafia careolari L. are also

rare.

'The p alanine content of different mangrove species differed. The analyses

ofAcatrrl~rr.s tlic.ifi,lir~s 1.. illustrates that alanine content is not constant for a species. It

varies with the salinity of the aquatic environment of mangrove ecosystem. Higher

salinity promoted the accumulation o f f \ alanine. It was also noted that irrespective of the

species, fi alanine content was constant in a particular locality. These results indicate a

strong correlation between salinity, f3 alanine and the distribution of the mangroves. j3

alanine does have a decissive role in the salt tolerance mechanism of the mangroves.

Physical conditions such as salinity and oxygen gradients determine the distribution of

plants (Robertson L,/ a / , 1991)

'Mangro.ves thrive in a very peculiar environment and serve as a bridging

ecosystem between lieshwater and marine systems. This has imposed several

modifications in these plants Accordins to Bhosale and Mulik (1992), aminoacids,

carbohydsates and polyphenols appear to be modified under saline conditions in

mangroves I'olyphenols seen to have a protective role to play under polluted conditions.

The polyphenoles are related to pollution stress, the nature of action is obscure (Kadam

and Bhosale. 1989). Similarly Mulik (1989) and Mulik and Bhosale (1989) reported that

the anlinoacid (3 alanine is found to play an important role in salt tolerance of these

plants The present results also suppons this view. In the investigation alanine content

in the leaves of ~L,'xcoecnr;tr itrdiccr Willd., Acuttfhr~s i/ic!folirts L., and Brtrgrriera

L ~ / / ~ I ~ / ~ I L ~ L I WA were found to be highest. The saline resistance of Acatrthrrs may be due

to its !?I alanine concentration. Nrrrgt~iertr cylitldrica WA, is observed in station 1, 6 and 7

where salinity value ranges between 29 to 33.9 x 10" . So this plant also has tolerated

high sal~nity stress. (3 alanine content is comparatively low in Sotrrrerafia ca.seolrrris L.

and Brrrprr~era jynr~~or,hizu Lamk. Both these species are found only in 3 stations.