Sains Malaysiana 50(10)(2021): 3085-3094

http://doi.org/10.17576/jsm-2021-5010-21

Zingiber zerumbet (L.) Smith Hexane Crude Extract Caused DNA Damage on Leptospira spp.

(Ekstrak Mentah Heksana Zingiber zerumbet (L.) Smith Menyebabkan Kerosakan DNA pada Leptospira spp.)

FARAH WAHIDA IBRAHIM¹, NURUL AMIRAH ABDUL AZIZ¹, LATIFAH IBRAHIM², NURUL FARHANA JUFRI¹ & ASMAH HAMID¹*

¹Center for Toxicology and Health Risk Studies (CORE), Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur, Federal Territory, Malaysia

²Biotechnology Unit, Institute for Medical Research, Jalan Pahang, 50588 Kuala Lumpur, Federal Territory, Malaysia

Received: 25 March 2020/Accepted: 18 February 2021

ABSTRACT

Numerous attempts have been made to control leptospirosis by using chemoprophylaxis, but with limited success. The present study was done to investigate the antileptospiral potential of hexane, ethyl acetate and methanol extracts of Zingiber zerumbet rhizomes. The extracts were assayed for antileptospiral activity using broth microdilution method against Leptospira interrogans (serovar Batavie, Canicola, Australis) and Leptospira biflexa serovar Patoc. The Z. zerumbet hexane extract exhibited antileptospiral activity, with IC₅₀ values of 248 μg/mL against L. interrogans serovar Canicola, IC₅₀ of 125 μg/mL against L. interrogans serovar Australis, IC₅₀ of 15.63 μg/mL against L. interrogans serovar Batavie and IC₅₀ of 109 μg/mL against L. biflexa serovar Patoc. However, both ethyl acetate and methanol extracts did not show any distinct antileptospiral activity. Since the hexane extract of Z. zerumbet showed antileptospiral activity, the DNA-damaging properties of this extract were tested according to their IC₅₀ and IC₂₅ values that were specific to each serovars. The DNA-damaging properties were determined by treating the selected Leptospira spp. with the hexane extract and subjecting its DNA to electrophoresis and analysis on agarose gels. The results demonstrated that the hexane extract had DNA-damaging properties towards L. biflexa serovar Patoc and L. interrogans serovar Australis, as proven by the appearance of fragmented DNA on the gels. We conclude that the Z. zerumbet hexane extract could inhibit the growth of Leptospira spp. serovar Patoc and Australis through DNA-damaging activity and thus, could be a potential antileptospiral agent. Further studies are needed to investigate the potential of this hexane extract as an antileptospiral agent using in vivo rat models of leptospirosis.

Keywords: Antileptospiral; DNA damage; growth inhibition; hexane crude extract; Zingiber zerumbet

ABSTRAK

Banyak usaha telah dibuat untuk mengawal leptospirosis dengan menggunakan kemoprofilaksis namun kejayaannya terhad. Penyelidikan ini dijalankan untuk mengkaji potensi antileptospira bagi ekstrak kasar heksana, etil asetat dan metanol rizom Zingiber zerumbet. Ekstrak telah diuji untuk aktiviti antileptospira menggunakan kaedah mikropencairan kaldu ke atas Leptospira interogans (serovar Batavie, Canicola, Australis) dan Leptospira biflexa serovar Patoc. Ekstrak heksana Z. zerumbet menunjukkan aktivti antileptospira dengan nilai IC₅₀ pada 248 μg/mL terhadap L. interrogans serovar Canicola, IC₅₀ pada 125 μg/mL terhadap L. interrogans serovar Australis, IC₅₀ pada 15.63 μg/mL terhadap L. interrogans serovar Batavie dan IC₅₀ pada 109 μg/mL untuk L. biflexa serovar Patoc. Walau bagaimanapun, kedua-dua ekstrak etil asetat dan metanol tidak menunjukkan sebarang aktiviti antileptospira yang ketara. Oleh kerana ekstrak heksana Z. zerumbet menunjukkan aktiviti antileptospira, keupayaan merosakkan DNA bagi ekstrak ini diuji mengikut nilai IC₅₀ dan IC₂₅ khusus yang diperoleh untuk setiap serovar. Hasil kajian menunjukkan bahawa ekstrak heksana mempunyai keupayaan untuk merosakkan DNA L. biflexa serovar Patoc dan L. interrogans serovar Australis seperti yang terbukti dengan kehadiran DNA yang berfragmen pada gel. Sebagai kesimpulan, ekstrak heksana Z. zerumbet boleh menghalang pertumbuhan Leptospira spp. serovar Patoc dan Australis melalui aktiviti merosakkan DNA dan dengan itu, berpotensi sebagai agen antileptospira. Kajian lanjut diperlukan untuk mengkaji potensi ekstrak heksana sebagai agen antileptospira dalam model tikus leptospirosis secara in vivo.

Kata kunci: Antileptospira; ekstrak kasar heksana; kerosakan DNA; merencat pertumbuhan; Zingiber zerumbet

REFERENCES

Abdul, A., Siddiq, I.A., Adel, S.A., Manal, M.E. & Syam, M.M. 2008. Anti-cancer and anti-microbial activities of zerumbone from the rhizomes of Zingiber zerumbet. Int. J. Pharm. 4(4): 301-304.

Abdul Hamid, Z., Budin, S.B., Ng, W.J., Hamid, A., Husain, K. & Mohamed, J. 2012. Nephroprotective effects of Zingiber zerumbet Smith ethyl acetate extract against paracetamol-induced nephrotoxicity and oxidative stress in rats. J. Zhejiang Univ. Sci. B 13(3): 176-185.

Adler, B. & de la Pena Moctezuma, A. 2010. Leptospira and leptospirosis. Vet. Microbiol. 140(3-4): 287-296.

Arzouni, J.P., Parola, P., La Scola, B., Postic, D., Brouqui, P. & Raoult, D. 2002. Human infection caused by Leptospira fainei. Emerg. Infect. 2002: 865-868.

Bahaman, A.R. & Ibrahim, A.L. 1988. A review of leptospirosis in Malaysia. Vet. Res. Commun. 12(2): 179-189.

Benacer, D., Siti, N.M.Z., Zhu Sim, S., Nizam, M.K.M.K., Galloway, R.L., Souris, M. & Thong, K.L. 2016. Determination of Leptospira borgpetersenii serovar Javanica and Leptospira interrogans serovar Bataviae as the persistent Leptospira serovars circulating in the urban rat populations in Peninsular Malaysia. Parasit Vectors 9: 117.

Benacer, D., Who, P.Y., Mohd Zain, S.N., Amran, F. & Thong, K.L. 2013. Pathogenic and saprophytic Leptospira species in water and soils from selected urban sites in Peninsular Malaysia. Microbes. Environ. 28(1): 135-140.

Cushnie, T.P. & Lamb, A.J. 2005. Antimicrobial activity of flavonoids. Int. J. Antimicrob. Agents 26(5): 343-356.

da Silva, T.M., Pinheiro, C.D., Orlandi, P.P., Pinheiro, C.C. & Pontes, G.S. 2018. Zerumbone from Zingiber zerumbet (L.) Smith: A potential prophylactic and therapeutic agent against the cariogenic bacterium Streptococcus mutans. BMC Complement Altern. Med. 18: 301.

Dive, A.M., Bigaignon, G. & Reynaert, M. 1987. Adult respiratory distress syndrome in leptospira icterohaemorrhagiae infection. Intensive Care Med. 13(3): 214.

Echeverría, J., Opazo, J., Mendoza, L., Urzúa, A. & Wilkens, M. 2017. Structure-activity and lipophilicity relationships of selected antibacterial natural flavones and flavanones of Chilean flora. Molecules 22(4): E608. doi: 10.3390/molecules22040608.

Evangelista, K.V. & Coburn, J. 2010. Leptospira as an emerging pathogen: A review of its biology, pathogenesis and host immune responses. Future Microbiol. 5(9): 1413-1425.

Faine, S., Adler, B. & Ruta, G. 1974. A mechanism of immunity to leptospirosis. Aust. J. Exp. Biol. Med. Sci. 52(2): 301-310.

Garba, B., Bahaman, A.R., Khairani-Bejo, S., Zakaria, Z. & Mutalib, A.R.  2017. Retrospective study of leptospirosis in Malaysia. EcoHealth 14(2): 389-398.

Guerra, M.A. 2013. Leptospirosis: Public health perspectives. Biologicals 41(5): 295-297.

Hamid, A., Ibrahim, F.W., Teoh, H.M., Nasrom, M.N., Eusoff, N., Husain, K. & Abdul Latif, M. 2018. Zingiber zerumbet L. (Smith) extract alleviates the ethanol-induced brain damage via its antioxidant activity. BMC Complement Altern. Med. 18(1): 101.

Hamid, A., Budin, S.B., Mohamed, R.A.P., Manaf, N.A., Yuhana, N.Y., Husain, K., Abd Hamid, Z. & Mohamed, J. 2011. Role of oxidative stress in the protective effects of Zingiber zerumbet Smith ethyl-acetate extract against paracetamol-induced hepatotoxicity in Sprague-Dawley rat. Aust. J. Basic Appl. Sci. 5(8): 1519-1525.

Haque, M.A., Jantan, I. & Harikrishnan, H. 2018. Zerumbone suppresses the activation of inflammatory mediators in LPS-stimulated U937 macrophages through MyD88-dependent NF-κB/MAPK/PI3K-Akt signaling pathways. Int. Immunopharmacol. 55: 312-322.

Husna, M., Nabilah, I. & Wan Nor Amilah, W.A.W. 2018. Anti-microbial activity of aqueous Quercus infectoria gall extract against pathogenic Leptospira. Malays. J. Med. Sci. 25(4): 42-50.

Kader, G., Nikkon, F., Rashid, M.A. & Yeasmin, T. 2011. Antimicrobial activities of the rhizome extract of Zingiber zerumbet Linn. Asian Pac. J. Trop. Biomed. 1(5): 409-412.

Kader, M.G., Habib, M.R., Nikkon, F., Yeasmin, T., Rashid, M.A., Rahman, M.M. & Gibbons, S. 2010. Zederone from the rhizomes of Zingiber zerumbet and its anti-staphylococcal activity. Boletín Latinoamericano y del Caribe de Plantas Medicinales y Aromáticas 9(1): 63-68.

Latifah, I., Abdul Halim, A., Rahmat, M.S., Faten, N.M., Ubil, Z.E., Asmah, H., Shafariatul Akmar, I., Picardeau, M., Siti Haslina, O. & Nasir, M.A. 2017. Isolation by culture and PCR identification of LipL32 gene of pathogenic Leptospira spp. in wild rats of Kuala Lumpur. Malays. J. Pathol. 39(2): 161-166.

Matsui, M., Rouleau, V., Bruyère-Ostells, L. & Goarant, C. 2011. Gene expression profiles of immune mediators and histopathological findings in animal models of leptospirosis: Comparison between susceptible hamsters and resistant mice. Infect. Immun. 79: 4480-4492.

Mérien, F., Amouriaux, P., Perolat, P., Baranton, G. & Saint Girons, I. 1992. Polymerase chain reaction for detection of Leptospira spp. in clinical samples. J. Clin. Microbiol. 30(9): 2219-2224.

Mohd Radi, M.F., Hashim, J.H., Jaafar, M.H., Hod, R., Ahmad, N., Nawi, M., Baloch, G.M., Ismail, R. & Farakhin Ayub, N.I. 2018. Leptospirosis outbreak after the 2014 major flooding event in Kelantan, Malaysia: A spatial-temporal analysis. Am. J. Trop. Med. Hyg. 98(5): 1281-1295.

Muhammad Aklil, A.R., Aznida, M.Z., Azman, A., Muhammad Haneef, A., Shaifuddin, N.H., Syed Syarizman, S.A.R., Mohammad Safree, J., Norfazilah, A. & Mohd Rohaizat, H. 2018. Effectiveness of antibiotic prophylaxis for leptospirosis among adults: A systematic review. Malays. J. Appl. Sci. 3(2): 46-56.

Murray, C.K. & Hospenthal, D.R. 2004. Determination of susceptibilities of 26 Leptospira sp. serovars to 24 antimicrobial agents by a broth microdilution technique. Antimicrob. Agents Chemother. 48(10): 4002-4005.

Musa, A.M., Ibrahim, M.A., Aliyu, A.B., Abdullahi, M.S., Tajuddeen, N., Ibrahim, H. & Oyewale, A.O. 2015. Chemical composition and antimicrobial activity of hexane leaf extract of Anisopus mannii (Asclepiadaceae). J. Intercult. Ethnopharmacol. 4(2): 129-133.

Nally, J.E., Whitelegge, J.P., Bassilian, S., Blanco, D.R. & Lovett, M.A. 2007. Characterization of the outer membrane proteome of Leptospira interrogans expressed during acute lethal infection. Infect. Immun. 75(2): 766-773.

Nelson, J., Chairman, K., Ranjit Singh, A.J.A., Padmalatha, C. & Hepsibah, B. 2013. Cytomorphological changes and inhibition of inclusion body formation in Leptospira Interrogans on treatment with the extracts of Adhatodavasica. Adv. Tech. Biol. Med. 1(1): 1-4.

Paster, B.J., Dewhirst, F.E., Weisburg, W.G., Tordoff, L.A., Fraser, G.J., Hespell, R.B., Stanton, T.B., Zablen, L., Mandelco, L. & Woese, C.R. 1991. Phylogenetic analysis of the spirochetes. J. Bacteriol. 173(19): 6101-6109.

Patel, S.S. & Savjani, J.K. 2015. Systematic review of plant steroids as potential antiinflammatory agents: Current status and future perspectives. J. Phytopharmacol. 4(2): 121-125.

Patial, P.K., Sharma, A., Kaur, I. & Cannoo, D.S. 2019. Correlation study among the extraction techniques, phytochemicals, and antioxidant activity of Nepeta spicata aerial part. Biocatalysis and Agricultural Biotechnology 20: 101275.

Picardeau, M., Bulach, D.M., Bouchier, C., Zuerner, R.L., Zidane, N., Wilson, P.J., Creno, S., Kuczek, E.S., Bommezzadri, S., Davis, J.C., McGrath, A., Johnson, M.J., Boursaux-Eude, C., Seemann, T., Rouy, Z., Coppel, R.L., Rood, J.I., Lajus, A., Davies, J.K., Médigue, C. & Adler, B. 2008. Genome sequence of the saprophyte Leptospira biflexa provides insights into the evolution of Leptospira and the pathogenesis of leptospirosis. PLoS ONE 3(2): e1607.

Prakash, V. 2017. Terpenoids as source of anti-inflammatory compounds. Asian J. Pharm. Clin. Res. 10(3): 68-76.

Saengjaruk, P., Sakolvaree, Y., Maneewatch, S., Tomanakan, K., Tongtawe, P., Tapchaisri, P. & Chaicumpa, W. 2007. Components of pathogenic Leptospira spp. with potentials for diagnosis of human leptospirosis. Asian Pac. J. Allergy Immunol. 25(4): 225-232.

Schneider, A.G., Casanovas-Massana, A., Hacker, K.P., Wunder, E.A.J., Begon, M., Reis, M.G., Childs, J.E., Costa, F., Lindow, J.C. & Ko, A.I. 2018. Quantification of pathogenic Leptospira in the soils of a Brazilian urban slum. PLoS Negl. Trop. Dis. 12(4): e0006415.

Schneider, M.C., Jancloes, M., Buss, D.F., Aldighieri, S., Bertherat, E., Najera, P., Galan, D.I., Durski, K. & Espinal, M.A. 2013. Leptospirosis: A silent epidemic disease. Int. J. Environ. Res. Public Health 10(12): 7229-7234.

Shafariatul Akmar, I., Syahirah, S., Fifi Fariza, A., Asmah, H., Latifah, I. & Dayang Fredalina, B. 2019. In vitro antileptospiral activity of Canarium odontophyllum Miq. (Dabai) leaves extract. Malays. J. Microb. 15(3): 220-225.

Vinod, K.K., Lall, C., Vimal, R.R., Vedhagiri, K., Sunish, I.P. & Vijayachari, P. 2018. Can subminimal inhibitory concentrations of antibiotics induce the formation of biofilm in Leptospira? Microb. Drug Resist. 24(7): 1040-1042.

Voravuthikunchai, S.P., Phongpaichit, S. & Subhadhirasakul, S. 2005. Evaluation of antibacterial activities of medicinal plants widely used among aids patients in Thailand. Pharm. Biol. 43(8): 701-706.

Wong, S.K., Lim, Y.Y. & Chan, E.W.C. 2010. Evaluation of antioxidant, anti-tyrosinase and antibacterial activities of selected Hibiscus species. Etnobotanical Leaflets 14: 781-796.

Yob, N.J., Jofrry, S.M., Affandi, M.M.L.K., Salleh, M.Z. & Zakaria, Z.A. 2011. Zingiber zerumbet (L.) Smith: A review of its ethnomedicinal, chemical, and pharmacological uses. Evid. Based Complement Alternat. Med. 2011: 543216.

Zaki, A.M., Rahim, M.A.A., Azme, M.H., Mahmood, N.A., Jeffree, M.S., Ghazi, H.F. & Hassan, M.R. 2018. Animal reservoirs for Leptospira spp. in South-East Asia: A meta-analysis. J. Adv. Res. Med. 5(3): 23-31.

Zhang, W., Xie, X., Wu, D., Jin, X., Liu, R., Hu, X., Fu, Y., Ding, Z., Zhang, N. & Cao, Y. 2017. Doxycycline attenuates leptospira-induced il-1β by suppressing nlrp3 inflammasome priming. Front Immunol. 8: 857.

*Corresponding author; email: asmah0901@ukm.edu.my