Formation and Bioremediation Performance of Freeze-Dried Pseudomonas aeruginosa and Bacillus subtilis Co-Application Using Concurrent and Sequential Methods for Diesel Contaminated Liquid Media

Kartika, Jasmine (2026) Formation and Bioremediation Performance of Freeze-Dried Pseudomonas aeruginosa and Bacillus subtilis Co-Application Using Concurrent and Sequential Methods for Diesel Contaminated Liquid Media. Other thesis, Institut Teknologi Sepuluh Nopember.

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Abstract

Pencemaran diesel di perairan merupakan masalah lingkungan yang serius karena bersifat toksik dan sulit dihilangkan. Metode pembersihan konvensional sering kali mahal dan dapat menyebabkan pencemaran sekunder, sehingga bioremediasi menjadi alternatif yang lebih berkelanjutan. Pseudomonas aeruginosa dan Bacillus subtilis merupakan bakteri yang efektif dalam mendegradasi diesel, tetapi penerapannya terbatas karena stabilitasnya yang rendah. Freeze-drying membantu mempertahankan bakteri tersebut untuk penggunaan jangka panjang. Penelitian ini mengevaluasi pengaruh metode inokulasi konkuren dan sekuensial terhadap kelangsungan hidup P. aeruginosa dan B. subtilis hasil freeze-drying serta kemampuannya dalam menurunkan Total Petroleum Hydrocarbon (TPH) pada media cair yang terkontaminasi diesel. Kultur murni kedua bakteri dikultivasi, dicampur dengan liofiloprotektan, kemudian menjalani proses freeze-drying sebelum direaktivasi. Percobaan biodegradasi dilakukan menggunakan reaktor Erlenmeyer 250 mL yang berisi Mineral Salt Medium (MSM) dengan 2% diesel selama 14 hari. Konsentrasi TPH, populasi bakteri (CFU/mL), pH, dan suhu diamati pada Hari ke-0, 3, 7, dan 14. Konsentrasi TPH dianalisis menggunakan metode gravimetri, sedangkan pertumbuhan bakteri diukur dengan metode Total Plate Count (TPC). Perlakuan sekuensial menghasilkan penyisihan TPH tertinggi, yaitu sebesar 88%, sedangkan perlakuan konkuren mencapai 73%. Reaktor blanko hanya menunjukkan penurunan TPH sebesar 24%, yang mengindikasikan bahwa degradasi hidrokarbon terutama disebabkan oleh aktivitas bakteri. Uji One-Way ANOVA menunjukkan bahwa metode inokulasi berpengaruh signifikan terhadap konsentrasi TPH (p < 0,001), sedangkan uji Tukey HSD menunjukkan tidak terdapat perbedaan yang signifikan antara perlakuan sekuensial dan konkuren (p > 0,05), meskipun keduanya berbeda signifikan dibandingkan blanko. Perlakuan konkuren mencapai populasi bakteri yang sedikit lebih tinggi dibandingkan perlakuan sekuensial pada Hari ke-7. Analisis korelasi Pearson menunjukkan hubungan yang lemah, negatif, dan tidak signifikan antara TPC dan TPH, yang menunjukkan bahwa jumlah bakteri saja tidak menentukan tingkat biodegradasi. Disimpulkan bahwa ko-aplikasi P. aeruginosa dan B. subtilis hasil freeze-drying berhasil direaktivasi pada media cair yang terkontaminasi diesel, dan metode inokulasi memengaruhi degradasi TPH setelah 14 hari.
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Diesel pollution in water is a serious environmental problem because it is toxic and difficult to remove. Conventional cleanup methods are often expensive and cause secondary pollution, making bioremediation a more sustainable alternative. Pseudomonas aeruginosa and Bacillus subtilis are effective bacteria for degrading diesel, but their application is limited by poor stability. Freeze-drying helps preserve these bacteria for long-term use. This study evaluated the effects of concurrent and sequential inoculation methods on the survival of freeze-dried P. aeruginosa and B. subtilis and their ability to reduce TPH in diesel-contaminated liquid media. Pure cultures of both bacteria were cultivated, mixed with lyoprotectants, and freeze-dried before reactivation. Biodegradation experiments were conducted in 250 mL Erlenmeyer reactors containing Mineral Salt Medium (MSM) with 2% diesel for 14 days. TPH concentration, bacterial population (CFU/mL), pH, and temperature were monitored on Days 0, 3, 7, and 14. TPH was determined by gravimetric analysis, while bacterial growth was measured using the Total Plate Count (TPC) method. The sequential treatment achieved the highest TPH removal, reducing TPH by 88%, while the concurrent treatment achieved 73% removal. The blank reactor showed 24% TPH reduction, indicating that hydrocarbon degradation was mainly because of bacterial activity. One-Way ANOVA showed that inoculation treatment significantly affected TPH concentration (p < 0.001), while Tukey HSD found no significant difference between the sequential and concurrent treatments (p > 0.05), although both differed significantly from the blank. The concurrent treatment reached a slightly higher bacterial population than the sequential treatment on Day 7. Pearson correlation showed a weak, negative, and non-significant relationship between TPC and TPH, indicating that bacterial abundance alone did not determine biodegradation. It is concluded that the freeze-dried co-application of P. aeruginosa and B. subtilis was successfully reactivated in diesel-contaminated liquid media and that the inoculation method influenced TPH degradation after 14 days.

Item Type:	Thesis (Other)
Uncontrolled Keywords:	Bioremediasi, Efisiensi Penurunan TPH, Gravimetri, Liofilisasi, TPC, Bioremediation, Gravimetric Analysis, Lyophilization, TPC Analysis, TPH Removal Efficiency
Subjects:	Q Science > QR Microbiology Q Science > QR Microbiology > QR74.8 Bacteria T Technology > TD Environmental technology. Sanitary engineering > TD192.5 Bioremediation
Divisions:	Faculty of Civil, Planning, and Geo Engineering (CIVPLAN) > Environmental Engineering > 25201-(S1) Undergraduate Thesis
Depositing User:	Jasmine Kartika
Date Deposited:	22 Jul 2026 06:08
Last Modified:	22 Jul 2026 06:08
URI:	http://repository.its.ac.id/id/eprint/136225

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