Indian Journal of Engineering & Materia ls Sciences Vol. 10. April 2001. pp. 143- 147

Biosusceptibility studies on carbon fibre composites for aerospace applications

R B Sri vastava, M C Upreti, Mi nu Awasthi & G N Mathur

Defence Materi als & Stores Research & Development Establi shment. Kanpur. 208 0 13. India

Received 8 May 2()02: accepled / Jal/lwrv 2()()3

Advanced compositcs are w idely used as structu ral materi als in aerospace applications. Carbon fibre re in forced com­pos ites are recognized as structural units for integral fuel tanks. T he behav iour of carbon f ibre compos ites as influenced by microbiologica lly induced degradati on due to fuel resident microorganisms has been of considerable il1leresl. The present study incorporates characterization o f biofilm generated on carbon ti bre composi tes due to the growth of fue l resident mi · croorgani sms in single and combination cul tu re and their effect on the mechanical properties of carbon fibre composites. An exposure of 120 days revea led that the surface of f ibre and resin matrix gets un iformly co loni zed by microorgan isms. Aure­obosirlilllll pilI/II/a ilS as well as combinati on of cultures induced a signifi cant (P<0 .05) loss in fl exura l strength. However. no sign ifi ca nt eFFec t was rccorded on flex ural modulus and inter lam inar shear strength properti es of carbon fi bre composi tes undcr the influence of fuel residing microorganisms.

Microorganisms such as Cladosporilllll resinae. spe­c ies of Aspergillus, PenicilliulII. Pselldomonas and Bacillll s are known to grow in storage f ue l tanks and frequentl y contaminate j et fue ls I. I n the presence of condensed wa ter, microorgani sms may utili ze fue l hydrocarbons as sole carbon source fo r growth2

. The hot and humid areas of the tropi cs and sub tropics are considered mos t aggress ive enviro nment for microbi a l degradati on of fue l. The study of Scott and Fo rsy th )

revea ls th at the most prevalent spec ies at 30°C is Cla­dosporilllll resinae but at 45°C Aspergillus jilll1igat lls predominates, although other fun gal and bacteri a l spec ies are a lso capab le to grow at the latter te m­perature. Growth o f microorgani sms at the interface be tween the wate r co ndensates and the fue l in the j et aircrafts fue l tanks and radi ato rs have been reported to promote the corros ion o f a luminium a lloys used as structura l components in these appli cations-l ·

5. During

coloni zati on, the metabo li c ac ti vi ty of aerobi c and anaerob ic o rgani sms resul ts in a lterati o n o f mechani ­cal and chemica l characte ri st ics of materi alo.

Advanced co mposites are increas ing ly be ing used in aircrafts world over for the ir hi gh specific strength , process ing fl ex ibility, weight reduc ti o n and many other ad vantages over conventi onal structura l mate ri ­a ls7

. Though the compos ites are advanced eng ineering materi als, they are a lso suscepti ble to environmenta l degradati on8

. Fibre re in fo rced po lymeric compos ites have earl ier been examined fo r the ir susceptibility to microbiologica ll y in fl uenced deg radati on9

. Fibres in

compos ites may promote fungal coloni zati on by serving as capillari es fo r transporting nutrients from susceptible regions or ex terna l surface , stimul ating extensive microbio logical in vas io n. Composites may contain a range of che mi cals such as pl asti c izers, fl ame retardants, catalysts, colorants and organi cs in res ins. These chemicals can be utili zed as carbon and

b · . p ' . llo energy sources y mlcroorgal1l sms. atn sl n et a . reported preferenti a l colo ni zati on of res in fibre inte r­face in composites compri s ing epoxy and carbon fibre as constituents.

Carbon fibre re in fo rced composites are specifica ll y recogni zed as s tructura l units fo r integra l fue l tanks. The present study is an attempt to assess the effect of microbi a l species, iso lated from biocontamin ated fu e l, on the mechani cal properti es of composite materi a ls, consisting of carbo n fibre and epoxy res in matri x.

Materials and Methods Contaminated fue l sampl es were collec ted fro m

know n source and nine fung i, two bacte ri a, one yeast and two sulphate- reducing bacte ria (S RB ) were iso­lated . C ultures were brought into pure form and the ir identities were confirmed at Institute o f Microbial Techno logy (IMTEC H), C handi garh . The fun gal iso­lates were Aureobasidium pullulans, AcremoniulII strictum, Aspergillus niger, Aspergillus terricola. Cladosporium resinae, Paecilo11lyces varioti, Curvu­Laria lunata. Libertella heaveae and Asperigillus ga ­nus var. bremis, Bacteri al iso lates were Bacillus fir-

144 I DIAN J. ENG. MATER. SCI.. APR IL 20m

III is, Bacil/us lIIegaleriwlI , yeast isolate was Calldida lropicalis and SRB iso lates were Desu/p/wvibrio deslI/p/llIricall s and Desu/phovihrio vll/garis.

Epoxy res in based carbon fibre co mpos ite (CFC) sheet of thi ck ness 3.6 mm was cut in stipulated size and shape of test samples req uired for mechanica l tes ts with the help of slow speed diamond cutting whee l. All the coupons sides were sealed with epoxy resi n, weighed and steri I ized with rec ti fied ethanol. Cou pons were introduced in 250 mL conical flasks fill ed with 150 mL of sterili zed mineral sa lt medium containing CaCl" 1.8x I O '~, MgSO~ 1.7x I0·} and ( H.j)~S04 7 .6x I 0.3 M per I itre and 50 mL of steri Ie fi ltered av iati on turbine fuel (ATF). All the iso lated fungal cultures were grow n in potato dextrose agar (PDA ), aerob ic bacterial cultures in nutrient agar (N A) and yeast in malt ex trac t agar for four days. SRB cultures were grown in anaerobic environment in iron sulphite agar. Four types of inoculums were pre­pared employ ing following test cultures and flasks were inoculated separately with each type of inocu­lum along with abiotic control : (i) single fun gal cul­ture-AureobasidiulII pullulalls, (ii ) sulphate Reduc­i ng Bacteria- Desulphovihrio desu/phuricalls and DeslIlp/1Ovibrio vulgaris, (iii) mi xed culture-Acre­IlIOlliUIII striclulII , Aspergillus lIiger,Aspergillus lerri­cola, Cladospo riulII resinae, Paecilomyces varioti, CurvlIla ria Illnala, Lihertella heaveae and Aspergillus gallll s var. b reI/lis. Bacillus finllis , Bacillus megate­rilll/l and Calldida lropicalis and (iv) combination culture-A ll the fungal, yeas t and bacterial cultures including A. plil/Illans and SRB.

All the tl asks were kept at 32°C for 120 days in an incubator in dark conditions to simulate the environ­ment prevailing in aircraft fuel tanks. Viability of in­oculum was checked periodically. After completion of 120 days ex posure period, the coupons were taken out and the biofilm (developed as a result of microbial growth) settled on the test coupons ex posed to A. pulllllans, SRB, mi xed culture and combination cul­tures in replicates was extracted in steri I ized disti li ed water. The extract was shaken in vortex mixer and hundred times dilution was prepared. One millilitre of the extrac t was incubated in sterili zed PDA and NA petri plates . 0.2% streptomyc in was added in PDA medium to check the growth of bacteri al spores pres­ent in mi xed and combination inoculums. Bacteri al and fungal co lonies were counted on second and fo urth days of growth respect ively with the help of co lony counter and colonies/cm" settled on test cou-

pons were enumerated. [n similar manner, I mL me­dium from each exposure jar was di luted hundred times and microbial colonies per millili rre of medi um were determ i ned.

The biofilm deposited on the coupons ex posed to SRB was ex tracted in sterili zed disti lled wate r. The cell suspension was thoroughl y shaken. From the sus­pension I mL, 0.1 mL and 0.0 I mL of so lutions were taken and introd uced in 10 mL of Postgate medium B and made up to 20 mL in the test tu bes wh ich were maintained in tripli cates. The test tubes were fill ed up to brim and closed tightly. The inocu lum was main­tained in anaerobic conditi ons by placing them in ni­trogen fill ed anaerobic jars incubated at 30DC for 2 1 days for determining the most probable number (MPN) . The technique to determine the approximate number of bac teri al colonies/counts in term of MPN involves the formation of black precipitate of iron sulphide conforming the presence of SRB. Tubes showing SRB growth (black precipi tate of iron sul ­phide) are recorded as positi ve. The MPN of SRB present in the sample is calculated, from the pattern of positive and negative tubes, using standard MPN ta­ble. Moisture uptake was recorded fo r test coupons after drying them for three days at room temperature. Test coupons of I x I cm were used for scanning elec­tron microscopic (S EM) stud ies afte r fi xi ng them overnight in 2% glutaraldehyde.

ASTM methods were followed for measuring flex­ural strength , fl ex ural modulus 11 and inter lam inar shear strength 12 (I LSS). Tests were conducted at stan­dard laboratory atmosphere of 23 ± 2DC at 50 ± 5% relative humidity.

Results and Discussion [n the present study , for all the test organi sms ei­

ther indi vidually or collectively , a signifi cant bio­growth on the materi al surface was observed (Table I) . Bacteria formed biofilms readily and con­stituted a major component of the biofilm. The at­tachment of bacterial population on material surface indicates possible bacteri al utilization of chemi cals present on the surface of the fibres in the com posites.

During an assessment of biogrowth, max imum amount of biomass (Table 2) was observed in biofilm formed by combination culture foll owed by mi xed culture and A. pullu/alls. Analysis of biofilm revealed that genera lly the proportion of organ ic and inorgani c content was approximately around 80% and 20% in mixed culture and A. pullu/alls. [n case of SRB , how-

SR IVASTAVA e1 (II. : f3 10SUSCEPTIBILlTY STU DI ES ON CARBO FIBRE COMPOSITES 145

ever, the organ ic content const itutes nearly 96% of the biomass. The organi c content in the biofilm is indica­tive of the exten t o f biological growth in the system.

The characterization of biofilm reveal ed that maximum thickness of biofilill is generated by SRB followed by mixed culture and A. plllluions. Reduced moisture uptake was noted in CFC coupons exposed to all tes t organ isms except mixed culture. This may be attributed to the presence of biofilm, which may ac t as a diffus ion barrier for wa te r, reducing the

. k 1"\ mOisture upta e . . A SEM visualization (Fig. I) he lped in assess ing the topographical features of the material surface under

influence of different test fuel res iding microbial cul­tures. Biodegradatio n of plastic surface using SEM

has ea rl ier been reported l4. In the present s tudy , SEM

micrographs provide ev idence of active biofilm de­velopment on the mate ri a l surface. Profuse g rowth of cultures is observed on the fibre and resin matrix . Un­der the intluence of A. pullulans (Fi g. I b), rough sur-face areas were observed a lo ng with debonding be­

tween fibre and res in matrix . In presence of SRB (Fi g. Ic) bac terial colonization o n fibre surface was noted and some deep hidden areas due to high bacte-

Table I - Biogrowth on carbon fibre compos ite materi al exposed to test cullllres

Cu ltures

A. p lIl/lllalls

SRB

Mixed cullllre

Fungi

Bacteria

Combi nation cu lture Fungi

Bacteri a

Biofilm No. of colo-

nies/cm"

20.5x 104

I.7 x 10'

3I.3x I0'

4S.lxI0"

12.2x I0·

29.Sx I 04

M edium No. of

colonies/ mL

18.0x I0'

I Ax l02

o.5x 10'

19.3x I0'

17 .2x 10·

2 1Ax 104

rial growth were a lso observed. In case of mi xed cu l­ture (Fig. I d) profuse network of fungal mycel ium was observed covering the complete surface area. Combination culture also indicated profu se bi ogrowth and debonding between fibre and res in matri x. These o bservations suggest poss ible utili zati o n of chemical components of composites by microorganisms as en­ergy and carbon sources favouring profuse mi crobia l growth. Mercedes et al. 15 observed through SEM that bacteri a were interconnec ted by ex tra-cellular poly­mers and were attached to hydrocarbo n droplets as well as to the po lymeric material s. In the present study , simil ar extracellular polymer productio n was noted in case of A. pullulans supporting ex tensive colonization.

In an earlier reported work, Srivastava et al. 16 ob­ta ined sig nificant decrease in mechanica l properti es of glass fibre re inforcements in vinyl ester resin as com­pared to epoxy resin exposed to SRB. Pankhurst and Davies l7 also observed decrease in tensile strength of adhesive PVC tapes exposed to two bacterial and one fun gal cultures. In contrast to this finding , Seal and Pantke l8 through their experiments showed signifi cant increase in tensile strength of PVC films. The present study relates to mechanical properties of CFC cou­pons (six replicates), such as fl ex ural strength , flex ­ural modulus and ILSS , exposed to test organisms and is compared with abiotic controls. In case of fl exura l strength , max imum reducti on was recorded 2 .92% in combination culture foll owed by 2.89% in A. pullu­lans, 1.99% in mixed culture and only 0 .24% in SRB. Stati stically a significant (P < 0.05) reduction in flex­ural strength was observed in case of combination culture, A. pullulans and mi xed culture. SRB , how­ever, did not show any significant effect. A sli ght re­duction was also observed in flexural modulus where combination culture indicated max imum loss 0.90% followed by 0 .70% in mixed culture, 0.60% in SRB and 0 .57% in A. pullulalls. Stati stica lly no significant

Table 2- Moi sture uptake and analysis of biofilm sett led on carbon fibre composi te material

Cu ltures Moisture uptake Biotilm weight analysis Thickness o f bio-(%) Biomass wI. (g) Organic content Inorganic film ( !-l )

(%) content (%) Contro l 0.80 A. pil l/illalls 0.48 0.01 2 80.00 20.00 35.93 SRB 0.65 0.0054 96.00 4.00 235.55 Mixed culture 0.83 O.OIS 78.60 2 1.40 193.33 Combinati on OA6 0.023 73.90 26. 1 nd

Nd- Ilot determined.

146 I DIAN J. ENG. MATER. SCI .. APRIL 2003

effect was recorded as regards flexural modulus of composites exposed to all sets of microbial cultures as well as in abiotic controls.

Results of ILSS reveal that reduction in presence of combination culture was maximum 12.93% followed by 4.44% in A. pullulans, 4.29% in mixed cu lture and 1.71 % in SRB. Though the percentage of reduction in

Fig. I - SEM microg raphs of carbon fiber compos it es in prescnce of diffcrent microbial cu ltures (a) CFC control coupon. (b) CFC coupon in A. pllllllialls cultures, (c) CFC coupon in su lphate re­ducing bacteria (S RB) cu lture. (d) CFC coupon in mixed cu lture and (e) CFC coupon in combination cu lture.

ILSS as compared to control as well as with other sets of cultures was much greater in combination culture, it was found stat istica lly non-significant.

These results indicate that combination culture as compared to others caused considerable loss in me­chan ical properties. It is considered that there may be a synergist ic effect amongst the bacteria and fungi present in combination culture which may enhance the abi lity of SRB and A. puilulal/s in particular, to attack carbon fibre composites.

In a similar study related to aluminum alloy (2024) wh ich is generally used as structural component in aircraft fuel tanks, it was observed that its mechanical properties were significantly affected by the fuel re­siding microorganisms. Moreover, SRB was noted as the main causative microorgani sm next to combina-

SRIV ASTAVA el al.: BIOSUSCEPTIBILITY ST UDI ES ON CA RB ON FIBRE COMPOSITES 147

ti on culture (unpubli shed wo rk). Under the present study, it is observed that SRB as well as o ther sets o f cultures have not affec ted the mechanical properti es o f carbon fibre composites to that ex tent.

Conclusions Thi s study sugges ts that tho ugh the surfaces of f i­

bres and res in matri x get uni fo rml y coloni zed by a ll types of fue l res iding microorgani sms, microbi a l co lonizati on do not adverse ly effect mechanical prop­e rti es of carbo n fibre composites . Carbon fibre com­posites the refore can be considered as suitable struc­tural mate rial s fo r aerospace applications.

Acknowledgement The authors thank Aeronauti cal Development

Agency , Bangalore , for awardi ng the project. T he he lp rendered by Shri K N Pandey in SEM observa­tio ns is grate full y acknow ledged . Thanks are due to Shri R K Gupta and Shri K M Ch audhary for carry ing out mechanica l tes ts. Thanks are a lso due to Dr N Tejo Prakash and Dr Ranj ana Prakash fo r the ir he lp in carry ing out this work .

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