Malaysian Journal of Analytical Sciences Vol 24 No 5 (2020): 649 - 656

 

 

 

 

ANALYSES ON TOXICITY OF Pb2+ TOWARDS CHLOROPHYLL A, TOTAL SOLUBLE PROTEIN AND CASPASE-3-LIKE ENZYME ACTIVITY OF Scenedesmus regularis

 

(Analisa Kesan Ketoksikan Pb2+ Terhadap Klorofil A, Jumlah Protein Terlarut dan Aktiviti  Enzim Bak Kaspase-3 Scenedesmus regularis)

 

Hazlina Ahamad Zakeri1, 3, Nakisah Mat Amin1, Nur Hidayah Kamilia Rassman1, Wan Bayani Wan Omar1, 2*

 

1Faculty of Science and Marine Environment,

2Institute of Marine Biotechnology,

3Biological Security and Sustainability (BioSeS) Research Group,

Faculty of Science and Marine Environment,

Universiti Malaysia Terengganu, 21030 Kuala Nerus, Terengganu, Malaysia

 

*Corresponding author:  bayani@umt.edu.my

 

 

Received: 24 December 2019; Accepted: 28 July 2020; Published:  12 October 2020

 

 

Abstract

Heavy metal pollutions, including lead (Pb), has become an increasing concern to humans due to their adverse effects and the fact that they are not easily degraded or destroyed. Microalgae are aquatic organisms that can be used in metal bioremediation since they can accumulate and detoxify metals. This study reported on the responses of a freshwater microalga, which was Scenedesmus regularis, when exposed to the inhibitory concentrations (IC) of Pb2+ at 25%, 50% and 75%. The tolerance level of S. regularis against Pb2+ at IC25, IC50 and IC75 was determined to be 3.5 mg/L, 7.2 mg/L and 10.9 mg/L, respectively. Then, the microalga was treated with these inhibitory concentrations. The concentration of Chlorophyll A (Chl A) and total soluble protein (TSP), as well as caspase-3-like enzyme activity of the alga were analysed. It was observed that Chl A concentration of the alga significantly decreased as more cells were inhibited by Pb2+. The highest concentration of Pb2+ significantly reduced the TSP concentration of the alga. However, no changes were observed amongst the concentrations of Pb2+, which inhibited 25% and 50% of the alga population. Activity of caspase-3-like enzyme was significantly induced by more than 3-fold of control in IC25 of Pb2+, while the activity of this enzyme was observed to be supressed in both the IC50 and IC75. In conclusion, the alga has the potential to be a good indicator for Pb2+ toxicity and the Chl A concentration and caspase-3-like enzyme activity can be applied as biomarkers.

 

Keywords:  lead(II) ion, metal toxicity, microalgae, biochemical analyses, inhibitory concentration  

 

Abstrak

Pencemaran logam berat termasuk plumbum menjadi semakin menarik perhatian manusia akibat kesan buruknya dan hakikat bahawa ia tidak mudah diurai atau dimusnah. Mikroalga ialah organisma akuatik yang boleh digunakan dalam bioremediasi logam kerana ia boleh mengumpul dan menyahtoksik logam. Kajian ini melaporkan tentang tindak balas mikroalga air tawar, iaitu Scenedesmus regularis apabila didedahkan kepada ujian toksik Pb2+  pada kepekatan rencatan (IC) 25%, 50% dan  75%. Tahap toleransi S. regularis terhadap Pb2+ pada IC25, IC50 dan IC75 ialah masing-masing pada 3.5 mg/L, 7.2 mg/L dan 10.9 mg/L. Kemudian mikroalga dirawat dengan kepekatan ini dan kepekatan Klorofil A. Jumlah protein terlarut (TSP) serta aktiviti enzim bak kaspase-3 dianalisis. Didapati bahawa kepekatan Klorofil A alga berkurangan apabila sel-sel lebih banyak direncat oleh Pb2+. Kepekatan tertinggi Pb2+ mengurangkan kepekatan TSP alga dengan ketara. Namun, tiada perubahan diperhatikan antara kepekatan Pb2+ yang merencat 25% dan 50% populasi alga. Aktiviti enzim bak kaspase-3 telah meningkat lebih daripada tiga kali ganda dalam IC25 Pb2+ berbanding kawalan, manakala aktiviti enzim ini berkurangan pada IC50 dan IC75. Kesimpulannya, alga mempunyai potensi untuk menjadi petunjuk yang baik untuk ketoksikan Pb2+ manakala kepekatan Klorofil A dan aktiviti enzim bak kaspase-3 boleh digunakan sebagai biopenanda.

 

Kata kunci:  ion plumbum(II), ketoksikan logam, mikroalga, analisa biokimia, kepekatan rencatan
 

References

1.      Kumar, K. S., Dahms, H-U., Won, E. J., Lee, J. S. and Shin, K-H. (2015). Microalgae - a promising tool for heavy metal remediation. Ecotoxicology and Environmental Safety, 113: 329-352.

2.      Cao, D.J., Shi, X.D., Li, H., Xie, P.P., Zhang, H.M., Deng, J.W. and Liang, Y.G. (2015). Effects of lead on tolerance, bioaccumulation, and antioxidative defense system of green algae, Cladophora. Ecotoxicology and Environmental Safety, 112: 231-237.

3.      Piotrowska-Niczyporuk, A., Bajguz, A., Talarek, M., Bralska, M. and Zambrzycka, E. (2015). The effect of lead on the growth, content of primary metabolites, and antioxidant response of green alga Acutodesmus obliquus (Chlorophyceae). Environmental Science and Pollution Research, 22: 19112-19123.

4.      Luqman, A. B. and Hazlina, A. Z. (2015). Accumulation of Cu(II) and Pb(II) in three rhodophytes of the genus Gracilaria and the impact of the metals on the algae physiology. Bioscience and Bioengineering, 1: 106-111.

5.      Dao, L. H. and Beardall, J. (2016). Effects of lead on growth, photosynthetic characteristics and production of reactive oxygen species of two freshwater green algae. Chemosphere, 147: 420-429.

6.      Saleh, B. (2016). Lead (Pb) heavy metal impacts in the green Ulva lactuca (Chlorophyceae) marine algae. Journal of Stress Physiology and Biochemistry, 12: 62-71.

7.      Carfagna, S., Giovanna, N. L., Adriana, S., Sorbo, B. S. and Vona, V. (2013). Physiological and morphological responses of lead or cadmium exposed Chlorella sorokiniana 211-8K (Chlorophyceae). SpingerPlus, 2: 147-152.

8.        Szivák, I., Behra, R. and Sigg, L. (2009). Metal-induced reactive oxygen species production in Chlamydomonas reinhardtii (Chlorophyceae).  Journal of Phycology, 45: 427-435.

9.      Sui, L., Zhang, R-H., Zhang, P., Yun, K-L., Zhang, H-C., Liu, L. and Hu, M-X. (2015). Lead toxicity induces autophagy to protect against cell death through mTORC1 pathway in cardiofibroblasts. Bioscience Reports, 35: 1-9.

10.   Chatterjee, S., Sarkar, S. and Bhattacharya, S. (2014). Toxic metals and autophagy. Chemical Research in Toxicology, 27: 1887-1900.

11.   Darehshouri, A., Affenzeller, M. and Lütz-Meindl, U. (2008). Cell death in unicellular green alga Micrasterias upon H2O2 induction. Plant Biology, 10: 732-745.

12.   Segovia, M. and Berges, J. A. (2009). Inhibition of caspase-like activities prevents the appearance of reactive oxygen species and dark-induced apoptosis in the unicellular chlorophyte Dunaliella tertiolecta. Journal of Phycology, 45: 1-11.

13.   Johnson, J. G., Janech, M. G. and Van Dolah, F. M. (2014). Caspase-like activity during aging and cell death in the toxic dinoflagellate Karenia brevis. Harmful Algae, 31: 41-53.

14.   Hii, Y. S., Shia, K. L., Chuah, T. S. and Hing, L. S. (2009). Physiological responses of Chaetoceros sp. and Nannochloropsis sp. to short-term 2,4-D, Dimethylamine and Endosulfan exposure. Aquatic Ecosystem & Health Management, 12: 375-389.

15.   Ritchie, R. J. (2008). Universal chlorophyll equations for estimating chlorophylls a, b, c, and d and total chlorophylls in natural assemblages of photosynthetic organisms using acetone, methanol, or ethanol solvents. Photosynthetica, 46: 115-126.

16.   Faurobert, M., Mihr, C., Bertin, N., Pawlowski, T., Negroni, L., Sommerer, N. and Causse, M. (2007). Major proteome variations associated with cherry tomato pericarp development and ripening. Plant Physiology, 143: 1327-1346.

17.   Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72: 248-254. 

18.   Bouchard, J. N. and Yamasaki, H. (2009). Implication of nitric oxide in the heat-stress-induced cell death of the symbiotic alga Symbiodinium microadriaticum. Marine Biology, 156: 2209-2220.

19.   Cenkci, S., Cigerci, I. H., YildIz, M., Ozay, C., Bozdag, A. and Terzi, H. (2010). Lead contamination reduces chlorophyll biosynthesis and genomic template stability in Brassica rapa L. Environmental and Experimental Botany, 67: 467-473.

20.   Pourraut, B., Shahid, M., Dumat, C., Winterton, P. and Pinelli, E. (2011). Lead uptake, toxicity and detoxification in plants. Reviews of Environmental Contamination and Toxicology, 213: 113-136.

21.   Shakya, K., Chettri, M. K. and Sawidis, T. (2008). Impact of heavy metals (copper, zinc and lead) on the chlorophyll content of some mosses. Environmental Contamination and Toxicology, 54: 412-421.

22.   Drazkiewicz, M. (1994). Chlorophyllase: occurrence, functions, mechanism of action, effects of external and internal factors. Photosynthetica, 30: 321-331.

23.   Gill, M. (2014). Heavy metal stress in plants: A review. International Journal of Advanced Research, 2: 1043-1055.

24.   Villiers, F., Ducruix, C., Hugouvieux, V., Jarno, N., Ezan, E. and Garin, J. (2011). Investigating the plant response to cadmium exposure by proteomic and metabolomic approaches. Proteomics, 11: 1650-1663.

25.   Cryns, V. and Yuan, J. (1998). Proteases to die for. Genes & Development, 12: 1551-1570.

26.   Wan Bayani, W. O., Hazlina, A. Z., Nakisah, M. A. and Nur Hidayah, K. R. (2017). Responses of a freshwater microalga, Scenedesmus regularis exposed to 50% inhibition concentration of Pb2+ and Hg2+. Malaysian Applied Biology, 46(4):213-220. 

27.   Yedjou, C. G., Milner, J. N., Howard, C. B. and Tchounwou, P. B. (2010). Basic apoptotic mechanisms of lead toxicity in human leukemia (HL-60) cells. International Journal of Environmental Research and Public Health, 7: 2008-2017.

28.   Wu, H., Che, X., Zheng, Q., Wu, A., Pan, K., Shao, A., Wu, Q., Zhang, J. and Hong, Y. (2014). Caspases: a molecular switch node in the crosstalk between autophagy and apoptosis. International Journal of Biological Sciences, 10: 1072-1083.

29.   Yu, L., Wan, F., Dutta, S., Welsh, S., Liu, Z. H., Freundt, E., Baehrecke, E. H. and Lenardo, M. (2006). Autophagic programmed cell death by selective catalase degradation. Proceedings of the National Academy of Sciences of the United States of America, 103: 4952-4957.

30.   Kroemer, G. and Levine, B. (2008). Autophagic cell death: the story of a misnomer. Nature Reviews Molecular Cell Biology, 9: 1004-1010.

31.   Thorburn, A. (2008). Apoptosis and autophagy: regulatory connections between two supposedly different processes. Apoptosis, 13: 1-9.

32.   Majno, G.and Joris, I. (1995). Apotosis, oncosis and necrosis: an overview of cell death. American Journal of Pathology, 146: 3-15.