Sains Malaysiana 50(2)(2021): 461-473

http://dx.doi.org/10.17576/jsm-2021-5002-17

Aqueous Polyphenol Semi-Purified Fractions from Galls of Quercus infectoria Increase ALP Activity and Mineralisation of hFOB 1.19 Cells

(Fraksi Separa Tulen Polifenol Akueusdaripada Puru Quercus infectoria Meningkatkan Aktiviti ALP dan Pemineralan Sel hFOB 1.19)

HERMIZI HAPIDIN¹*, FAIRUZA MUNIRAH MAZLAN¹, HASMAH ABDULLAH¹ & IMA NIRWANA SOELAIMAN²

¹Biomedicine Programme, School of Health Sciences, Universiti Sains Malaysia, Health Campus, 16150 Kubang Kerian, Kelantan Darul Naim, Malaysia

²Department of Pharmacology, Universiti Kebangsaan Malaysia Medical Centre, Jalan Yaacob Latif, Bandar Tun Razak, 56000 Cheras, Kuala Lumpur, Malaysia

Diserahkan: 14 April 2020/Diterima: 5 Ogos 2020

ABSTRACT

The galls of Quercus infectoria (QI) have been reported to possess numerous medicinal values and give a positive effect on bone metabolism. This study investigated the effects of semi-purified fractions of QI gall extract on the proliferation, alkaline phosphatase (ALP) activity, and mineralisation of human foetal osteoblast cell line (hFOB 1.19). The semi-purified fractions (fraction A and B) were prepared by column chromatography methods. MTT assay was used to measure cell proliferation activity to obtain half maximal concentration (EC₅₀) of the cells treated with fraction A (phenolic components and contained amide), fraction B (phenolic components with the presence of alkene), and pamidronate (drug control). The most potent or lowest EC₅₀ was further used to measure ALP activity in the treated and untreated cells at day 1, 3, 7, 10, and 14 by ELISA. Cell mineralisation was determined by von Kossa staining for phosphate depositions and Alizarin Red S staining for calcium depositions. The EC₅₀ values for fraction A and B were 8.86 and 9.92 μg/mL, respectively, which showed a greater effect compared to the pamidronate (15.27 μg/mL). The ALP activity of both fractions in the treated cells were also greater compared to the two control groups (cells treated with pamidronate and untreated cells), starting from day 3 onwards. The calcium depositions appeared as red spots, while phosphate depositions appeared as black spots. Interestingly, the calcium depositions of cells treated with both fractions were higher than those of the two control groups. In conclusion, semi-purified fractions of QI gall extract enhanced proliferation, improved mineralisation, and increased ALP activity of hFOB 1.19 cells.

Keywords: Alizarin Red S; alkaline phosphatase (ALP); hFOB 1.19 cell line; Quercus infectoria; von Kossa

ABSTRAK

Puru Quercus infectoria (QI) telah dilaporkan mempunyai nilai perubatan yang banyak dan memberi kesan positif pada metabolisme tulang. Penyelidikan ini dijalankan untuk mengkaji kesan fraksi separa tulen QI pada proliferasi, aktiviti alkalin fosfatase (ALP) dan pemineralan sel osteoblas fetal manusia (hFOB 1.19). Fraksi separa tulen QI (fraksi A dan B) telah disediakan melalui kaedah kromatografi kolum. Asai MTT digunakan untuk mengukur aktiviti proliferasi sel bagi menentukan kepekatan separa maksimum (EC₅₀) sel yang dirawat dengan fraksi A (komponen fenolik dan mengandungi amida), fraksi B (komponen fenolik dengan kehadiran alkena) dan pamidronat. Nilai EC₅₀ paling poten atau rendah yang diperoleh digunakan untuk mengukur aktiviti ALP dalam sel yang dirawat dan juga sel yang tidak dirawat pada hari 1, 3, 7, 10 dan 14 melalui ujian ELISA. Pemineralan sel ditentukan dengan pewarnaan von Kossa bagi pemendapan fosfat dan pewarnaanAlizarin Red S bagi pemendapan kalsium. Nilai EC₅₀ bagi sel yang dirawat dengan fraksi A fraksi B masing-masing adalah 8.86 dan 9.92 μg/mL yang menunjukkan kesan lebih hebat berbanding pamidronat (15.27 μg/mL). Kedua-dua fraksi juga menunjukkan aktiviti ALP yang lebih tinggi berbanding kedua-dua kumpulan kawalan (sel yang dirawat dengan pamidronat dan sel yang tidak dirawat), bermula pada hari ke-3 dan hari-hari berikutnya. Pemendapan kalsium kelihatan sebagai tompokan merah manakala pemendapan fosfat kelihatan tompokan hitam. Menariknya, pemendapan kalsium oleh sel yang dirawat dengan kedua-dua fraksi adalah lebih tinggi berbanding dua kumpulan kawalan. Kesimpulannya, fraksi separa tulen QI menambahkan proliferasi, memperbaiki pemineralan dan meningkatkan aktiviti ALP sel hFOB 1.19.

Kata kunci: Alizarin Red S; alkalin fosfatase (ALP); Quercus infectoria; sel hFOB 1.19; von Kossa

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*Pengarang untuk surat-menyurat; email: hermizi@usm.my