- Original article
- Open Access
Mould-resistance of bamboo treated with the compound of chitosan-copper complex and organic fungicides
© The Japan Wood Research Society 2012
- Received: 20 February 2011
- Accepted: 5 September 2011
- Published: 18 January 2012
Bamboo has received increasing attention as an alternative raw material for wood in the late twentieth century for its fast growing nature and good mechanical properties. But bamboo is readily discolored by mould fungi, which greatly limits the applications of bamboo. In this paper, mould-resistance of moso bamboo treated with chitosan-copper complex (CCC®), propiconazole (PPA), tebuconazole (TBA), the compound of CCC® and PPA or TBA was reported. Results showed that CCC® or PPA used alone as bamboo-mould inhibiter could defer or restrain the growth of Penicillium citrinum Thom (P. citrinum), while not being effective against Trichoderma viride Pers. ex Fr (T. viride) and Aspergillus niger V. Tiegh (A. niger). However, the compound of CCC® and PPA could inhibit all mould fungi in the test, showing a good synergetic effect. Additionally, TBA had better resisting effect against P. citrinum and T. viride than against A. niger, but showed no synergetic effect with CCC®.
- Chitosan-copper complex
- Mould-resistant effect
- Organic fungicides
- Synergistic effect
The ecological crisis facing the world brings sustainable development ideas to the people. A lot of countries imposed restrictions on felling natural forests, which caused the diminishing of wood resources . Therefore, the world’s attention is focused on the need to identify substitute materials. In view of its rapid growth (exceeding most fast growing trees) and wide availability, bamboo emerges as a very suitable alternative [2, 3]. Bamboo products have found more and more applications in daily necessities, furniture, crafts, indoor decoration, constructions, and so on [4–6]. With the development of adhesive bonding and processing technology, strand woven outdoor flooring has drawn more and more companies’ attentions in China, and the products were loved in the United States, Europe, Southeast Asia, and so on [7, 8]. But as a natural material, bamboo is readily deteriorated by mould fungi, decay fungi, and insects, especially mould fungi, which greatly limits the application of bamboo, leading to much more economic losses and wastage of bamboo resources.
To protect bamboo from mould fungi, sodium pentachlorophenate (PCP-Na) was widely used for its high effectiveness and relative low cost, but faced rejection due to the public concerns on the environment. The implementation of the Stockholm Convention on Persistent Organic Pollutants in 2004 further limited the production and application of PCP-Na . The PCP-Na alternatives used widely are mainly from the successful wood preservatives, such as chromated copper arsenate (CCA), amine copper quat (ACQ) or copper azole (CA). But concerns on the leaching of copper from treated wood to the environment and their bad effects against mould fungi limited the application of these preservatives [10–12]. Non-metallic and organic preservatives such as borax, 2-decyl dimethyl ammonium chloride (DDAC), propiconazole, and tebuconazole 3-iodo-2-propyl-butyl carbamate (IPBC), and so on are developed or modified as mould-resistant chemicals. However, considerable research is still under way regarding the effectiveness against mould fungi and higher cost compared with PCP-Na.
Chitosan (deacetylated chitin) has a broad spectrum of unique biological activities [13–15]. Chitosan-copper complex (CCC®), produced from the chemical reactions between chitosan and copper salts, was shown to be a low-toxic preservative for wood and bamboo [16–21]. Bamboo treated with CCC® had better resistance against Penicillium citrinum Thom and Trichoderma viride Pers. ex Fr, but poor against Aspergillus niger V. Tiegh . In order to improve the resistance of CCC® against a wide variety of the mould fungi, a compound of CCC® and organic fungicides was prepared and tested under laboratory conditions.
Moso bamboo (Phyllostachys pubescens Mazel ex H. de Lehaie) was selected due to its popularity, abundance, and value in the world. Bamboo samples, 4 years old, were collected from Sankou town, Lin’an city, Zhejiang, China, on May 17th, 2009. A segment of 2–4 m of bamboo from bottom, with green and yellow faces planed off, were chosen and machined into specimens without wormhole and mildew. The dimension was 50 mm (length) by 20 mm (width) by 5 mm (thickness). The specimens were then weighed, measured, and marked.
Three kinds of mould fungi including Trichoderma viride Pers. ex Fr (T. viride), P. citrinum Thom (P. citrinum) and Aspergillus niger V. Tiegh (A. niger), separated from natural mildew bamboo by microorganism group of Zhejiang A&F University, were applied in mould-resistance test. These fungi are common species on bamboo . Fungal resistant test was conducted referring to “Standard Method for Testing Fungicides for Controlling Sapstain and Mould on Unseasoned Lumber”  and “Testing method for anti-mould chemicals in controlling mould and blue stain fungi on wood” .
Chitosan-copper complex (CCC®, weight ratio of copper ion 11.8%) was prepared by reacting copper chloride with chitosan at the ratio of 1.2:1.0, 50°C for 2 h. CCC®–propiconazole or CCC®–tebuconazole complexes were prepared by compounding CCC® with propiconazole or tebuconazole at the ratio of CCC® to organic fungicide being 1:0.5. Propiconazole and tebuconazole were prepared in water solution with the help of nonionic surfactants.
Potato dextrose agar (PDA) substrate was used as culture media in the cultivation of mould fungi. For the preparation of PDA substrate, 200 g peeled potato was cut into small pieces, and put into 1000 ml boiling water. After 30 min, the solution was filtrated quickly into 1000 ml beaker. Then water was complemented to the filtrate until 1000 ml, and 20 g glucose, 25 g agar was put into the solution. The solution was heated until the agar was melted. Thus the PDA substrate was prepared. PDA substrate was then poured evenly into petri dishes while it was hot.
Steam sterilizer was used to sterilize the specimens and petri dishes filled with PDA substrate at 121°C, 0.1 MPa for 30 min. Incubation cabinet set at a temperature of 28 ± 2°C, and a relative humidity between 90 ± 5%, was used to incubate fungi. Petri dishes with the size of 100 mm in diameter and 20 mm in height were used in mould-resistance tests.
Treatment of specimens
Six specimens per concentration of a fungicide were used for each fungus tested. Also, 6 untreated control specimens were used for each fungus tested.
The specimens to be treated with the same concentration of the same fungicide were put into 500 ml beaker, and a piece of glass was placed between the specimens to avoid clinging. After 30 min dipping in fungicide, the specimens were taken out, wiping out the superficial fungicide. The treated specimens were autoclaved at 121°C, 0.1 MPa for 30 min before inoculation.
Inoculation and incubation
Test fungi were activated by inoculating the test fungi onto test tubes filled with PDA substrate, and cultivating for several days at the temperature of 28 ± 2°C and relative humidity of 90 ± 5%. Purified water was prepared by infusing 10 ml water into test tube, stuffing with cotton plug, and wrapping in kraft, enlacing with cotton string, then sterilizing at 121°C, 0.1 MPa for 30 min.
The mycelium and spore were picked and put carefully into purified water with inoculation loop. The spore suspension was stirred frequently during inoculation. 0.25 ml spore suspension was streaked into the petri dishes filled with PDA substrate, and was scratched evenly on the substrate. The petri dishes were then placed into incubation cabinet, maintaining at a temperature of 28 ± 2°C, and relative humidity of 90 ± 5%. After 4–7 days, the fungi in petri dishes grew, and prepared to inoculate the specimens.
Evaluation of the test
Standard method for rating the infection value
The infection value
Mould coverage on specimens
The surface of specimens have no mycelium
The area of mould infection <25%
The area of mould infection 25–50%
The area of mould infection 50–75%
The area of mould infection >75%
The amount of chemicals absorbed in bamboo blocks
Different from wood, bamboo lacks radial transport systems, which has a significant influence on the efficacy of treatments. Therefore, the fungicides are mainly distributed on the surface of bamboo, and the absorption is closely related to the concentration and molecular weight of the fungicides .
Resistances of different fungicides against mould fungi
Mould-resistant effects of different fungicides against P. citrinum
Comparing the resistance of CCC® and PPA alone with that of the compound of CCC® and PPA (CCC®–PPA), one can find that the resistance of CCC®–PPA against P. citrinum improved significantly (Fig. 3). The initial infection of P. citrinum appeared after 25 days for CCC®–PPA treated bamboo, postponed 7–14 days comparing with CCC® or PPA-treated ones, and the final infection value was only 0.67. This phenomenon might result from a synergistic effect of the two fungicides. The resistance of CCC®–TBA was between CCC® and TBA, mainly from TBA, not the synergistic effect of both. The synergistic effect of organic fungicides with antioxidant or metal chelator has been reported in a number of literature [26–29]. CCC®, prepared by the reaction of chitosan and copper salts, may act as metal chelator which enhances the resistance of PPA against mould fungi. This inference can also be proved by the follows.
Mould-resistant effects of different fungicides against T. viride
Mould-resistant effect of different fungicides against A. niger
Results analyzed above showed that the organic fungicides (including PPA and TBA) resisted mould fungi more effectively than CCC®. Among the test mould fungi, A. niger was the most serious one, only the compound of CCC® and PPA or higher concentrations of PPA could restrain or inhibit it. The significant resistance of CCC®–PPA indicated that the two fungicides had good synergistic effect. However, CCC®–TBA, at the concentration of 1%, only had certain resistance against P. citrinum and T. viride, and the effects were mainly attributed to TBA, not to the synergistic effect of CCC® and TBA.
Overall appraisal of the test fungicides
Overall appraisal of the fungicides against the test fungi
Infection value of treated bamboob
Resisting effectiveness (%)
As could be seen from Table 2, CCC®, TBA, and CCC®–TBA had lower RE, even at the concentration of 2%, CCC®–TBA showed no synergistic effect against the test fungi. RE of PPA was higher than TBA, CCC® and CCC®–TBA, but when used alone, could not resist all of the three test fungi effectively. The compound of CCC® and PPA had the highest resistance against all of the test fungi, even at low concentrations, showing synergistic effect.
Mould-resistant effect of CCC®, PPA, TBA, CCC®–PPA, and CCC®–TBA against P. citrinum, T. viride, and A. niger indicated that CCC® or PPA used alone could not resist the test fungi effectively, but the compound of CCC® and PPA behaved the best against all of the test fungi, even at low concentrations, showing a good synergistic effect. However, no synergistic effect could be inferred from the resisting results of CCC®–TBA against the test fungi. Further tests on field are recommended for the outdoor applications of bamboo treated with these fungicides.
This research was supported by the International Foundation for Science (No. D/3284-2), the National Natural Science Foundation of China (No. 30972304) and Science and Technology Department of Zhejiang Province of China (No. 2009C32065). The authors thank Hangzhou bamfox bamboo products co., Ltd, for their kind help in the preparation of bamboo samples.
- Li WC (2000) Analysis on world forest resources protection and China’s forestry development polices. Resour Sci 22:71–76Google Scholar
- Du Y, Yu XJ (2008) The development and comparison of world’s bamboo industry. World Agric 7:18–21Google Scholar
- De Flander K, Rovers R (2009) One laminated bamboo-frame house per hectare per year. Constr Build Mater 23:210–218View ArticleGoogle Scholar
- Liese W (1987) Research on bamboo. Wood Sci Technol 21:189–209View ArticleGoogle Scholar
- Scurlock JMO, Dayton DC, Hames B (2000) Bamboo: an overlooked biomass resource? Biomass Bioenergy 19:229–244View ArticleGoogle Scholar
- Vesey DG (2002) Structural design of the bamboo pavilion, Berlin. In: Proceedings of the international conference on advances in building technology, 4–6. Hong Kong, China, pp 653–660Google Scholar
- Nugroho N, Ando N (2001) Development of structural composite products made from bamboo II: fundamental properties of laminated bamboo lumber. J Wood Sci 47:237–242View ArticleGoogle Scholar
- Van der Lugt P, van den Dobbelsteen AAJF, Janssen JJA (2006) An environmental, economic and practical assessment of bamboo as a building material for supporting structures. Constr Build Mater 20:648–656View ArticleGoogle Scholar
- The United Nations Environment Program (2004) The Stockholm convention on persistent organic pollutions. Stockholm ConventionGoogle Scholar
- Rapp AO, Brandt K, Peek RD, Schmitt U (1997) Quantitative measurement and chemical analysis of wood dust collected in German woodworking companies. Holz Roh Werkst 55:141–147View ArticleGoogle Scholar
- Tascioglu C, Cooper P, Ung T (2009) Effects of delayed drying and CO2 application on copper amine fixation in ACQ treated red pine. Eur J Wood Prod 67:7–12View ArticleGoogle Scholar
- Mourant D, Yang DQ, Lu X, Riedl B, Roy C (2009) Copper and boron fixation in wood by pyrolytic resins. Bioresour Technol 100:1442–1449PubMedView ArticleGoogle Scholar
- Kumar MNVR (2000) A review of chitin and chitosan applications. React Func Polym 46:1–27View ArticleGoogle Scholar
- Chirkov SN (2002) The antiviral activity of chitosan (review). Appl Biochem Microbiol 3(8):1–8View ArticleGoogle Scholar
- Chung YC, Chen CY (2008) Antibacterial characteristics and activity of acid-soluble chitosan. Bioresour Technol 99:2806–2814PubMedView ArticleGoogle Scholar
- Kobayashi T, Furukawa I (1995) Optimum conditions for the formation of chitosan–metal salts and their fixation in wood. J Antibact Antifung Agents 23:263–269Google Scholar
- Kobayashi T, Furukawa I (1995) Wood-preserving effectiveness of chitosan–metal salts against wood decaying fungi. J Antibact Antifung Agents 23:343–348Google Scholar
- Eikenes M, Fongen M, Roed L, Stenstrøm Y (2005) Determination of chitosan in wood and water samples by acidic hydrolysis and liquid chromatography with online fluorescence derivatization. Carbohydr Polym 61:29–38View ArticleGoogle Scholar
- Sun FL, Duan XF, Mao SF, Wen GF (2007) Decay resistance of bamboo wood treated with chitosan-metal complexes against the white-rot fungus Coriolous versicolor. Sci Silvae Sin 43:82–87Google Scholar
- Sun FL, Duan XF, Wen GF, Mao SF (2006) Anti-mould effects of CMC wood preservatives on bamboo wood. Sci Silvae Sin 42:40–43Google Scholar
- Kobayashi T, Furukawa I (1996) Antifungal effects of chitosan-metal salt. J Antibact Antifung Agents 24(3):191–193Google Scholar
- Wu KY, Weng YX (2000) Bamboo mildew-rotting and its relation with environmental condition. For Res 13:63–70Google Scholar
- ASTM: D4445-91 (1994) Standard method for testing fungicides for controlling sapstain and mould on unseasoned lumber (laboratory method), pp 495–498 (reapproved 1994)Google Scholar
- GB/T 18261-2000. Testing method for anti-mould chemicals in controlling mould and blue stain fungi on woodGoogle Scholar
- Lee AWC, Chen G, Tainter FH (2001) Comparative treatability of Moso bamboo and Southern pine with CCA preservative using a commercial schedule. Bioresour Technol 77:87–88PubMedView ArticleGoogle Scholar
- Goodell B, Nicholas DD, Schultz TP (2003) Wood deterioration and preservation advances in our changing world. ACS symposium series 845. Washington, USA, pp 378–398View ArticleGoogle Scholar
- Schultz TP, Nicholas DD (2002) Development of environmentally benign preservatives based on the combination of organic biocides with antioxidants and metal chelators. Phytochemistry 61:555–560PubMedView ArticleGoogle Scholar
- Mabicka A, Dumarcay S, Rouhier N, Linderc M, Jacquot JP, Gérardin P, Gelhaye E (2005) Synergistic wood preservatives involving EDTA, Irganox 1076 and 2-hydroxypyridine-N-oxide. Int Biodeterior Biodegrad 55:203–211View ArticleGoogle Scholar
- Green F, Kuster TA, Highley TL (1997) Targeted inhibition of wood decay. IRG/WP-10203. International Research Group on Wood PreservationGoogle Scholar