INTERNATIONAL . BIDDETERIORATION

BuLLETIN

CATOMANCE LIMITED

manufacturers of

mystox* for the preservation of

timber, .textiles paper, cordage

plastics and specialised applications

* mgsuix is the registered trade mark of

Catomance Limited 94 BRIDGE ROAD EAST, WELWYN GARDEN CITY,

HERTS., ENGLAND. Telephone: Welwyn Garden 24373/8

Volume 17 Number 1 Spring 1981 ISSN 0020-6164

INTERNATIONAL BIODETERIORATION

BuLLETIN

CONTENTS Page No.

Biodeterioration Society Newsletter

The Biodegradation of E·Caprolactam and some related compounds a review G. Shama and D.A.J. Wase La biodegradation du E-caprolactame et de quelques composes lies: revue Der biologische Abbau von E-Caprolactam und einigen verwandten Verbindungen • Eine Ubersicht La Biodegradacion de E-Captrolactama y Algunos Compuestos Relacionados - una revision

Photodegradation of the Biocide 1, 2-Benziothiazolin-3-one used in a paper-based jointing material M J Lugg Photodegradation du biocide 1,2-benzisothiazoline-3-one utilise dans un materiau de jointage a base de papier Photoabbau des Biocids 1, 2-Benzisothiazolin-3-on, welches fiir ein Dichtungsmaterial auf Papierbasis verwendet wird Fotodegradacion del Biocida 1,2-Benzisotiazolin-3-uno Usado en Material de Junta Basado en Papel

lll

1-9

l The use of Actinomycetes in textile biocide testing 15-17

T.R. Fermor and H.O. W. Eggins Utilisation d'actinomycetes dans l'essai des biocides pour textile Die Verwendung von Actinomyceten bei der Prilfung von Biociden fiir Textilien El Uso de Actinomicetos en Pruebas de Biocidas Para Textiles ·

Use of waste tea leaves as an aid to culture of some wood-rotting fungi 19-20 N.S. Bisht and N.S.K. Harsh Utilisation de feuilles usees de the pour aider a cultiver quelques champignons de pourriture du bois Verwendung von Abfaii-Teebliittern als Hilfsmaterial zur Zucht von holzzerstiirenden Pilzen Uso de Hojas de Te para el Cultivo de Hongos que Atacan Ia Madera

Feeding responses to wood and wood extracts by Bifiditermes Beesoni (Gardner) (Isoptera: Kalotermitidae) 21-25 Muhammad Saeed Akhtar Reponses a Ia consommation de bois et d'etraits de bois par Bifiditermes beesoni (Gardner) (Isoptere: Kalotermitidae) FraPreaktionen gegenuber Holz und Holzinhaltsstoffen von Bifiditermes beesoni (Gardner) (Isoptera: Kalotermitidae) Crecimiento de Bifiditermes Beesoni (Gardner) (Isoptera: Kalotermitidae) sobre Madera y Extractos de Madera

Book Reviews 26-27

"

INTERNATIONAL BIODETERIORATION

BULLETIN

BIODETERIORATION CENTRE UNIVERSITY OF ASTON ST. PETER"S COLLEGE, SAL TLEY, BIRMINGHAM BS 3TE.

Editor-in-Chief of Biodeterioration Centre Journals Dr. H.O.W. Eggins. Editor Professor T.A. Oxley. Business Manager Dr. D. Allsopp

The Editors are able to call upon the assistance of an Editorial Board whose members are in Britain, various countries of Europe, and the U.S.A.

NOTES FOR CONTRIBUTORS The International Biodeterioration Bulletin is pu·

blished four times per year (Spring, Summer, Autumn and Winter). Typescript contributions should be sent to the Editor, Professor T.A. Oxley, at the above address.

The Bulletin acts as a vehicle for the publication of original works, including reviews, on all aspects of bio­deterioration, i.e., deterioration of materials, artefacts or facilities, of economic importance by living organisms, which include microorganisms, insects, rodents, birds, higher plants, etc. Articles on biodegradation, that is conversion of materials to less objectionable, more easily disposable, or higher value products by living organisms, are also published.

Contributions are published only in English. Each article must be accompanied by a summary in 50-150 words which will be translated into French, German and Spanish. Native speakers of these languages are invited to submit their summaries in their own language; in certain circumstances complete articles may be sub­mitted in French, German or Spanish and will be trans· lated into English for publication.

Illustrations must be very clearly drawn, normally larger than the size finally desired. The suggested final size should be clearly indicated but the Editor reserves the right to vary this in the interests of economy and clarity.

As far as possible diagrams will be reduced to single column width (80 mm) or to half page (170 mm). In any event, neither these nor half tone photographs can exceed full page (260 by 170 mm). Authors should bear in mind that it is generally more convenient for readers if legends which accompany diagrams or photographs appear with them on the same page and should proportion their illustrations accordingly. Lettering on diagrams will normally be inserted by the printer; authors are therefore asked to insert lettering or symbols in pencil on the originals or in ink on a copy.

All articles are submitted by the Editor to one or more independent referees for advice on their clarity, originality, and general suitability for publication, but the final decision whether or not to publish an article rests with the editors. If articles are rejected the sub­stance of the referee's report will usually be communic· ated to the author and in suitable case• the Editor will be pleased to help authors to improve their papers with a view to possible publication.

Bibliographic references are indicated in the text by author names (no initials) and year only, viz: Reese and Levinson (1952); or: Darby et at., (1968) and in the bibliography in strict alphabetical order of first author's names, thus:

or:

Reese E.T. and Levinson H.G. (1952) Comparative study of the breakdown of cellulose by microorganisms. Physiologica Plantarum 5: 354-366

Darby R.T., Simmons E.G. and Wiley B.J. (1968) A survey of fungi in a military aircraft fuel supply system. International Biodeterioration Bulletin 4 (1): 39-41 References to books, conference proceedings, etc.

should quote first the author(s) or editor(s), then the year of publication and title followed by the name of the publisher and the city in which it is published. As far as possible titles of journals should be given in full except for such abbreviations as 'Journ.', 'Proc.', 'Trans.' etc.

20 reprints will be sent free of charge to the first named author unless otherwise instructed. Any number (normally not more than 150) of additional reprints may be purchased if ordered sufficiently in advance. An order form and price will be sent giving about one month's notice.

ACKNOWLEDGEMENTS TO SUSTAINING ORGANISATIONS

Financial support for the Biodeterioration Centre from the following organisations is gratefully acknowledged: ALBRIGHT & WILSON (MFG) LTD.. CIBA-GEIGY LIMITED,CH4002 Busle, microbiology and microbiological

Oldbury Division, P.O. Box 3. Switzerland; manufacturers of dye- deterioration. Oldbury, Warley, Wares., England.- stuffs, industrial chemicals, plastic

B.D.H. CHEMICALS LIMITED. Lab· oratory Chemicals Division, Poole, Dorset, England; manufacturers of laboratory chemicals, biochemicals, industrial fine chemicals and micro· biocides.

BRITISH INSULATED CALLENDERS CABLES LIMITED, 38 Wood Lane, London, W12, England.

BP CHEMICALS LIMITED.

INTERNATIONAL

CATOMANCE LIMITED, Welwyn Garden City, Hertfordshire, England; manufacturers of speciality chemicals for the textile, paper, timber,leather industries, etc., including fungicides, bactericides and insecticides.

additives, photochemicals, pharma­ceutical and agricultural che~micals,

COURTAULDS LIMITED, Coventry, Warwickshire, England.

FAABENFABAIKEN BAYER A.G .• Leverkusen, Germany; manufacturers of dyestuffs, industrial chemicals, synthetic fibres, pharmaceutiCal and agricultural chemicals and preserva· tives for wood, foodstuffs and technical products.

STEALING INDUSTRIAL LIMITED. Chapel town, Sheffield, England.

HALDANE CONSULTANTS LIMITED, 27 Dawkins Road, Poole, Dorset, BH15 4JB; consultants in industrial

ii

HICKSON & WELCH !HOLDINGS! LTD .. lngs Lane, Castleford, York­shire. England.

IMPERIAL CHEMICAL INDUSTRIES LIMITED, Agricultural Division, Billingham, Co. Durham, England.

LUCAS AEROSPACE LTD .• Shaftmoor Lane, Birmingham, England. B28 ssw.

NATIONAL COAL BOARD, Coal House Lyon Road, Harrow, Middlesex, England.

RENTOKIL LIMITED, East Grinstead, West Sussex, England.

REVEATEX LIMITED. Harlow, Essex, England.

BIODETERIORATION SOCIETY NEWSLETTER

5th. International Biodeterioration Symposium University of Aberdeen, Scotland, 7-11 September 1981

The Scientific Committee which is organising the programme for this Symposium reports that around 70 presentations had been offered or commissioned by the end of March, and more are expected. They are able to announce that a gap in the programme has been filled by Dr. Dickson Liu, of the National Water Research Institute of Canada, who will chair a Session on Biodegradation of Effluent Wastes (to include chemical as well as domestic wastes). In addition, the Committee hopes that Dr. William B. Jackson, of Bowling Green State University will be able to organise and chair a short session on Biodeterioration by Rodents and Birds.

The local Organising Committee, under the chairmanship of Dr. James Shewan (79, Duthie Terrace, Aberdeen, ABl 6LS) has made excellent arrangements. There will be ample residential accommodation in University residences provided that participants are able to reserve this early, preferably in April.

At least two sessions will run simultaneously throughout the week in adjacent meeting rooms, but care has been taken to avoid, as far as possible, any clash of interests.

Early registration is recommended. The registration fee of £65 (which entitles participants to a free copy of the Proceedings) should be sent directly to Dr. Shewan, and arrangements for accommodation can also be made with him.

Canadian Wood Preservation

Members will be interested to know that a Canadian Wood Preservation Association has been formed with objectives similar to the corresponding associations in other countries. There are over 40 pressure treating plants in Canada, numerous remedial treatment companies concerned with wood in buildings, and anti-sap-stain treatments are commonly used by lumber companies. In addition, of course, research into the biodeterioration oftimber, and the prevention of preservation, has long been very active in Canada, not least by Dr. R.S. Smith, the President of the Association. Enquiries should be directed to Dr. J.N.R. Ruddick, C.W.P.A., 6620,N.W. Marine Drive, Vancouver, B.C. V6T 1X2.

.iii

Secretary of the Biodeterioration Society

For some time Mrs. Joan Maw has been assisting the Secretary of the Society, Dr. Brian Hollingsworth. Due to his work having become more remote from Biodeterioration, Dr. Hollingsworth has been obliged to relinquish all work as Secretary and Mrs. Maw is now the acting secretary. Her appointment as Secretary must await the Annual General Meeting of the Society which will be delayed this year until September when it will be held in connection with the 5th. Symposium. Meanwhile members should communicate with Mrs. Maw on all matters requiring the attention of the Secretary. Mrs. Maw's address is:-

The Hatfield Polytechnic, C.P. Snow Building, P.O.Box 109, Hatfield, Herts, England. ALIO 9AB.

International Biodeterioration Bulletin ISSN 0020-6164 17(1) Spring 81

THE BIODETERIORATION SOCIETY

APPLICATION FOR MEMBERSHIP

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Cheques should be made payable to The Biodeterioration Society. Please send this completed form, with your remittance, to the honorary treasurer:

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£4.50 for members in Great Britain and Ireland (includes meetings fee) £3.00 for members in other parts of the world (who may be asked to pay separately for meetings orga-

nised in their areas)

As a member of the Biodeterioration Society you will be entitled to a reduced rate personal subscription to the International Biodeterioration Bulletin, and/or the bibliographic journal, Biodeterioration Research Titles (B.R.T.) The normal and reduced rate subscriptions are given on the inside back page of the International Biodeterloration Bulletin.

If you are interested in this possibility please tick the box:

I may be interested in a personal subscription to either or both of I.B.B. and B.R.T. D You will be asked to undertake that your subscription is for your own personal use and not for the Institution with which you are affiliated. Do not send money for a subscription to a journal with your Society subscription. You will be invoiced by the Biodeterloration Centre of the University of Aston which publishes the journals.

iv

International Biodeterioration Bulletin ISSN 0020-6164 17(1) S~ring 1981

THE BIODEGRADATION OF E-CAPROLACTAM AND SOME RELATED COMPOUNDS A REVIEW

G. Shama1 and D.A.J. Wase'

Summary

A brief report is given of the synthesis routes of E-caprolactam together with approximate tonnages produced. Relevant toxicological considerations are mentioned along with information on the polluting nature of effluents which contain &-caprolactam. Methods of biological treatment are then discussed and a survey of the microorganisms involved in such processes is presented, together with tabulated information on those strains degrading&· caprolactam and related oligomers. Finally, a discussion of the biochemical mechanisms by which &~aprolactam and related oligomers are degraded is provided.

La biodegradation du e-caprolactame et de quelques composes lifs : revue

On fait un rapport bref des voies de synthese du e-caprolactame avec les tonnages approximatifs produits. On mentionne les aspects toxocologiques appropriCs tout au long en informant sur la nature de Ia pollution des effluents contenant du e-caprolactame. On discute ensuite des mCthodes de traitement biologique et prCsente une revue des microorganismes impliquCs dans de tels processus en donnant sous forme de tableau des informations sur ces souches dCgradant le E-caprolactame et les oligomCres tiCs. Enfin, on d.iscute les mecanismes biochimiques par lesquels le - caprolactame et les oligomeres m~s sont degrades.

Introduction

&-caprolactam is not a naturally-occuring compound. Because its most important property is its capacity for polymerisation, it is extensively manufactured and used almost exclusively to produce polycaprolactam, or nylon 6. In the USA, for example, 92.2% of the annual production of &-caprolactam was used to produce nylon 6 fibres and 7.4% in the production of nylon resins and films. (Anon 1977).

The world tonnage produced is difficult to ascertain. The capacity of most plants in the West in 1979 was of the order of tens and occasionalJy hundreds of thousands of tons. For instance, the capacity of the three US plants in 1979 was 480,000 tons, and it is likely that the plants were operating at about 65-80% capacity: Japanese production for 6 plants is of the same order. In the USSR in 1979 13 plants were operating, but the capacity of only five is known, ranging from 10,000 tons to 100,000 tons.

Der blologische Abbau von e-Caprolactam und einigen verwandten Verbindungen- Eine Ubersicht

Es wird ein kurzer Bericht Ober die Synthese von e-Caprolactam gegeben, und un gefahre Produktionszahlcn werden genannt. In d.iesem Zusammenhang werden Betrachtungen Uber toxikologische Aspekte erwlihnt und Informationen Ober die Verunreinigung durch E-Caprolactam-enthaltcnde Abwasser gegeben. Verfahren filr cine biologische Behandlung werden dargestellt, und ein Oberblick Ober die Microorganismen, die an diesem ProzeJl beteiligt sind, wird gegeben mit tabellarischen Angaben Obersolche SUimme, die E-Captolactam und verwandte Oligomere abbauen. SchlieJllich werden die biochemischen Mechanismen, durch welche E­Caprolactam und verwandte Oligomere abgebaut werden, besprochen.

La Blodegradacion de t-Captrolactama y Algunos Compuestos Relaclonados • una revision

Se describe prevemente las rutas de sintesis de &-caprolactama junto con su producci6n aproximada.

Se haec menci6n de importantes propiedades toxicas junto con Ia naturaleza contaminate de los vertidos que contienen metodos de tratamiento biologico y una panoramica de los microorganismos que intervienen en tales procesos, con informaci6n sobre cepas que degradan el E-caprolactama y los oligomeros relacionados. Finalmente se haec una discusion de los mecanismos bioquimicos de degradaci6n de &-caprolactama y oligomeros relacionados.

Nevertheless, it is clear that the USSR is a major world producer of caprolactam.

The most common feedstoc~ for the production of&­caprolactam is cyclohexane which is first converted to cyclohexanone. Hydroxylamine then supplies the nitrogen for final conversion to &-caprolactam. Of the I million tons of cyclohexane produced in the USA in 1979 about 30% was reported as being converted into &-caprolactam (Anon, 1979a), confirming the production figure reported above (Anon, 1979b). There are variants to the 'conventional' process, and one new plant in the USSR is reported to convert toluene via nitrosation with nitrosyl sulphuric acid whilst a Japanese company converts cyclohexane via photonitrosation with nitrosyl chloride (Anon 197b). Clearly &-caprolactam is manufactured on a very large scale with a consequent risk of environmental contamination.

Department of Chemical Engineering and Chemical Technology, Imperial College of Science and Technology, Prince Consort Road, London, SW7 2BY.

Biochemical Engineering Section, Department of Chemical Engineering, University of Birmingham, P.O. Box 363, Edgbaston, Birmingham Bl5 2TI.(Please address reprint requests to Dr. Wase)

(Received, January 1981)

1

Bio~egradation of £-caprolactam and some related compounds G. Shama and D.A.J. Wase

The toxicology of e-caprolactam

The toxicological properties of &-caprolactam are considered in a number of papers (Hohensee (1951), Goldblatt et a/. (1954), Ferguson and Wheeler (1973), Anon (1979c)), but only certain findings are of direct relevance to the subject of this review. For instance, Goldblatt et a/. (1954) showed that rabbits were apparently able to metabolize &-caprolactam completely, whereas rats, by comparison, excreted relatively large amounts of &-aminocaproic acid. These results have in part been borne out by more recent studies. Hargreaves and Evered (1973) noted that rat tissues in vitro were unable to transaminate &­aminocaproic acid, but that the lower homologues of e-aminocaproic acid were further metabolized by transamination. It is thus evident that mammals differ in their response to e-caprolactam; the relevance of such studies to potential human responses is therefore likely to be doubtful. Possible reasons for such differences are discussed later in this paper.

The polluting nature of e-caprolactam wastes

An examination of the literature reveals that considerable interest and concern exists in Eastern European countries as to the efficiency of installations treating e-caprolactam-containing effluents.

This may be due to the fact that controls exercised in these countries over the release of &-caprolactam to the environment are less stringent than elsewhere. For example, the maximum permissible concentration of e-caprolactam dust in working areas is lmg/m3 in the USA and in the UK. (Anon (1978)), whereas it is as high as 10mglm3 in the USSR (Anon (1979)).

Even these somewhat high limits are not, it seems, strictly adhered to. The threshold limit value (TL V) of e-caprolactam was exceeded considerably at the Rusfavi plant in the USSR with the result that the workforce at the plant contracted a number of diseases and complaints, some of which were serious and relatively long lasting. (Kuchukhidze and Dolidze (1975), Kuchukhidze and Kakhidze (1975), Levina and Shlepakov (1975)). Some ofthis concern is centred in the USSR at least, around the practice of irrigating crops .with partially treated efflu_ents containing &­caprolactam. Thus Kovtun (1976) demonstrated that the effluents from a plant producing &-caprolactam were toxic to mice even after biological treatment. Moreover, Repetun (1971) showed that concentrations of E-caprolactam in irrigation waters greater than 1000 mgllitre could when applied to wheatfields, stop growth for up to 3 weeks, and Zlateeva (1975) measured the lipase activity of mustard seed irrigated with e-caprolactam solutions and classed all solutions as unsuitable if more than 10-12% of the activity was inhibited; this corresponded to &-caprolactam concentrations greater than 250 mg/litre.

2

Evidently Russian investigators have amassed a considerable amount of evidence as to the toxic nature of &-caprolactam-containing effluents. However Novoderzhkina eta/. (1974) claimed that rabbits fed with wheat irrigated with &-caprolactam-containing waste waters were indistinguishable from controls, and hence the effluents were harmless. Unfortunately they probably overlooked the work of Goldblatt eta/ (1954), (where it was shown that these particular mammals were unusual in completely metabolising the offending material).

On balance therefore, it seems very likely that the concern felt by some USSR scientists is justified.

The Biological Treatment of e-caprolactam containing wastes

Fitter (1976) carried out small scale experiments on the degree and rate of biological degradation of a number of organic compounds among which were &­caprolactam and cyclohexanone.

The test compounds were dissolved in a biological growth medium containing nitrogen, phosphorus (in the form of phosphates) and a number of essential trace elements, and were inoculated with a small quantity of activated sludge previously adapted to the compound under test. The percent removals based on COD, were for &-caprolactam and cyclcohexanone, 94.3 and 96.0 whilst the rates of biodegradation were (mg COD g-1 h"'). 16.0 and 30.0 respectively. This compares quite favourably with 98.5% removal at 180mg COD g"1 h-1 obtained for glucose.

Experience in the waste water treatment industry appears to be that &-caprolactam-containing wastes are amenable to conventional biological treatment provided that certain precautions are observed. Zlateeva (1974) found &-caprolactam to be toxic towards activated sludge if its concentration in the effluent to be treated exceeded 350 mgllitre and that nitrification was affected by levels of &-caprolactam above 200 mgllitre. It has frequently been reported that for the biological treatment of &-caprolactam­containing effluents to acceptable standards, certain nutrient additions must be made.

Kulakov et a/. (1966), for example, noted that &­caprolactam containing effluents could be adequately treated either by combining the effluent with domestic sewage or independently, provided that phosphate was added to the effluent.

Shabi and !lett (1977) described a two-stage biological treatment system treating effluent from an &­caprolactam polymerizing plant. High rate biological filtration using a tower packed with a plastic medium was followed by sedimentation with recycling of the clarified supernatant liquid through the tower thus diluting the new effluent. Excess supernatant was

overflowed to a diffused air activated sludge plant. Provided a P:N:BOD ratio of 1:5:100 was maintained, a high percentage BOD removal was claimed.

It therefore appears that provided due account is taken of the concentration-dependent toxicity of E­capr'Jlactam and the nutrient requirement of the microbial population involved, satisfactory procedures for the biological treatment of E­caprolactam-containing wastes are available.

Micro-organisms Involved

The ability of an organism to attack E-caprolactam requires that it possess a lactamase of some sort, and early work by Iizuka et a/. (1967) suggests that the possession of lactamase activity is the exception rather than the rule in nature. For this study Iizuka et a/.

Table 1

e-caprolactam-degrading micro-organisms

Absidia

Achromobacter

Arthrobacter

Aspergillus

Bacillus

Bacterium

Byssochlamys

Citrobacter

Corynebacterium

Micrococcus

Mycobacterium

Penicillium

Pseudomonas

Rhodotorula

Trichosporon

Tosa and Chibata (1965)

Fukumura (1966a, 1966b), Kinoshita et at. (1973)

Horikoshi and Kamiyama (1977)

Tosa and Chibata (1965)

Rotmistrov et a/. (1975) Tomita eta/. (1977)

Naumova and Belov (1968)

Wase and Shama ( 1979)

Stavskaya et a/. (1975)

Fukumura (1966a, 1966b)

Uemura et at. (1966)

Tosa and Chibata (1965)

Tosa ·and Chibata (1965), Wase and Shama (1979)

Tosa and Chibata (1965) Fukumura (1966a, 1966b) Iizuka et a/. ( 1967) Naumova and Be1ov (1968) Maekawa et a/. (1972) Mirura et a/. (1977)

· Shevtsova (1969)

Shevtsova (1979)

International Biodeterioratlon Bulletin ISSN 0020-6164 17(1) Spring 1981

3

( 1967) tested a number of micro-organisms in the lAM Culture Collection at Tokyo University for the ability to utilise E-capro1actam as sole carbon and nitrogen source, and found only one, a pseudomonad, capable of so doing.

The view was to some extent upheld by Arnaud et at. (1976) who, whilst finding general amidase activity to be widespread amongst a fairly diverse taxonomic group of micro-organisms, failed to detect any organisms capable of hydrolysing E-caprolactam, an "internal amide". Nevertheless, since the first reported isolation of micro-organisms capable of degrading E­caprolactam, (Kato and Fukumura, 1962), quite a large number of micro-organisms have been implicated in E-caprolactam breakdown. Whilst some of these micro-organisms can utilise E-caprolactam as sole carbon and nitrogen source, others require an auxiliary carbon source such as glucose, employing the E-caprolactam merely as a nitrogen source, whilst still others can use it as a carbon source only. Shevstova (1969), for example was able to isolate a number of yeasts falling into each of the categories listed above.

As Tables 1 and 2 show, the view of Iizuka and co­workers is probably too limited; a very wide range of organisms is now known to attack E-caprolactam (Table 1) and related oligomers (Table 2).

Table 2

Micro-organisms reported to degrade oligomers

Organism Compound(s) attacked

Achromobacter Cyclic dimer

Alcaligenes Cyclic dimer

Corynebacterium Cyclic trimer, tetramer and pentamer, linear dimer, trimer and tetramer

Flavobacterium • Cyclic dimer

Pseudomonas Cylic dimer

Reference

Nonomura et a/. (1974)

Nonomura et at. (1976)

Fukumura, (1966c,l966d)

Kinoshita et at. (1975) Negoro et a/. (1980)

Maekawa et a/ (1972)

• Originally classified as Achromobacter

Biodegradation of E~aprolactam and some related compounds G. Sharna and D.A.J. Wase

MONOMERS

H 0 ' I; _..N-C

u OLIGOMERS

E-caprolactam

£-aminocaproic acid

Fig. I t-caprolactam and related compounds

Biochemistry of Lactam Degradation

When considering the degradation of t-caprolactam, one finds that this is linked in the literature to the breakdown of other lactams, principally y­butyrolactam and 15-valerolactam. Whilst the principal aim of this section is to make clear the mechanisms by which the breakdown of t­caprolactam is effected, valuable evidence is often obtained from mechanisms of degradation of the homologues mentioned above.

For this reason, the short survey below is not in chronological order, and mentions the degradation of homologues where this is considered helpful to the understanding of the degradation of t-caprolactam itself.

A survey of the literature indicates that it is unanimously maintained that the first product of lactam hydrolysis is the corresponding co-amino acid, (Goldblatt et a/. 1954; Fukumura0 1966b; Uemura et a/. 1966; Naumova, 1969; Kinoshita et a/. 1973; Naumova et a/. 1978).

In some cases the co-amino acid has not been shown to have been present (Tosa and Chibata, 1965; Wase and Shama, 1979), in spite of efforts to identify it. In these cases it has been assumed to have been formed and then further metabolised without accumulation, thus preventing detection. Indirect evidence supports this view; for instance Noe and Nickerson (1953) working with a strain of Pseudomonas aeruginosa which utilized y-butyrolactam as sole carbon and nitrogen source failed to detect the presence of y-aminobutyric acid.

4

However, they demonstrated that a transaminase specific for y-aminobutyric and a-ketoglutaric acids was present in cell-free extracts of organisms previously cultured on y-butyrolactam, and, in spite of their failure to detect y-aminobutyric acid as intermediate, assumed that it was involved; incidentally they went on to suggest that succinyl semialdehyde, the second product of the transaminase reaction, was further converted to succinic acid.

Tosa and Chibata (1965) using a lactam as nitrogen source showed that sometimes the corresponding co­amino acid accumulated and sometimes it did not depending on the micro-organism involved.

The question arises as to whether a common enzyme system exists capable of utilising as substrate each of the three unnatural lactams previously mentioned, (namely t-caprolactam, y-butyrolactam and 15-valerolactam), or, alternatively whether distinct systems operate.

Studies indicate that whilst the former situation may well be so for certain micro-organism, not surprisingly, it is by uo means universally true.

For example, Fukumura (1966b) observed that some of his isolates could utilie y-butyrolactam and 15-valerolactam in addition to t-caprolactam, whereas Noe and Nickerson (1958) noted that their strain of P.aeruginosa mentioned above, was unable to metabolise t-caprolactam. Nevertheless Fukumura (1966b) expressed his belief that the initial hydrolysis of lactams shared some common features. More recent genetic studies (Penfold eta/. (1977), Arst eta/. (1978), on Aspergillus nidulans have shown that mutations in a single gene-lam A - prevent the conversion of both exogenous y-butyrolactam and 15-valerolactam to their respective co-amino acids. (They do not state whether they presented their strains of A.nidu/ans with t­caprolactam).

It is moreover these workers' opinion that lam A specifies a lactamase rather than a lactam permease.

This might explain the differences in metabolic responses obtained earlier. The activity of the permeases determines whether and at what rate substrates are transferred into the cell. They are then subjected to a common metabolic system the relative activities of the enzymes compared with substrate availability determining whether intermediates accumulate or not.

It is pertinent to indicate at this point that of the assorted bacterial strains isolated by Kato and Fukumura (1962), some metabolised t-caprolactam but not t-amino caproic acid, others preferred t­amino caproic acid and still others utilised both substates indiscriminately, a finding which can now be explained on the basis of the activities of the various permeases involved.

Further evidence of a single linked enzyme system arises from the finding of Arst eta/. (1978) that the lam A gene comes under the control of an integrator gene, int A, which controls expression of three other genes. Of these, gat A specifies a y-aminobutyric acid transaminase and gab A a y-aminobutyric acid specific permease; it would therefore appear that permeases are substrate-specific but the main metabolic route is common. Confirmation of this arises from the finding that recessive mutations in gat A eliminate the y­aminobutyric acid transaminase which is also apparently active in the transamination of o­aminovaleric acid (Arst 1976).

Working along somewhat different lines, Naumova et a/. (1978) using cell extracts derived from pseudomonads and studying the transamination of a number of w-amino acids, noted that transamination activity decreased with increase in chain length from four to eight carbon atoms. As y-aminobutyric acid is a naturally occurring w-amino acid it appears likely that y-aminobutyric acid transaminase is active in the transamination of the higher homologues.

One may perhaps draw a parallel here with an observation made by Fukumura (1966a) who noted that whilst one of his strains of Corynebacterium aurantiacum lost the ability to metabolise £­

caprolactam and &-aminocaproic acid on storage, it nonetheless retained the ability to utilise ammonium adipate as sole carbon and nitrogen source.

Again this is explicable on the basis of elimination of transaminase activity by mutation. Fukumura et a/. (1964), using cell free extracts of their isolates, demonstrated transaminase activity between &­aminocaproic acid and a-ketoglutaric acid. On this basis, and by analogy with the pathway of y­butyrolactam dissimilation described by Noe and Nickerson (1958), (parts of which have since been shown to be identical to the pathways of y­aminobutyric acid metabolism by E. coli Kl2, (Dover and Halpern 1972) and to that of y-butyrolactam metabolism by Aspergillus nidulans (Arst eta/. 1978)), Fukumura (1966a) postulated the following pathway for &-caprolactam metabolism:- Fig. 2.

International Biodeterioration Bulletin ISSN 00~6164 17(1) Spring 1981

At about the same time, Naumova and Belov (1968) proposed an identical pathway for &-caprolactam metabolism. In addition to confirming the existence of transaminase-mediated transfer of the (-NH1) group between £-aminocaproic acid and a-ketoglutaric acid, they detected glutamate dehydrogenase activity. Furthermore, they succeeded in detecting adipic acid and proceeded to show that the breakdown products of adipic acid were oxidised in the tricarboxylic acid cycle.

So although the exact pathways followed by intermediates of t-caprolactam degradation after adipic semialdehyde have not been precisely elucidated for many species, and appear indeed to have been deduced from studies on the degradation of y-butyrolactam, it would nonetheless still appear that the overall scheme first proposed by Fukumura (1966a) is essentially correct.

Biochemistry of Degradation of Oligomers

Fukumura (1966a) noted that two of his strains of Corynebacterium aurantiacum possessed the ability to hydrolyse the linear di-, tri- and tetramers of £­aminocaproic acid and the cyclic tri-, tetra- and pentamers of t-aminocaproic acid; &-aminocaproic acid being the ultimate product of hydrolysis in each case. However, neither of the strains was able to attack the cyclic dimer and he ascribed the molecules' resistance to degradation to stabilisation by hydrogen bonds.

In a further paper (Fukumura 1966c), he demonstrated that exactly the same pattern of oligomer hydrolysis was obtained in cell-free extracts of C.aurantiacum. He also observed that the pH optima of ring opening activity and linear dimer hydrolysis differed and this led him to postulate that there were two enzymes involved in oligomer hyrolysis. In an additional paper, (Fukumura 1966d), he was able to confirm that this was in fact the case and succeeded in separating and purifying the two enzymes. One of these, he claimed, acted only on cyclic oligomers and performed two functions, the first of which was ring opening and the second of which was

Fig. 2 Scheme for the degradation of £-carpolactam

e:-ca.pro 1 actam

a-ketoglutaric acid

glutamic acid

e:-aminocaproic acid transaminase

e:-aminocaproic acid

•

5

CHO

I 'IH2)4 ------­cooH

adipic semi aldehyde

adipic semi aldehyde dehydrogenase

adipic acid

Biodegradation of &-caprolactam and some related compounds G. Shama and D.A.J. Wase

CYCLIC TRIMER

I I

I I

I I

t

I I

t

~

CYCLIC TETRAMER

I

I I

I I

t

I ~ I I

t

I

I I

CYCLIC PENTAMER

' I

I I ~ t I

I

t I I I

I I

t I I

I

I ~

r=::l

LINEAR PENTAMER

I ' I I I

I I

' IC:J I ' I I

I I

r=::l I!,

I

I I

r=::l ~ I I

t

r=::l r=::l Fig.} The scheme postulated by Fukumura ( 1966d) for degradation of oligomers. One enzyme(~ acts on cyclic compounds both for ring-opening and subsequent cleavage; the other enzyme •) acts only on linear oligomers. I::::J, represents a molecule of E-aminocaproic acid. (After Fukumu~a (1966d)).

cleavage of linear dimer units from the "linearised" oligomer. The second enzyme acted only on linear oligomers and served to remove only single molecules of E-aminocaproic acid. (Fig. 3).

Fukumura 's experience of the resistance of the cyclic dimer to bacterial degradation does not appear to be shared by other workers. For instance, Maekawa eta/. (1972) patented a process for the manufacture of E­aminocaproic acid from the cyclic dimer, using a strain of Pseudomonas aeroginosa, and Nonomura et a/. (1974) noted two bacteria, identified as belonging to the genera Achromobacter and Alcaligenes, which again hydrolysed the cyclic dimer to E-aminocaproic acid. 6-oxo-7-aza-tridecanedioic acid was shown to be present as an intermediate degradation product in the culture broths of both species.

Moreover, Kinoshita et a/. (1975) working with a strain of Achromobacter guttatus noted that the cyclic dimer was readily hydrolysed, in addition to hydrolysis of a number of linear oligomers.

Furthermore they showed that the oligomer­hyrolysing activity of A. guttatus relied on two separate enzymes (a similar mechanism to that found by Fukumura (1966d) for his strain of Corynebacterium aurantiacum) and succeeded in demonstrating that these two enzymes were present and active in cell free preparations. One enzyme hydrolysed the cyclic dimer to the linear dimer, whilst the second hydrolysed linear oligomers only.

Kinoshita et a/. (1977) went on to describe the isolation, purification and some of the properties of

6

the cyclic amide hydrolase. The hydrolytic action of the enzyme appears to be directed solely towards the cyclic dimer of &-aminocaproic acid, as when it was tested against a large number of related compounds including E-caprolactam, its linear oligomers and its lower homologues, no degradation products were detected.

In a recent paper, (Negoro eta/. 1980), Kinoshita and his co-workers have reclassified the organism with which much of their earlier work was done as belonging to the genus Flavobacterium and not Achromobacter as reported earlier. They show that their strain of Flavobacterium contains three plasmids and give evidence to support their theory that the genes specifying the two enzymes required for the degradation of the cyclic dimer of E-aminocaproic acid to E.aminocaproic acid are located on one of these plasmids - pOAD2. As they themselves point out, however, they have not been entirely able to discount the possibility that a regulator of the structural genes rather than the structural genes themselves is located on pOAD2. They attempted, but failed, to transfer the cyclic dimer degradation ability to a number of Pseudomonas strains. ·

It is of incidental interest that a parallel can be drawn between the possibility of these genes being plasmid encoded, and the knowledge that the genes for aerobic dissimilation of aromatic compounds are also plasmid-encoded (Fisher et a/. 1978; Pemberton and Fisher, 1977; Williams, 1980), both of these pathways being of the ring-fission type and involving substances not normally thought of as naturally-occurring.

Finally it is significant to note that whilst cell free oligomer-hydrolysing activity has been isolated. (Fukumura, 1966d; Kinoshita et a/., 1975), of the workers who have attempted to detect in vitro lactamase activity, namely Noe and Nickerson (1958), Fukumura (1966b) Kinoshita and Okada (1973) and Artst et al. (1978), only Kinoshita and Okada have achieved some degree of success.

Conclusions

I. A wide range of microorganisms is now known to degrade e-caprolactam and related oligomers.

2. The ability to degrade oligomers appears less widespread than that of e-caprolactam degradation.

3. The toxicity of e-caprolactam varies widely, and is particularly dependent on the organism involved.

4. It is unwise to employ untreated e-caprolactam containing effluents to irrigate crops for subsequent human consumption.

5. Parts of the degradative pathway of e­caprolactam are likely to be common to at least some of its homologues.

6. Provided nutrition and toxicity are accommodated, aqueous wastes containing e­caprolactam and related compounds can be successfully treated in biological treatment plants.

Acknowledgements

The authors are particularly grateful for the help given by members of the acetate and Synthetic Fibres Laboratories, Courtaulds Ltd., Coventry. In addition, they wish to place on record their gratitude to Dr. Christine Trollope, who laboriously translated numbers of papers from Russian.

References

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Anon (1977) Synthetic organic chemicals, US Production and Sales 1976. U.S. International Trade Commission, USITC Publication 833 Washington D.C. U.S. Govt. Printing Office, Washington D.C. pp.229, 304.

International Biodeterioration Bulletin ISSN 0020-6164 17(1) Spring 1981

7

Anon (1978) Registry of toxic effects of chemical substances. National Institute for Occupational Safety and Health; NIOSH 1978 edition. Editor: R.J. Lewis sen.: 137.

Anon ( 1979a) Cyclohexane, Chemical Engineering News, March 26th.: 13

Anon (1979b) Chemical Plant Data, I.P.C. Chemical Data Base. I.P.C. Business Press, London.

Anon (1979c) Caprolactam and nylon 6. !ARC Monographs on the evaluation of the carcinogenic risk of chemicals to humans. 19: 115-130.

Arnaud, A., Galzy, P. and Jallageas, J.C. (1976) Amidase activity of some bacteria. Folia Microbiologica 21: 178-184.

Arst, H.N. (1976) Integrator gene in Aspergillus nidulans. Nature, London 262: 232-234.

Arst, H.N., Penfold, H.A. and Bailey, C.R. (1978) Lactam utilization in Aspergillus nidu/ans: evidence for a fourth gene under the control of the integrator gene intA. Molec. gen. Genet. 166: 321-327.

Dover, S. and Halpern, Y.S. (1972) Utilisation of y-aminobutryic acid as the sole carbon and nitrogen source by Escherichia coli K-12 mutants. Journal of Bacteriology 109: 835-843.

Feguson, W.S. and Wheeler, ,)?..D. (1973) Caprolactam vapor exposures. American Industrial Hygiene Assoc. Journ. 34: 384-389.

Fisher, P.R., Appleton, J. and Pemberton, J.M (1978) Isolation and characterisation of the pesticide­degrading plasmid pJPl from Alcaligenes paradoxus. . Journal of Bacteriology 135: 798-804.

Fukumura, T., Shimizu, H. and Kato, K. (1964) On the enzyme hydrolysing oligomers of the eaminocaprioic acid and e-aminocaproic acid-transaminase. · The Symposia on Enzyme Chemistry (Japan) 16: 307-310.

Fukumura, T. (1966a) Bacterial breakdown of e-caprolactam and its cyclic oligomers. Plant and Cell Physiology 7: 93-104.

Biodegradation of &-caprolactam and some related compounds G. Shama and D.A.J. Wase

Fukumura, T. (1966b) Splitting of &-caprolactam and other lactams by bacteria. Plant and Cell Physiology 7: 105-114.

Fukumura, T. (1966c) Hydrolysis of cyclic and linear oligomers of 6-aminocaproic acid by a bacterial cell extract. Journal of Biochemistry 59: 531-536.

Fukumura, T. (1966d) Two bacterial enzymes hydrolysing oligomers of 6-aminocaproic acid. Journal of Biochemistry 59: 537-544.

Goldblatt, M.W., Farquharson, M.E., Bennett, G. and Askew, B.M. (1954)

&-caprolactam British Journal oflndustrial Medicine 11: 1-10.

Hargreaves, B.M.C. and Evered, D.F. (1973) Metabolism of co-amino acids by rat tissues in vitro Xenobiotica 3: 219-223.

Hohensee, F. (1951) Pharmacologic and physiological action of &­caprolactam. Faserforch. Textiltech. 8: 299-303.

Horikoshi, K. and Kamiyama, H. (1977) Bacterial Cells. Japan Kokai 77, 108, 81: Appl. 76, 24, 740.

Iziuka, H., Tanabe, I., Fukumura, T. and Kato, K. (1967)

Taxonomic study on the &-caprolactam-utilising bacteria. Journ. of Applied Microbiology 13: 125-137.

Kato, K. and Fukumura, T. (1962) Bacterial breakdown of &-caprolactam. Chemistry and Industry 1146.

Kinoshita, S., Kobayashi, E. and Okada, H. (1973) Degradation of &-caprolactam by Achromobacter gutta/us KF71. Journ. of Fermentation Technology 51:719-725.

Kinoshita, S. and Okada, H. ( 1973) Degradation of &-caprolactam by subcellular fraction of Achromobacter guttatus. Journ. of Fermentation Technology 51:753-756.

Kinoshita, S., Kageyama, S., Iba, K., Yamada, Y. and Okai, H. (1975)

Utilisation of a cyclic dimer and linear oligomers of &-aminocaproic acid by Achromobacter guttatus KI72. Agricultural and Biological Chemistry 39: 1219-1233.

8

Kinoshita, S., Negoro, S., Muramatsu, M., Bisaria, V.S., Sawada, S. and Okada, H. (1977)

6-aminohexanoic acid cylic dimer hydrolase. A new cyclic amide hydrolase produced by Achromobacter gutta/us KI72. European Journal of Biochemistry, 80:489-495.

Kovtun, V.P. (1976) Sanitary-toxicological evaluation of caprolactam production effiuents treated in biological installations. Gig. Sanit. 8: 15-18.

Kuchukhidze, G.E. and Dolidze, T.G. (1975) Effect of cyclohexanone, caprolactam and dinil on the reproductive function of workers in industry. Sb. Tr. Nauchno-Issled. Inst. Gig. Tr. Profzabol. Tiflis. 14: 188-198.

Kuchukhidze, G.E. and Kakhidze, L.P. (1975) Effect of working conditions on the health status of workers of the Rustavi synthetic fiber plant. Sb. Tr., Nauchno-Issled. Inst. Gig. Tr. Profzabol., Tiflis 14: 179-188.

Kulakov, E.A., Ishkavova, E.B. and Vandyuk, N.Y. (1966)

Biochemical clarification of waste waters containing caprolactam. Tr. Vses. Nauch.-Jssled. Jnst. Vodosnabzh., Kanliz., Sooruzhenii. Inzh. Gidrogeo 14: 18-23.

Levina, O.L. and Shlepakov, E.R. (1975) Sanitary-hygienic monitoring of working conditions at the Rustavi Chemical Plant. Sb. Tr., Nauchno-Issled. Inst, Gig. Tr. Profzabol., Tiflis 14: 84-89.

Maekawa, Y., Takenaka, N. and Kihara, R. (1972) Fermentative manufacture of 6.ACA. Japan patent 72 33157.

Mimura, A., Hayakawa, S. and Iguchi, T. (1973) Cultivation of epsilon-caprolactam-utilising microorganisms. Canada Patent 981197.

Namuova, R.P. and Belov, I.S. (1968) Conversion of &-aminocaproic acid by bacterial decomposition of caprolactam. Biokhimiya 33: 946-952.

Namuova, R.P. (1969) Bacterial degradation of &-caprolactam Prikl. Biochim. Mikrobiol. 5: 43-46.

Namuova, R.P., Giniatullin, I.M., Zakharova, N.G, and Shishkova, N.A. (1978)

Peculiarities of the amino acid composition in bacteria utilising unnatural compounds. Mikrobiologiya 47: 689-692.

j

Negoro, S., Shinigawa, H., Nakata, AS., Kinoshita, S., Hatozaki, T. and Okada, H. (1980)

Plasmid control of 6-aminohexanoic acid cyclic dimer degradation enzymes of Flavobacterium sp. KI72. Journal of Bacteriology 143: 238-245.,

Noe, F.F. and Nickerson, W.J. (1958) Metabolism of 2-pyrrolidone and y­aminobutryic acid by Pseudomonas aeruginosa. Journal of Bacteriology 75: 674-681.

Nonomura, S., Kotani, E., Urakabe, R., Shima, S. and Sakai, H. (1975)

A new metabolite, 6-oxo-7-aza tridecanedioic acid, a microbial product from cyclic dimer of E­caprolactam. Agricultural and Biological Chemistry 38: 1755-1756.

Nonomura, S., Kotani, R. and Urakabe, R. (1976) Microbial production of 6-oxo-7-aza­tridecanodicarboxylic acid Japan Kokai 76 22, 879

Novoderzhkina, Yu, G., Kigel, T.B. and Makarenki, E.V. (1974)

Harmlessness of grain grown on sewage­irrigated fields. Vop. Pit. 2: 86-88.

Patel, M.D. and Patel, D.R. (1977) Biodegradability and treatability of caprolactam waste. Indian Journal of Environmental Health 19: 310..318.

Pemberton, J.M. and Fisher, P.R. (1977) 2, 4-D plasmid and persistence. Nature (London) 267: 732-733.

Penfold, H.A., Bailey, C.R. and Arst, H.N. (1977) An integrator gene in Aspergillus nidu/ans, regulating and metabolic roles of m-amino acids. Heredity 39: 433.

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Repetun, S.l. (1971) Maximum permissible concentration of caprolactam in irrigation waste water. Melior. Vod. Khoz. 16: 9-14.

Rotmistrov, M.N., Roi, A.A. and Gvozdyak, P.l. (1975)

Spore forming bacteria of the Bacillus genus which decompose caprolactam. Mikrobiologiya 44: 727-731.

International Biodeterioration Bulletin ISSN 0020-6164 17(1) Spring 1981

9

Shabi, F.A. and Ilett, K.J. (1977) The role of biological processes in the treatment of effluents from the chemical industry. The Chemical Engineer, April: 247-248, 250.

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Study of caprolactam and p-nitroaniline degradation by bacteria. Nauchn. Osn. Tekhnol. Obrab. Vody2: 139-142.

Tomita, K., Hanamoto, H. and Kageyama, M. (1977) Treatment of caprolactam-containing waste water. Japan Patent 34551.

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Uemura, T., Kawai, I. and KoMing, V. (1966) Ninth International Congress on Microbiology. Abstracts of Reports (In Russian). Meditsina, Moscow. (Cited by Naumova, R.P. and Belov. I.S. (1968))

Wase, D.A.J. and Shama, G. (1979) The utilisation of E-caprolactam by moulds. Biotech. Letters. 1: 327-330.

Williams, P. (1980) University College of N. Wales, Bangor. Personal Communication.

Zlateeva, P. (1974) The effect of caprolactam on the biological treatment of sewage water. Tr. Vodosnabdyavane, Kanaliz. Sanit. Tekh. 10: 175-182.

Zlateeva, P. (1975) Lipase activity of seeds - a new index for determining the possibility of using industrial waste waters for irrigation and treatment by a natural biological method. · Tr. Vodosnabdyavane, Kanaliz. Santi. Tekh. 11: 105-114.

SPECIALISED

Prepared and published by the Biodeterioration Centre

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Specialised bibliographies are produced from the Biodeterioration Information Centre's document collection from 1965 as listed in the bibliographic journals Biodeterioration Research Titles (B.R.T.) and Waste Materials Biodeterioration (W.M.B.).

Bibliographies may be updated by use of B.R.T. and W.M.B. or purchase of new editions of existing titles. Copies of papers listed may be purchased from the Biodeterioration Information Centre.

Following are Bibliographies currendy available- all currendy updated

SB1

SB2

SB3

SB4 SB5

SB6

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SBS

SB9

SB10

SB11

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SB15

SB16

SB17

SB18

SB20

SB21

SB22

SB23

Glass, Ceramics and Optical Equipment-· 32 references

Stone, Brick and Concrete - 93 references

·Microbial Corrosion of Metals - 263 referencas

Microorganisms in Metal Extraction- 128 referencas

Marine Fouling- 774 referencas

Fouling of Pipelines and Water Systems - 207 references

Aquatic Weeds and Fouling of Waterways - 150 references

Rubber and Plastics Biodeterioration - 376 referencas

Fuels, Hydrocarbons and Lubricants Biodeterioration - 539 references

Pharmaceutical Products and Cosmetics- 180 references

Termites, Behaviour and Control -· 350 references

Composting of Agricultural and Municipal Wastes·· 130 references

Paints - 490 referencas

Books and Museum Specimens- 80 references

Rodents - 782 references

Single-cell Protein from Wastes - 294 referencas

Food Wastes, including Fermentation, Canning and Dairy Industries-· 406 references

Pulp and Paper Wastes- 400 references

Algae as Deteriogans- 215 references

Fungal Attack on Timber- 677 references

IN PREPARATION

SB24 Wood Preservation - 1500 references approximately

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Orders for published Specialised Bibliographies, or requests for the preparation of Specialized Bibliographies in other subjects should be addressed to:

Dr. D. Allsopp, Biodeterioration Centre, University of Aston, St. Peter's College, College Road, Birmingham B8 3TE England

10

International Biodeterioration Bulletin ISSN 0020-6164 17(1) Spring 1981

PHOTODEGRADATION OF THE BIOCIDE 1, 2-BENZIOTHIAZOLIN-3-0NE USED IN A PAPER-BASED JOINTING MATERIAL

M J Lugg

Summary

A paper·based jointing (gasket) material proofed with a biocide containing I, 2-benzisothiazolin-3-one lost its ability to resist mould growth after three months' storage exposed to sunlight. Exposure of the active ingredient to either natural or simulated sunlight caused changes in the absorption spectrum after one month and one week respectively. The role of accelerated weathering regimes in the selection of biocides for protection of stored materials is discussed.

Photodt!gradation du biocide 1,2-bcnzisothiazoline-:.H)Oe utilise dans un materiau de jointage a base de papier.

Un materiau de jointage a base de papier mis a l'essai avec un biocide contenant du 1,2-bensizothiazoline-3-one a perdu sa possibilite de resistance aux moisissures a pres trois mois de stockage en exposition a Ia lumif:re solaire. L'exposition de Ia matil:re active a Ia lumiCre solaire naturelle ou simulee a produit des changements dans le spectre d' absorption apres respectivement un mo_is et une semaine. On discute du rOle des regimes de vieillissement accelere dans Ia selection des biocides pour Ia protection des mathiaux stockes.

Introduction

Paper jointing, ie., a paper-based gasket material manufactured to BS 4249:1967, contains not only cellulose but also gelatine, glycerol and sorbitol. Since all these constituents are microbial nutrients a biocide is incorporated into the material during manufacture to prevent subsequent biodeterioration. To assess the resistance of the finished paper jointing to biodeterioration the material is challenged with an a~eous suspension of fungal conidia and incubated at 30 C in an elevated relative humidity for 28 days. This procedure is specified in BS 4249:1967.

It has been noted in this laboratory that paper jointing manufactured to the above specification was able to resist colonization by fungi when tested immediately after manufacture, but was susceptible to biodeterioration after a period of storage.

Other workers have reported the weathering action of light upon biocides (Siu, 1951; Lukens, 1973; Neihof, 1977, and Neihof, Bailey, Patouillet and Hannan, 1979). The paper jointing studied in the present work was treated with a proprietary biocide containing I, 2-benzisothiazolin-3-one as the active ingredient. In order to assess whether the reduction in efficacy of this biocidal treatment was a result of photodegradation, the effects of sunlight upon both the treated paper jointing and the isolated active ingredient were investigated.

Photoabbau des Blocids 1, 2-Benzisothiazolin-3-on, welches fUr ein Dichtungsmaterial auf Paplerbasis Yerwendet wird

Ein Dichtungsmaterial au Papierbasis, welches mit einem Biocid, das I, 2-Benzisothiazolin-3-one enthielt, ausgerOster war, verlor nach 3 monatiger Langerug bei Sonnenlicht seine F3higkeit, gegen Schimmelbefall zu schUtzen. Einwirkung von natUrlichem oder kunstlichem Sonnenlicht auf die Wirksubstanz nach einem Monat bzw. Einer Woche bewirkte Anderungen im Absorptionsspektrum. Die Rolle der beschleunigten Witterungsverfahren bei der Auswahl von Biociden zum Schutz von gelagertem Material wird besprochen.

Fotodegradacion del Biocida 1,2-Benzisotiazolin-3-uno Usado en Material de Junta Basado en Papel

Material de juntas en Ia que se utiliza papel tratado con un biocida que contenia 1,2-benzisotia~olin-3-1 perdio su resistencia al crecimiento de bongos tras tres meses de almacenajc expucsto a Ia luz solar. La exposici6n del componente activo a Ia luz natural o simulada causaron cam bios en el- espectro de absorci6n al mes y a Ia semana respectivamentc. Se discute el papel de los sistemas de envegecimiento acelerado en Ia solecci6n de biocidas para proteger materiales almacenados.

Experimental

Exposure of Paper Jointing

Squares (side 15 mm) of the paper JOmting were enclosed in either clear or black polyethylene film and exposed in duplicate on the ledge of an east facing window. A third set was stored unwrapped on shelving not exposed to direct sunlight, as a control.

Samples were assessed in triplicate for resistance to mould growth in accordance with the method described in the Introduction (BS 4249:1967), initially after 3 months' exposure and subsequently after 6 months' exposure. The exposure period was between October and April. The extent of any mould growth was recorded as 'light' 'medium' or 'heavy'.

Exposure of Biocide

I, 2-benzisothiazolin-3-one was extracted in water from a proprietary formulation and purified by evaporation to remove the water, leaving a brown solid which was recrystallised from methylated spirit, yielding a pale brown solid.

Successful separation was confirmed by nuclear magnetic resonance and mass spectroscopy. An aqueous solution of the biocide (0.015 g/litre) was prepared and the absorption spectrum recorded using a Pye Unicam SP8 100 spectrophotometer with

Biodeterioration Laboratory, M.Q.A.D., Royal Arsenal East, LONDON, SE18 6TD. (Received, October 1980)

II

Photodegradation of 1,2-bcnzisothiazolin-3-one in a paper-based material. M. Lugg

distilled water as reference. 3 ml aliquots were placed in tubes made of 'Spectrosil' glass which has very low absorption in the UV and visible spectrum above 200 om. Spectrosil is supplied by Thermal Syndicate Ltd., Howe Bros. Gateshead-on-Tyne. Samples were exposed in triplicate as below:

I. To sunlight on a flat roof, tubes supported at 45° facing south.

2. To light emitted by an Original Hanau Xenotest 150 to induce accelerated light ageing (Schafer, 1967).

3. In darkness, as a control.

Additional tubes were wrapped in aluminium foil and exposed as above to act as station controls. All tubes were sealed with 'Cling Film'.

Absorption spectra of samples exposed in the Xenotest were recorded after one and after two weeks. Those exposed on the roof were recorded after one month. The absorption spectra of the dark controls were recorded on all three occasions. Before spectra were recorded, the volumes were made up to 3 ml with distilled water where any loss due to evaporation had occurred.

2.0 --STOCK 1\ 2.0 --sTOCK

--- XENOTEST II -o--ROOF

: t.61

I I I I 1.6

I I I

1.2

~ 0 I .J c I •• 0.8 e. 0 I 11 <

I ;> ::> I 0.4

I /

'/

400 300 200 400 300

Results

Paper Jointing

The results are given in Table I. Initially, samples were resistant to biodeterioration as shown by their inability to support mould growth when subjected to the test procedure of BS 4249:1967. After three

Table 1

Effect of exposure to light on the mould resistance of paper jointing. Scores of triplicate specimens

Initial After 3 After 6 months months

Control + +-

Black polyethylene + + (+)

+++ +++ +++ +++

Clear polyethylene

= no growth + = medium growth

+ = heavy growth +

(+) = light growth

2.0 - -- XENOTEST 1\

A -•-- ROOF AI

I ·f.! t1 l, I I 1.6 !t l • I I I • ' • I I I • \ • I

0 1.2 •

I I • • I I

0.8

0.4

200 400 300 200

wavelength (nm) wavelength (nm) wavelength (nm)

A B c

Figure 1. U.V. Absorption spectra of 1,2-benzisothiazolin-3-one after exposure to daylight and to the Xenotest. I

A. Stock control compared with material exposed to xenotest.

B. Stock control compared with material exposed on the roof to daylight.

c. Material exposed to Xenotest light superimposed on material exposed to daylight.

12

i I

..--------------------------------~ -

. . •

months' exposure to sunlight, however, samples stored in clear polyethylene film had lost this resistance and showed heavy fungal growth when subjected to the mould growth test. Samples stored in black polyethylene and samples stored away from direct sunlight were still resistant to biodeterioration . Three months' further storage under these latter conditiOns led to a slight decrease in fungus resistance .

Biocide

The stock solution of I, 2-benzisothiazolin-3-one, extracted from the biocide formulation, was found to have absorption maxima at 318 nm and 226 nm (Figure 1). After one week's exposure to simulated sunlight in the Xenotest all three samples showed changes in their absorption spectra (Figure lA). The peak at 318 nm had disappeared and that at 220 nm had shifted to 194 nm. The station and dark controls showed no changes. A second week's exposure caused no further spectral changes. After one month's exposure to natural sunlight all three samples showed similar changes in absorption spectra to those exposed in the Xenotest (Figure IB). Station controls were again unaffected. When superimposed, the changes occuring during the two regimes of exposure were seen to be practically identical (Figure I C).

Discussion

The results of the first experiment with proofed paper jointing indicate that although the efficacy of the biocide had decreased slightly after 6 months' storage, a complete loss occurred after only 3 months' exposure to sunlight. The fact that the sunlight had to pass through both the window glass and the clear polyethylene film did not prevent this breakdown from occurring.

Although there was no attempt during these storage trials to control temperature within the polyethylene film wrapping, Neihof eta/. (1979) concluded that the photodegradation of pyridine-N-oxide 2-sulphonic acid biocides was unaffected by ambient temperature or pH. Further, they concluded that the wavelengths between 320 nm and 355 nm were most effective in bringing about photolysis, a range of wavelengths passed by window glass.

The absorption spectrum of the extracted active ingredient of the biocidal formulation showed changes indicative of structural alterations in the molecule after exposure to natural or simulated sunlight. Since the station controls were exposed under identical temperatures to those showing spectral changes in both the roof and Xenotest exposure trials, it may be concluded that the changes are not temperature related.

Neihof, (1977) found that biocides based on derivatives of pyridinethione were very susceptible to photodegradation. Lukens, (1971) states that coloured

lnternDtiona/ Biodeterioration Bulletin ISSN 00~6164 17(1) Spring 1981

13

compounds absorbing light in the visible or near ultra­violet part of the spectrum, are susceptible to photodegradation. In the work reported here the extracted material was a light brown compound which absorbed in the ultra-violet region of the spectrum.

It is recommended that the selection of biocidal treatments for stored materials should take into account the effects of possible photodegradation. If materials may be exposed to sunlight during storage, tests of biocidal efficacy immediately after incorporation may not be adequate to predict performance during storage.

Although direct comparisons can not be made between exposure of active ingredients and proofed materials due to the effects of medium and presence of other chemicals, information on the susceptibility of biocides to photodgradation may be gained by exposure of active ingredients to either natural or simulated sunlight. The method ·described is rapid; ultra-violeUvisible absorption spectra provide a convenient technique for detecting light-induced changes.

References

BS 4249:1967 Specification for paper jointing British Standards Institution, London.

Lukens, R.J. (1971) Chemistry of Fungicidal Action Chapman and Hall, London. P. 28.

Neihof, R.A. (1977) Photodegradation of biocides. 25th Conference on microbiological deterioration of military materiel, 16-18 Nov 1976 US Army NARADCOM Technical Report Natrick/TR-77/014 Natick, Massachusetts

Neihof, R.A., Bailey, C.A., Patouillet, C. and Hannan, P.J. (1979)

Photodegradation of mercaptopyridine-N-oxide biocides. Archives of Environmental Contamination and Toxicology 8: 355-368

Schafer, V. (1967) Accelerated light exposure to the Xenotest; results, limits, and comparison with other equipment. Applied Polymer Symposium No. 4 Weatherability of Plastics Materials. John Wiley & Son, New York.

Siu, R.G.H. (1951) Microbial Decomposition of Cellulose Reinhold, New York p.381.

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International Biodeterioration Bulletin ISSN 0020-6164 17(1) Spring 1981

THE USE OF ACTINOMYCETES IN TEXTILE BIOCIDE TESTING

by T.R. Fermor1 and H.O.W. Eggins2

Summary

Mesophilic actinomycetes were grown on cotton textile test strips (from T.N.O., Netherlands) to investigate the cellulolytic and staining abilities of these organisms. The efficacy of biocides in protecting a starch finished cloth against biodeterioration by actinomycetes was assessed both visually and by tensile strength measurements.

Utilisation d'actlnomycetes dans l'essal des biocides pour textile

Des actinomycetes mesophiles ont CtC cultivCs sur les bandes d'essai en coton du TNO pour rechercher les possibilitCs cellulolytiques et de coloration de ces organismes. L'efficacitC des biocides a protCger des tissus finis a !'amidon contre Ia biodeterioration par les actinomycetes a CtC estimCe visuellement et par des mesurcs de resistance en tension.

Introduction

Next to wood products, cotton textiles utilise a greater quantity of industrial biocides than any other product. It is most important that the efficiency of such biocides is rigorously tested before they are marketed. Pure culture trials ofbiocides are mainly restricted to using fungi as the test organisms, very little notice has been taken of other organisms which may be significant. For example, actinomycetes possess a large battery of enzymes and produce a wide range of metabolic products which may possibly give them some resistance to biocides, and thus explain some of the variable results obtained by soil-burial testing.

Many actinomycetes which occur in soils, composts, manures and fodders can degrade cellulose (Lacey, 1973). Stutzenberger (1972) describes the production of cellulases by the thermophilic Thermomonospora curvara isolated from compost, but its activity was much lower than the fungal cellulases, as was that of cellulases from a mesophilic Streptomyces species (Mandels, 1975). Siu and Reese (1953) detected cellulase activity in filtrates of Streptomyces in contact with cotton duck. The first report of in service deterioration of cotton by actinomyces was by Betrabet eta/. (1968); they found that the deterioration of untreated cloth stored in Bombay warehouses was caused by actinomycetes.

This paper describes experiments to determine the ability of Streptomyces species to degrade starch sized and unsized cotton and to measure their susceptibility to textile biocides.

Ole Verwendung von Actlnomyceten bel der Priifung von Bloclden ffir Textllien

Mesophile Actinomyceten worden auf TNO Baumwollstreifen gezilchtet, um die ccllulolytischen und foirbenden Eigenschaften dieser Organismen zu untersuchen. Die Wirksamkeit von Biociden zum Schutz von mit St!l.rke ausgeriistetem Gcwebc gegen Acinomyceten wurde visuell und durch Messung der Zugfestigkeit beurteilt.

El Uso de Actinomicetos en Pruebas de Blocldas Para Textiles

Para investigar Ia celulolisis y tinci6n de actinomicetos mesofilicos, se hicieron crecer sobrc tiras de algod6n TNO.

La eficiencia de los biocidas para proteger unos - tejidos almidonados se detecto visualmente y por medidas de tracci6n.

Materials and Methods

Organisms

Mesophilic Streptomyces species were previously isolated and characterized using standard techniques (Fermor and Eggins, 1981). Streptoverticillium waksmanii (Waksman) Baldacci, Farina and Locci was the gift of Mr. A. Barr, Catomance Ltd., Welwyn Garden City.

Test cloth and biocide treatment

Pure cotton standard test cloth was supplied by Central Laboratory, T.N.O., Delft, Netherlands, its specification was as given by Walton and Allsopp (1977). In all experiments loomstate cloth and sized loomstate cloth (wheat starch added at I% w/w of cloth) were used as controls. The following biocides were added to the cloth as a percentage ofthe weight of starch finish on the cloth:

(i) sodium pentachlorophenate, 0.5%

(ii) salicylanilide, 0.1%

(iii) tributyltin oxide (T.B.T.O.), 0.1 and 0.2%

It must be emphasised that the biocide doses used were of a concentration designed to preserve the starch sizing agent only, not the cloth itself. All biocides were

, applied at these recommended concentrations to the loomstate cloth by Catomance ltd., who also checked cotton samples for biocide residues after microbial testing.

Microbiology Department, Glasshouse Crops Research Institute, Worthing Road, Littlchampton, West Sussex, BN16 3PU.

· Biodeterioration Centre, St. Peter's College, University of Aston in Birmingham, College Road, Sattley, Birmingham, B8 3TE.

15

Usc of Actinomycetes in textile biocide testing T.R. Fermor and H.O.W. Eggins

Unprotected and biocide treated T.N.O. test cloth strips (2.5 x 5.0 em) were placed on the surface of cellulose agar (Eggins and Pugh, 1962). The strips were dampened with sterile distilled water. Streptomyces spores were inoculated onto the textile and the agar surround, and growth was visually assessed (Fermer. and Eggins, 1981) after incubation at 30°C for 5 days.·

Tensile strength

The tensile strength of cotton strips (2.5 x 15.0 em) was measured to see whether Streptomyces were active cellulose degrading organisms or merely surface colonizers. Strips were inoculated with spore suspensions of Streptomyces BS, B28, Streptomyces griseo/us (W8), or Streptoverticillium waksmanii in dilute organic salts solution (Waksman, 1967). The cotton strips were not sterilized as this caused leaching or degradation of the biocides. Rigorous checks by light microscopy and plating replicate samples were performed to ensure that only the organism inoculated

colonised the test strips. Colonisation by other organisms was infrequent and contaminated strips were immediately removed from test tanks. Inoculated strips were suspended over water in sterile tanks at 25°C. Separate tanks were used for each combination of organism and biocide. After 28 days strips were removed, washed and then air-dried for 24 hours. Prior to testing strips were conditioned overnight at 25°C and 65 ± 1% relative humidity. Ten replicate strips were individually broken using a motor-driven lensometer (Houndsfield).

Results and Discussion

All mesophilic Streptomyces species tested were able to colonise cellulose agar and with the exception of S. a/bus and S. jlavogriseus could colonise the surface of pure loomstate cloth (Table 1). All species colonised starch finished cloth, greater growth occurring because all species easily assimilated the starch size.

Table 1

Growth of Streptomyces species on untreated and biocide treated cotton textile

Streptomyces species Loomstate Loom state Loomstate cloth + starch finish (I% wlw) + biocide t

~cloth cloth+ 0.1% 0.5% 0.1% 0.2% starch Salicylanilide sodium penta- TBTO TBTO

finish (1% wlw) chlorophenate

S. a/bus B12 s S+

S. atroo/ivaceous B20 s S+IA s

S. jlavogriseus A3 S+IA -· S. griseo/us W8 A+ A+ s -· A+ AlA+

S. griseorubens S+IA A A S+IA

S. griseus B 30 s S+ -· S+

S. /avendulae B29 A A+ AlA+ AlA+

S. nigrifaciens A 72 A A -· S. setonii B I s s -· Streptomyces sp. B8 Unidentified A A AlA+ AlA+

Streptomyces sp. B28 Unidentified A A+ -· S+

Streptoverticillium waksmanii R14 .J

p p p

s sparse substrate mycelium ·A sparse aerial mycelium no mycelial growth

S+ strong substrate mycelium A+ mature aerial mycelium • inhibition zone formed by biocide in agar

p pink staining t Biocide expressed as % (wlw) of starch finish on loomstate cloth

16

.,

' I I

Only Streptomyces sp. B8 caused significant (P.;;;O.Ol) tensile strength loss of loomstate cloth. However, tensile strength losses of starch sized strips by S. griseo/us W8, B8 and B28 were in the range 7-24% (P ..:;0.1). This confirmed the cellulolytic ability of these species in producing transparent 'clearing' zones in opaque cellulose agar. Streptoverticillium waksmanii caused considerable pink staining of unprotected strips but only slight growth could be seen on examination at the microscope. Streptomyces waksmanii appeared to be non-cellulolytic and did not produce 'clearing' zones in cellulose agar.

Sodium pentachlorophenate and salicylanilide were found to be completely effective biocides for the protection of starch size (and hence the sized cotton cloth) against Streptomyces degradation. T.B.T.O., although used at the manufacuturer's recommended concentration to give protection to the size, was less effective. T.B.T.O. (0.1%) allowed Streptomyces growth (Table I) and consequent small tensile strength losses by the cellulolytic species. T.B.T.O. (0.2%) prevented tensile strength losses, but did not reduce pigment production and staining of the fabric.

In view of their cellulolytic and staining abilities, Streptomyces should be included in future lists of standard test organisms when textile biocides are evaluated by means other than soil-burial.

Acknowledgements

We are most grateful for the financial support given (to T.F.F.) by Catomance Ltd., Welwyn Garden City. Mr. Aleyn Barr (Catomance) and Dr. Dennis Allsopp (Biodeterioration Centre) are thanked for their constructive criticism of the manuscript.

References

Betrabet, S.M., Dasani, U.P. and Bhat, I.G. (1968) Studies on cellulolytic micro-organisms. Part l: Microflora associated with the degradation of cotton in storage in Bombay. Textile Research Journal 38: 1189-1197.

Eggins, H.O.W. and Pugh, G.J.F. 1962) Isolation of cellulose decomposing fungi from the soil. Nature, London 193: 94-95.

Fermor, T.R. and Eggins, H.O. W. (1981) The effect of water activity on growth of Streptomyces species. International Biodeterioration Bulletin 16 (4).

Lacey, J. (1973) Actinomycetes in soils, composts and fodders. In: Actinomycetales: Characteristics and Practical Importances, pp.231-251. Edited by G. Sykes and F.A. Skinner.

· London: Academic Press.

/nternOtional Biodeterioration Bulletin ISSN 0020-6164 17(1) Spring 1981

17

Mandels, M. (1975) Microbial sources of cellulases. In: Cellulose as a Chemical and Energy Resource. pp.81-105. Edited by C.R. Wilke . New York: John Wiley.

Siu, R.G.H. and Reese, E.T. (1953) Decomposition of cellulose by microorganisms. Bacteriological Review 19: 377-416.

Stutzenberger, F.J. (1972) Cellulolytic activity of Thermomonospora curvata: Optimal assay conditions, partial purification, and production of the cellulase. Applied Microbiology 24: 83-90.

Waksman, S.A. (1967) The Actinomycetes - a summary of current knowledge. Ronald Press, New York.

Walton, D.W.H. and Allsopp, D. (1977). A new test cloth for soil burial trials and other studies on cellulose decomposition. International Biodeterioration Bulletin, 13 (4): 112-115.

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TITLES

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CY FRUIT AND VEGETABLE JUICES

77/1-CY-0261 Growth and toxin production by

1 1 1 1 1 1 1 2 2 3 3 3 3 4 4 5

Clo:::tl'idium botul.inum in mouldy tomato juic~. HUHTANEN, C.N.; NAGHSKI, J.; CUSTER, C.S.; RUSSELL, R~W. (East. Reg. Res. Cent., ARS, Philadelphia, PA 19118, U.S.A.). AppZ. •. ~."nViT'On. tfl:c:l'obiol., 1976, 32(5): 711-715. En.

DA FRUIT AND VEGETABLES

77/1-DA-0391 The storage and transport of fruits and vegetables. THORNE, S. (Dep. Food Sci., Queen Elizabeth Coll., Univ. London, England). Orahal'dist N. ·;. 1976, 49( 6):

EX Animal fibres EY Adhesives FB Lignin FE Cellulose .. FG Vegetable fibres FJ Timber FL Bacterial attack FM Terrestrial fungal attack FN Marinejaquatic fungal attack. FO Insect attack FP Marine borer attack FR Preservation FT \food pulp .. FU Paper FV Books, etc .. FX

control of mango anthrac­nose with benomyl and hot water. MUIRHEAD, I. F. (Dep. Primary Ind., Indooroopilly, Queensland, Australia). Aw::t. ,}, Exp. Agria. Anim. Husb. 1976, 16:

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Dl SOFT FRUIT - OTHER

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Effect of time of planting on fruit yield and runner production of cold stored and freshly lifted strawberry plan~s. COX, J.E. (Dep. Agric., Gosford, New South Wales, Australia). Aust. J. ::.·xp. Agl"ic. Anim. 1/usb. 1976, 16: 604-607. En.

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International Biodeterioration Bulletin ISSN 0020-6164 17(1) Spring 1981

USE OF WASTE TEA LEAVES AS AN AID TO CULTURE OF SOME WOOD-ROTTING FUNGI

N.S. Bisht1 and N.S.K. Harsh1

Summary

When tea leaves have been extracted they are normally treated as waste. It has been found that waste tea leaves can be used as the basis for an effective and economical medium for isolation, oxidase reaction test, and stock culture of wood-rotting fungi.

Utilisation de feullles usees de the pour alder B cultiver quelques champignons de pourriture du bois

Quand on a elttrait des feuilles de thC:, on les traite normalement comme de dCchets. On a trouve qu'elles pouvaient i:tre utilisCes comme base d'un milieu efficace et Cconomique pour l'isolement,le test de reaction de l'oxydase et Ia culture de champignons de pourriture.

Introduction

Tea, as a beverage, is a Chinese discovery; it is obtained from the leaves of Camellia sinensis. India and China are the major tea producing countries while England is one of the chief consumers. Tea contains 2-5% theine and 13-18% tannin.

When an infusion is made with hot water, the alkaloid and oil readily dissolve out giving a refreshing drink (tea). The residual, extracted tea leaves which remain after straining are then thrown away as unavoidable waste. Cellulose is, of course, the main component, but the used leaves also contain some tannin as well as other compounds.

In view of the fact that wood-rotting fungi exhibit oxidase reaction on media containing tannic acid, and have a good capacity for degrading cellulose, we considered that used tea leaves, which provide a basis for a useful and economical laboratory medium.

We tested this hypothesis in three ways:

Isolation

Isolation of wood-rotting fungi is somewhat difficult for they are slow growing and liable to contamination by bacteria, moulds and other fungi.

Conical flasks (150 ml) each containing 5g of dried used tea leaves and 25m! distilled water, were autoclaved and inoculated. It was found that the fungi grew moderately rapidly on this medium, but since tea leaves are not readily spreadable on petri dishes, they were mixed with agar. Different ratios were tested and the final formula adopted for tea agar medium was:

Verwendung von Abfali-TeebHittern als HUfsmaterial zur Zucht von holzzerstlirenden Pilzen

Wenn TeebUI.tter extrahiert worden sind, werden sie normalerweise als Abfall angesehen. Es wurde festgestellt, daB AbfallteebUI.tter als Giundlage fUr ein wirksames und billiges Medium zur Isolierung von Pilzen, fUr den Oxidase-Reaktions-Test and fUr die Haltung von holzzerstOrenden Pitzen verwendet werden kOnnen.

Uso de Hojas de Te para el Cultivo de Hongos que Atacan Ia Madera

Cuando las hojas de Te han side extraidas son normalmente desechadas. Se ha encontrado que estas hojas pueden ser usadas como base para un medio efectivo y economico para el aislamiento, prueba de oxidasa, y cultivos de conservacion de bongos que atacan Ia madera.

dried, used, tea leaves 150 g

agar agar

distilled water

15 g

1000 ml

This was found to provide all the necessary nutritional requirements. It is especially interesting that many of the fungi grown on this medium eventually produced fruit bodies (basidia and basidiospores). No bacterial contamination has been observed on this medium, neither have other common contaminants appeared.

Oxidase reaction

The presence or absence of extracellular phenolic oxidases has been used as a criterion to separate wood­rotting fungi into two broad groups, the white rot and the brown rot fungi, respectively. Nobles (1948) gives the following medium for observing the oxidase reaction:

malt extract 15 g

agar agar 20 g

gallic/tannic acid 5g

distilled water 1000 ml

On this medium white rot fungi exhibit a diffusion zone due to the presence of phenolic oxidases. To use the tannin of used tea leaves for this purpose we have added an extract to two basic media, malt agar and potato dextrose agar, as follows:

·Department of Botany. Kumaun University, Naaini Tal263 002 India. (Received, January 1981)

19

Use of waste tea leaves in culture of some wood·rotting fungi. N.S. Bisht and N.S.K. Harsh.

IOOg of dried used tea leaves were mixed with 250 ml of distilled water and boiled for 10 minutes; the filtered extract was then added to either malt extract agar or to potato dextrose agar. Malt extract agar was prepared by dissolving 20 g of malt extract in I 00 ml of distilled water and added to 15 g of agar agar dissolved in 500 ml distilled water. The tea leaf extract was added to this and the whole made up to one litre. The potato dextrose agar was prepared in a similar way.

The autoclaved medium was poured into sterilized petri dishes and tested with five white rot and five brown rot fungi. The results are given in Table I. They show that used tea leaves can successfully be used in place of tannic acid for the oxidase reaction test.

Stock culturing

Cultures of wood-rotting fungi are normally maintained on malt extract agar or potatoe dextrose agar. As these media incur some cost, the possibility that used tea leaves could be used for this purpose was tested. We have found that they provide a complete nutrition source for these fungi and they can also be used as a medium for maintenance of the cultures. For this purpose we use 15 gof dried used tea leaves with 25 ml of distilled water. This will maintain cultures for six months at room temperature. All the hypha! characters and various physiological activities

(cellulose and lignin decomposition) were well retained during this period.

We are aware that other authors have published lists of selective media suitable for isolation and culture of wood-rotting fungi but all these are relatively expensive and contain relatively expensive or not readily available inhibitors (Hale and Savory, 1976). We therefore believe that it will be of interest to other workers to know that a free waste material can be used with good effect.

Acknowledgement

We are grateful to Professor B. S. Mehrotra, Head of the Department of Botany, Kumaun University, for providing laboratory facilities.

References

Hale, M.D.C. and Savory, J.G. (1976) Selective media for the isolation of basidiomycetes from wood. International Biodeterioration Bulletin 12(4): 112-115

Nobles, Mildred K. (1948) Studies in forest pathology VI, Identification of cultures of wood-rotting fungi. Canadian Journal of Research 26: 281-431.

Table 1

Tests for oxidase reaction on malt agar (MA) and on potato dextrose agar (PDA) containing extract of used tea leaves

Fungi

white rot organisms:-

Coria/us hirsutus (Wulf.ex.Fr.) Que!

C. versicolor (L.ex Fr.) Que!.

Daedalea jlavida Lev.

Phellinus gilvus (Schw.ex. Fr) Pat.

Polyporus arcularius Baisch ex Fr.-

Brown rot organisms:-

G/oephyllum striatum (Sow. ex. Fr.) Murr.

G. subferrugineum (Berk.) Bond & Sing.

Serpula /acrymans (Wulf.ex Fr.) Karst.

Trametes sepium Berk.

T. serialis Fr.

20

Presence or absence of diffusion zone

on MA on PDA

PRESENT

PRESENT

PRESENT

PRESENT

PRESENT

ABSENT

ABSENT

ABSENT

ABSENT

ABSENT

PRESENT

PRESENT

PRESENT

PRESENT

PRESENT

ABSENT

ABSENT

ABSENT

ABSENT

ABSENT

..

International Biodeterioratlon Bulletin ISSN 0020-6164 17(1) Spring 1981

FEEDING RESPONSES TO WOOD AND WOOD EXTRACTS BY BIFIDITERMES BEESON! (Gardner) (ISOPTERA: KALOTERMITIDAE)

Muhammad Saeed Akhtar'

Summary

The heartwoods of three wood species were treated for their natural resistance to attack by Bifiditermes beesoni; these were: Dalbergia sissoo, Pinus wallichiana and Cedrus deodara. Comparisons were made of the survival and feeding of Bffiditermes beesoni on wooden blocks, unextracted sawdusts, solvent extracted sawdusts and extracts applied to filter papers. Extraction was with acetone-hexane-water (53:44:3). The upper layer of the extract of Cedrus deodara proved toxic to the test termite. Minimum feeding was recorded on the heartwood block or Dalbergia sissoo, but survival of the test termite on filter paper treated with the upper layer of the extract of Dalhergia sissoo was not affected. The natural resistance of Da/hergia sissoo is therefore presumed to be because of its hardness.

Reponses a Ia consommation de bois et d'extraits de bois par Blfiditermes beesoni (Gardner) (lsoptere : Ka1otermltidae).

On a essaye Ia resistance naturelle des bois de coeur de trois especies de bois quant a l'attaque par Bifiditermes bessoni; il s'agit de : Dalbergia sissoo, Pinus wallichiana,et Cedrus deodara. On a fait les comparaisons de Ia survie et de Ia nutrition de Bifiditermes beeson/ sur blochets de bois, sur sciures non extraites, sur sciures extraites a aux solvantset sur les extraits appliques a des papiers filtre. L'extraction a CtC faitc au moyen d'acCtone- hexane- eau (53-44-3). La couche supCrieure de l'extrait de Cedrus deodora s'est reveiec toxique au test termite. La consommation minimum a CtC notCe sur le blochet de bois parfait de Dalbergia sissoo, mais Ia survie n'a pas etC affectCe dans l'extrait de Dalbergia sissoo. On prCsume que Ia resistance naturelle de Dalbergia sissoo est due A sa duretC.

Introduction

Dalbergia sissoo Roxb., Pinus wallichiana A.B. Jackson, and Cedrus deodara (D.Don) G. Don f. are the most important timbers of Parkistan and are commonly used for woodwork in buildings. Among these, Cedrus deodara is preferred.

Wood of Da/bergia sissoo is moderately hard and heavy with an average weight of 800 kg per m3 (SOlb per ft 3

) at 12% moisture content. Pinus wallichiana is soft and light with an average weight of(512 kg per m3

(32lb per ft3) at 12% moisture content, whereas the

wood of Cedrus deodara is moderately hard and somewhat heavier with an average weight of 560 kg per m3 (35lb per ft3

) (Ahmad and Ayaz, 1970).

During earlier field experiments on the narural resistance of various timbers (Akhtar and Ali, 1979), Dalbergia sissoo was found to be highly resistant to attack by the termite Odontotermes obesus but wooden blocks of Cedrus deodara previously dried at 60°C for

Fra8reaktionen gegenuber Holz und Holzlnhaltsstoffen von Blfidltermes beesoni (Gardner) (Isoptera: KalotermiUdae)

Das Kernholz von den drei Holzarten Dalbergia sissoo, Pinus wallichiana und Cedrus deodara wurde auf seine naturliche Dauerhaftigkeit gegen Bifidltermes beeson/ geprUft. Es wurde cine vergleichcnde Untersuchung der Uberlebensrate und der fraBaktivit!it von B. bcesoni mit HolzkiOtzchen, nicht extrahiertem Siigemchl, mit Siigemehl, das mit einem LOSungsmittel extrahiert war, und mit Filterpapier, welches mit Holzinhaltsstoffen getriinkt war, durchgefUhrt Die Extraktion erfolgte mit einem Gemisch aus Azeton, Hexan und Wasser (53:44:3). Die obere Schicht des extraktes a us C. deodara wirkte auf die gepruften Termiten giftig. KcmholzklOtzchen a us D. sissoo wurden am wenigsten angergriffen; die Uberlebcnsrate der Termiten auf Filterpapier, welches mit der obcren Schicht des. extraktes aus D, sissoo getrankt war,wurde jedoch nicht vermindert. Die natOrliche Dauerhaftigkeit von D. sissoo wird daher auf die Hiirtc dieser Holzart zurfickgefahrt.

Creclmiento de Bifiditermes Beesonl (Gardner) (lsoptera: Kalotermitidae) sobre Madera y Extractos de Madera

La resistencia natural de Ia medula de Dalbergia sissoo, Pinus wallichiana y Cedrus deodara fue estudiada ante el ataquc de Bifiditermes beesoni. Se com para Ia supervivencia y alimentacion de Bifiditermes beesoni sabre bloques de madera, serrin sin extraer, serrines extraidos con solventes y extractos aplicados a papcl filtro. La cxtracci6n fue hecha con acetona, hexano, agua (53,44,3). La parte superior del extracto de Cedrus deodara es toxica al termita testigo. La alimentaci6n fue minima sabre Ia medula de Dalbergia slssoo, pero Ia supervivcncia del termita testigo no se afecta sabre papel de filtro tratado con Ia cepa superior del extracto de Delbergia slssoo. La resistencia natural de Dalbergia sissoo es en consecuencia debida presumiblemente a su dureza.

48 hours were severely damaged by the same termite. However, Akhtar and Ali found that the acetone­hexane-water extract of Cedrus deodar a was somewhat toxic which suggests that the toxic principle is volatile.

Bifiditermes beesoni (Gardner) has been selected for experiments because it can be reared and maintained easily in the laboratory and is the best material for experimental studies. In Pakistan it is common in the subtropical continental lowland (Akhtar, 1974), attacking mostly trees in orchards. Its preferred hosts are Bauhinia variegate and Prunus bokhariensis. It has also been collected from Dalbergia sissoo (Akhtar, 1972). At first Bifiditermes beesoni attacks the dead portions of a tree but later on the damage is extended to the live portion also. In spite of the notable work on its biology by Ahmad et a/. (1979), little is known about its wood preferences. This paper reports the effect of extracts of heartwoods of Da/bergia sissoo, Pinus wa//ichiana and Cedrus deodara on the feeding and survival of Bifiditermes beesoni for the first time.

Department of Zoology, University of the Punjab, Lahore, Pakistan (Received September 1980: in final form December 1980).

21

Feeding responses to wood and wood extracts by Biftditermes beesoni. M. Saeed Akhtar

Materials and Methods

Nymphs of Bijiditermes beesoni (Gardner) used in the laboratory studies were obtained from colonies infesting Bauhinia variegata and Prunus bokhariensis growing in the Botanical garden, Punjab University campus. The nymphs were fed on blotting paper for three days and unhealthy and inactive individuals noticed during the period were eliminated. Approximately last instar nymphs were used for the experiments.

Wood pieces of Dalbergia sissoo, Pinus wallichiana and Cedrus deodara were purchased from a commercial lumberyard. Heartwoods of the timbers were used for the recovery of extracts.

Five grams of sawdust of each wood was extracted with 120 ml of solvent consisting of a 53:44:3 mixture of acetone-hexane-water (AHW). Carteret a/., (1972, 1979) have found AHW very efficient in extracting termicidal components from wood. The extraction was made in a soxblet apparatus according to Vogel (1964). The soxhlet was allowed to siphon six times for the extract from each timber.

The extract recovered by means of the soxhlet apparatus was placed in a separating funnel and the upper layer separated from tbe lower layer. The upper layer was usually approximately 80 ml and the lower layer (mainly aqueous) approximately 20 mi. The extract solutions were coloured, the colour varying with the type of timber and the layer.

The test materials were as follows:

I. Five grams of unextracted sawdust. 2. Extracted sawdust. 3. Filter paper treated with 80 ml of the upper layer

of extract.

4. Filter paper treated with 20 ml of lower layer of extract.

5. A wooden block weighing 5 grams previously dried at 60°C for 48 hours.

Treated filter papers and sawdust were air dried at 25±1°C for 48 hours. Filter papers treated with 120 ml of AHW were used as control. Three replicates of 50 termites each were used for each test material. The petri dishes containin.f. the test material were kept suitably moist at 25±1 C. At the end of the test period, all the test materials were dried by the same procedure as had been used before they were exposed to the termites. They were then weighed again and the amounts consumed thus determined. Daily observations were made during the exposure period and dead termites were removed from the cages.

Results

Feeding

Of the three species of wooden blocks, m1mmum feeding was recorded on the wood of Dalbergia sissoo. Bijiditermes beesoni showed maximum feeding on the wooden block of Cedrus deodara, but consumed the least amount of filter paper treated with the upper layer of its extract which proved highly unpalatable and toxic. Bearing in mind the toxic effect of the upperlayer, the natural resistance of Cedrus deodara can be attributed to the presence of certain chemicals which are lost during heating because the wooden blocks dried at 60°C were damaged by Bijiditermes beesoni.

Among the treated filter papers, maximum feeding was noted on that treated with the upper layer of Dalbergia sissoo which can be contrasted with the least feeding on its wooden block. The natural resistance of

Table 1

Amount consumed (milligrams) of each test material in 4 weeks by Bijiditermes beesoni. Each figure is the mean of three replicates and the figure in brackets is the standard error of that mean.

Wood species Control • Wooden Upper layer Unextracted Lower layer Solvent extracted and block of extract sawdust of extract sawdust vernacular name

Dalbergia sissoo 149.8 21.4 53.3 30.0 60.0 37.4 Shish am (15.7) (1.9) (13.2) (10.3) (5.8) (0.7)

Pinus wallichiana 265.0 23.2 48.3 22.1 84.7 50.9 Kail (50.8) (4.5) (2.6) (2.9) (I 1.8) (3.4)

Cedrus deodara 232.9 30.3 15.8 53.3 59.3 98.6 Deodar (31.4) (4.2) (3.9) (7.4) (14.0) (5.8)

• Although the controls were filter paper in each experiment the different amounts consumed could be due to differences in the termites themselves since they were collected from a different source for each experiment.

22

,]

Da/bergia sissoo wood appears, therefore, not to be due to any extractable constituents; it may thus be due to its hardness.

The feeding behaviour of Bijiditermes beesoni on the various test materials of Pinus wal/ichiana was similar to that on those of Dalbergia sissoo (Table 1).

Survival

Amongst the wooden blocks the maximum survival was noted in cages containing blocks of Pinus wal/ichiana and the minimum in those containing Cedrus deodara. The upper layer of the extract from Cedrus deodara proved toxic to Bijiditermes beesoni, whose survival after 4 weeks zero as against 77.2% and 95.6% in the wooden block and control respectively. The lower layer of the extract of Cedrus deodara was also toxic to termites but its termicidal potency was very much less than that of the upper layer.

The upper layer of the extract from Da/bergia sissoo did not seriously affect the survival of termites and that of Pinus wal/ichiana had no effect. In fact none of the test materials from Pinus wallichiana had any significant effect on the survival of Bijiditermes beesoni (Table 2).

Discussion

The natural resistance of certain woods to termite attack results primarily from extractive constituents (Wolcott, 1957; Saeki eta/., 1971; Carter and Smythe, 1974). Neither hardness nor high lignin content of wood prevent it from being eaten by dry wood termites (Wolcot, 1951). Behr eta/., (1972) report that there is a strong correlation between hardness of the wood and its resistance to attack by termites other than dry wood termites. Heartwood of Dalbergia sissoo is very hard and Akhtar and Ali (1979) have reported that, during field experiments, this timber was not damaged even

International Biodeterioration Bulletin ISSN 00~6164 17(1) Spring 1981

slightly when placed in the mounds of Odontotermes obesus. It has also been shown by Akhtar (unpublished) than an AHW extract of Dalbergia sissoo does affect the survival of Odontotermes obesus to some extent but, as shown in the present work, does not affect the survival of Bijiditermes beesoni. The natural resistance of Dalbergia sissoo may, therefore, be due solely to its hardness or it may be due to the presence of a toxic principle not extracted by AHW. A non-polar solvent might, perhaps, prove more efficient in extracting such an antitermitic material, if one is present.

As regards the natural resistance of Cedrus deodara, its extract in AHW is toxic to termites. Further studies on its extracts in non-polar solvents and its essential oil might reveal more toxic material.

Conclusions

Cedrus deodara is highly resistant to the attack of Bijiditermes beesoni, but loses its resistant quality if heated. The upper layer of its extract in AHW is highly termicidal. The natural resistance of Dalbergia sissoo appears probably to be due to its hardness as the upper layer of its extract has no effect on the survival of Bijiditermes beesoni. Pinus wallichiana is not resistant to attack by Bijiditermes beesoni.

Acknowledgement

This work has been conducted under the Pakistan Science Foundation Grant No. PSF/RES/P-PU, Bio 79.

Table 2

Percentage survival of Bijiditermes beesoni (Gardner) after 4 weeks Each figure is the mean of three replicates and the figure in brackets is the standard error of that mean ,

Wood species Control Wooden Upper layer Unextracted Lower layer Solvent extracted and block of extract sawdust of extract sawdust vernacular name

Dalbergia sissoo 94.6 75.2. 86.8 84.0. 88.6 92.6 Shisham (0.6) (5.2) (0.6) (4.0) (1.2) (1.7)

Pinus wal/ichiana 93.2 90.0 92.6 91.2 95.2 97.2 Kail (1.2) (2.4) (1.6) (0.6) (1.2) (I. 7)

Cedrus deodara 94.6 77.2. 0.0. 52.0. 43.2. 96.6. Deodar (2.8) (5.4) (0.0) (7.2) (7.6) (1.2)

• For a wood species (horizontal line) means asterisked differ significantly at P = 0.05 from the control (Duncan's multiple range test)

23

Feeding responses to wood and wood extracts by Bijulitermes beesoni. M. Saeed Akhtar

A 0. SISSOO [H.W.I

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80

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U : UPPER LAYER OFT~ EXTRACT

N ; UIIUTRACT£0 SAWOUST

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Figure 1 Feeding (D) and mean percent survival of Bifiditermes beesoni (Gardner) on different ,test materials ,of Dalbergia sissoo (A), Pinus wa/ichiana (B) and Cedrus deodara (C).

I Vertical bars indicate standard error of the mean.

References

Ahmad, S.S. and Ayaz, M. (1970) Identification of some of the common timbers of Pakistan. Pakistan Forest Institute, Peshawar, Bulletin No.I.

Ahmad, M., Sheikh, K.H., Jafri, R.H., Kyani, S.A., Afzal, M., Kahn, K.I., Salihan, Z. a_nd Fazal, Q. (1979)

Biology, Pathology and Distribution(in relation to soil conditions) of Termites ofPakistan. Final technical report, PL-480, project No. Al7-FS-2l Grant No. FG-Pa-201. pp 424.

Akhtar, M.S. (1972) Studies on the Taxonomy and zoogeography of the Termites of Pakistan. PhD Thesis, University of the Punjab, Lahore.

Akhtar, M.S. (1974) Zoogeography of the termites of Pakistan. Pakistan J ourn. of Zoology 6: 85-104.

24

Akhtar, M.S. and Ali, S.S. (1979) Wood preferences and survival of Coptotermes heimi (Wassmann) and Odontotermes obesus (Rambur) (Isoptera). Pakistan Journ. Zoology. ll(2): 303-314.

Behr, E.A., Behr, C.T. and Wilson, L.F. (1972) Influence of wood hardness on feeding by the subterranean termite Reticulitermes flavipes (Isoptera: Rhinotermitidae) Annals of the Entomological Society of America 65(2): 457-460.

Carter, F.L. and Smythe, R.V. (1972) Extractives of Baldcypress, Black Walnut, and Redwood and survival of the eastern subterranean termite, Reticulitermes flavipes. Annals of the Entomological Society of America 65(3): 687-689.

Carter, F.L. and Smythe, R.V. (1974) Feeding and survival responses of Reticulitermes flavipes (Koller) to extractives of wood from eleven coniferous genera. Holsforschung 28(2): 41-45.

Carter, F., Stringer, C.A. and Taras, M.A. (1979) Termicidal properties of slash pine wood related to position in the tree. Wood Science 12(1): 46-51.

Saeki, I.M., Sumimoto, M. and Kondo, T. (1971) The role of essential oil in resistance of coniferous woods to termite attack. Holzforschung 25(2): 57-60.

Vogel, A.I. (1964) A Text Book of Proctical Organic Chemistry Including Qualitative Organic analysis. Longmans, Green & Co. London.

Wolcott, G.N. (1951) Termite-repellent wood extractives. Journ. of Agriculture, University of Puerto Rico. 27: 224-227.

Wolcott, G.N. (1957) Inherent natural resistance of woods to the attack of the West Indian termite, Cryptotermes brevis Walker. Journ. of Agriculture, University of Puerto Rico 41: 259-311.

International Biodeter/oratlon Bulletin ISSN 0020-6164 17(1) Spring 1981

25

BOOK REVIEWS PESTICIDE MANUFACTURING AND TOXIC MATERIALS CONTROL ENCYCLOPAEDIA

Chemical Technology Review No. 168 Environmental Health Review No. 3 Pollution Technology Review No. 69

Edited by Marshall Sitting. Noyes Data Corp. N.J. USA 1980

ISBN 0-8155-0814-X pp. xix + 810 US 96

The success of the predecessor to this book, Pesticide Progress Encyclopaedia, published in 1977, which contained 558 subject entries, can be judged by the fact that it is now out of print. The present volume is essentially an up-dated and expanded version, with greater accent on environmental and possible pollution aspects, as the associated review titles indicated.

A short introductory survey of pesticide manufacture and formulations is followed by even shorter sections on toxic materials control in both manufacture and use, and on environmentally acceptable alternatives to pesticide usage, including biodegradable compounds, slow release formulations, biological control with, finally, a note on the concept of integrated pest management.

Deletion of some 100 suprseded pesticides from the earlier work, together with the addition of recently introduced compounds, means that the present volume comprises 514 entries, arranged alphabetically by common or generic names, those peculiar to the USA being preferred when there is any conflict. For each compound there is given, in regular sequence, its function, chemical name,structural formula, and trade names (or code numbers),followed by details of the manufacturing process. These include a description of the raw materials involved and of synthetic procedures for the intermediates, including reaction conditions,often accompanied by flow charts. Each entry is completed by the addition of toxicological data, usually in the form of acute oral LDso values for rats, together with some comments on any limitations on exposure and on tolerances set by the Evironmental Protection Agency of the USA. Finally, relevant references are cited, including invariably one to the 6th. edition of the Presticide Manual (published by the British Crop Protection Council), and to U.S. and British patents. The contents are again completed by two useful indexes: raw materials and trade names. The small bibliography, which refers to the earlier preliminary sections, might well have been more appropriately sited with them rather than at the end of the book.

When I received the earlier volume in 1977, I commented that the absence of analytical,

26

formulation and toxicological data limited its usefulness. I am pleased to note that the latter deficiency has been made good, but regret that references to appropriate analytical methods are conspicuous by their absence, and there is still no reference to minor components in the technical products. Degradation and metabolism information, both of extreme importance today, are also lacking, though space (or secrecy?) considerations have perhaps precluded any attempt to cover these vital aspects.

The book is extremely well produced and free from errors; it will prove a useful reference work, complementary to the Pesticide Manual, for many workers in the pesticide field.

D. Woodcock

THE STABILITY OF INDUSTRIAL ORGANISMS

B.E. Kirsop, Editor. Proceedings of a Symposium held at the University of Newcastle, 20th July 1979. Commonwealth Mycological Institute (Commonwealth Agricultural Bureaux) Farnum House, Farnum Royal, Slough, SL2 3BN England. 57 pages. £2.50 (£3.00 overseas).

This book consists of the papers presented at a Symposium organized by the UK Federation for Culture Collections in Newcastle 1979. The authors are leading authorities in their respective fields and have produced a very useful inexpensive reference guide to the Stability of Selected micro-organisms, including bacteria, brewing yeasts, actinomyces and fungi.

Some points are worthy of comment; first, there are few typographical errors and generally the text is easy to read but the writing styles are different. Secondly, there is no consistency about bibliography, some authors did not include any references, whereas some papers had a comprehensive literature list. Thirdly, it is apparent that there are still problems and differences of microbial stability. For example, Walker states that manufacturers have devised methods to ensure stability of vaccine-producing strains and stability does not present a serious problem. Whereas Chater points out that there is little, if any, ability to control 'instability' despite some knowledge of genetic instability and gene cloning systems. He further suggested that perhaps more additional insight into molecular mechanisms would help with understanding 'instability'. When discussing yeasts, Gilliland made the very valid point that there are many strain differences of Saccharomyces cerevisiae. Therefore, as many strains as possible should be thoroughly tested and stored. He further stressed that strains should be obtained from a wide range of sources. These comments could well apply to other yeasts as well as many other micro-organisms.

To sum-up this book despite the limitation oflack of references for some contributions is good value for money. It should serve both Student and Practising Microbiologist with an insight into stability of some micro-organisms. In addition, specialists from other fields (Biochemists and Technologists) might benefit from knowing of this volume.

R.R. Davenport \

27

THE BIODETERIORATION CENTRE

SHORT COURSE PROGRAMME 1981

6th-10thApri/1981 B.C. Course VC811 British Pest Control Association, 3rd Operator Course A course for operators and supervisors in pest control. An approved course which is required for eventual BPCA Certification. Applications for places to be made via BPCA.

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. organisms, insects {including termites), rodents and higher plants, and their effects on a wide range of materials of economic importance. The course will be of interest to research students, industrial technical staff and all who wish to explore the field of materials biology. One week residential course. Total cost£150.00

17th-20th August 1981 B.C. Course VC 814 Safety and Handling of Microorganisms Module 1 - 17/18 August (Basic) A two module course Module 2- 19/20 August (Intermediate) Module 1. Basic laboratory procedures, handling of organisms and use of safety equipment. Mainly practical work.

Module cost £120.00 Module 2. Further instruction on safety equipment, lectures on organisms, safety and legislation and regulations.

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Both modules as one course. £200.00

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All fees include accommodation and meals but not travel

More detailed programmes and booking forms for these courses can be obtained from the Biodeterioration Centre.

The Centre reserves the right to cancel any of these courses if there are insufficient bookings to run the course economically.

For further details of these courses please contact:

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28

Telephone 021 . 328 5950

LITERATURE COLLECTION

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lnternational Biodeterioration Bulletin (IBB) A scientific journal for publication of original works, including reviews and book reviews on all aspects of biodeterioration and biodegradation. Also contains the Biodeterioration Society Newsletter and short abstracts of papers presented at meetings of the Society in Great Britain and Ireland.

Biodeterioration Research Titles (BRT) A bibliographic journal which presents, in classified form, references to pub· fished literature on all aspects of biodeterioration and biodegradation. About 2000 references per annum.

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