Pressurized Cyclic Solid–Liquid (PCSL) Extraction of Sea Buckthorn Leaves for Microbiologically Safe, Value-Added Kombucha Production
Abstract
1. Introduction
2. Materials and Methods
2.1. Materials
2.2. Proximate Composition Analysis of Sea Buckthorn Leaves and Their Aqueous Extracts
2.3. Preparation of Sea Buckthorn Leaves Extracts
2.4. Determination of Proanthocyanidins by the Bate-Smith Assay
2.5. Total Phenolic Content (TPC) by Folin−Ciocalteu’s Assay
2.6. Preparation of Kombucha
2.7. pH, Titratable Acidity, and Total Soluble Solids
2.8. Shotgun Metagenomic Analysis of M. gisevi MI-2 and Fermented Product
2.9. Assessment of Antioxidant Activity
2.10. Ethanol Analysis
2.11. Assessment of Antimicrobial Activity
2.12. Sensory Evaluation
2.13. Statistical Analyses
3. Results and Discussion
3.1. Sea Buckthorn Leaves Characteristic
3.2. Effect of Extraction Conditions on Sea Buckthorn Leaves
3.3. Preparation and Characterization of Sea Buckthorn Leaves Kombucha
3.4. Antimicrobial Activity of Kombucha Evaluation
3.5. Shotgun Metagenomic Analysis of M. gisevii MI-2 and Fermented Kombucha
3.6. Sensory Evaluation
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
Abbreviations
AA | Auxiliary Activity |
AAB | acetic acid bacteria |
AACC | American Association of Cereal Chemists |
AOAC | Association of Official Analytical Collaboration |
CAZy | Carbohydrate-Active Enzymes |
CBM | Carbohydrate-binding modules |
CE | (+)-catechin equivalent |
DPPH | 2,2-Diphenyl-1-picrylhydrazyl |
EFSA | European Food Safety Authority |
FP | fermented product |
GAE | gallic acid equivalents |
GH | glycoside hydrolases |
GT | glycosyl transferases |
KEGG | Kyoto Encyclopedia of Genes and Genomes |
LAB | lactic acid bacteria |
TPC | Total phenolic content |
MG | Medusomyces gisevii MI-2 |
PCSL | Pressurized Cyclic Solid–Liquid |
PL | polysaccharide lyases |
Scoby | symbiotic culture of bacteria and yeast |
TSS | Total soluble solids |
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Parameters | |
---|---|
Moisture content, % | 5.37 ± 0.23 |
Crude protein content, % | 15.4 ± 0.57 |
Fat content, %, | 7.30 ± 0.36 |
of which: | |
saturated fatty acids, % | 2.49 ± 0.11 |
monounsaturated fatty acids, % | 0.37 ± 0.04 |
polyunsaturated fatty acids, % | 4.45 ± 0.07 |
omega-3 fatty acids, % | 3.37 ± 0.03 |
omega-6 fatty acids, % | 1.08 ± 0.09 |
omega-9 fatty acids, % | 0.30 ± 0.02 |
Carbohydrates, % | 27.85 ± 1.51 |
Total sugar content, % | 2.94 ± 0.09 |
of which: | |
fructose, % | 0.34 ± 0.11 |
glucose, % | 1.28 ± 0.09 |
sucrose, % | 1.34 ± 0.07 |
Dietary fiber content, % | 38.73 ± 1.05 |
of which: | |
soluble | 3.51 ± 0.16 |
insoluble | 35.22 ± 0.98 |
Mineral content, % | 5.35 ± 0.16 |
of which: | |
Potassium (K), mg/100 g | 66.29 ± 2.91 |
Magnesium (Mg), mg/100 g | 59.04 ± 2.78 |
Calcium (Ca), mg/100 g | 353.05 ± 5.45 |
Sample | Extraction Cycles | Cycle Duration, min | Proanthocyanidins, gCE/L | TPC, gGAE/L |
---|---|---|---|---|
L-1 | 20 | 10 | 2.47 ± 0.10 a | 8.39 ± 0.19 a |
L-3 | 20 | 5 | 3.46 ± 0.08 b | 8.81 ± 0.11 b |
L-4 | 20 | 2 | 4.44 ± 0.06 d | 9.85 ± 0.02 d |
L-2 | 60 | 10 | 5.51 ± 0.08 e | 12.42 ± 0.25 e |
L-6 | 60 | 5 | 3.92 ± 0.11 c | 9.30 ± 0.10 c |
L-5 | 60 | 2 | 4.36 ± 0.06 d | 9.98 ± 0.06 d |
Sample | Fermentation, Days | Proanthocyanidins, gCE/L | TPC, gGAE/L | DPPH, % |
---|---|---|---|---|
L4-100 | 0 | 7.30 ± 0.21 bc | 1.05 ± 0.02 cd | 84.75 ± 0.23 k |
1 | 8.28 ± 0.34 d | 0.89 ± 0.01 a | 74.39 ± 0.57 hi | |
3 | 7.74 ± 0.35 cd | 1.03 ± 0.03 bc | 80.29 ± 0.13 jk | |
5 | 7.01 ± 0.29 b | 1.09 ± 0.02 de | 77.06± 0.29 ij | |
7 | 6.11 ± 0.22 a | 1.19 ± 0.02 f | 74.75 ± 0.08 hi | |
10 | 5.76 ± 0.28 a | 1.21 ± 0.03 f | 72.83 ± 0.20 g | |
L4-75 | 0 | 7.39 ± 0.35 bc | 1.03 ± 0.02 bc | 83.92 ± 0.12 lk |
1 | 7.66 ± 0.27 c | 0.98 ± 0.01 ab | 76.25 ± 1.00 hi | |
3 | 8.99 ± 0.38 e | 0.96 ± 0.02 ab | 81.53 ± 0.36 jk | |
5 | 7.47 ± 0.31 bc | 1.03 ± 0.01 bc | 79.87 ± 0.36 j | |
7 | 7.62 ± 0.35 c | 1.08 ± 0.03 de | 77.02 ± 0.07 i | |
10 | 6.46 ± 0.17 ab | 1.11 ± 0.01 e | 75.53 ± 0.06 hi | |
L4-50 | 0 | 7.81 ± 0.36 cd | 0.98 ± 0.01 ab | 83.30 ± 0.12 l |
1 | 8.20 ± 0.39 d | 0.99 ± 0.01 ab | 77.26 ± 3.25 hi | |
3 | 8.52 ± 0.25 de | 0.94 ± 0.02 ab | 76.78 ± 0.95 h | |
5 | 7.53 ± 0.28 bc | 1.02 ± 0.04 bc | 72.09 ± 0.91 g | |
7 | 7.51 ± 0.11 bc | 1.01 ± 0.04 bc | 75.89 ± 0.01 h | |
10 | 6.38 ± 0.20 ab | 1.12 ± 0.03 e | 67.94 ± 0.08 f | |
L4-25 | 0 | 7.75 ± 0.21 cd | 1.01 ± 0.01 bc | 82.63 ± 0.35 l |
1 | 6.81 ± 0.28 b | 0.93 ± 0.01 ab | 78.03 ± 0.57 ij | |
3 | 8.35 ± 0.39 d | 0.92 ± 0.01 ab | 60.57 ± 0.57 e | |
5 | 7.53 ± 0.28 bc | 1.01 ± 0.02 bc | 48.50 ± 0.28 c | |
7 | 7.06 ± 0.31 b | 0.99 ± 0.02 ab | 32.53 ± 0.31 a | |
10 | 6.38 ± 0.17 ab | 1.05 ± 0.02 cd | 31.88 ± 0.13 a | |
G-100 | 0 | 7.19 ± 0.31 b | 0.90 ± 0.01 a | 86.97 ± 0.03 m |
1 | 7.88 ± 0.29 cd | 0.99 ± 0.02 ab | 78.15 ± 1.14 ij | |
3 | 8.36 ± 0.36 d | 1.03 ± 0.02 bc | 64.33 ± 0.18 e | |
5 | 7.06 ± 0.29 b | 0.91 ± 0.04 a | 55.77 ± 0.26 d | |
7 | 5.97 ± 0.21 a | 0.91 ± 0.04 a | 60.62 ± 0.09 e | |
10 | 6.13 ± 0.27 a | 0.93 ± 0.03 ab | 43.73 ± 2.02 b |
Sample | Fermentation, Days | Glucose, % | Fructose, % | Saccharose, % | Total Saccharide, % | TSS, °Brix |
---|---|---|---|---|---|---|
L4-100 | 0 | 0.02 ± 0.00 a | 0.02 ± 0.00 a | 8.69 ± 0.31 def | 8.73 ± 0.31 bcd | 9.56 ± 0.40 a |
1 | 0.02 ± 0.00 a | 0.06 ± 0.00 c | 8.87 ± 0.32 ef | 8.95 ± 0.32 cd | 9.56 ± 0.39 a | |
3 | 0.10 ± 0.00 g | 0.14 ± 0.00 k | 8.75 ± 0.36 ef | 8.99 ± 0.36 cd | 9.59 ± 0.41 a | |
5 | 0.21 ± 0.01 k | 0.16 ± 0.00 l | 8.45 ± 0.34 def | 8.82 ± 0.35 bcd | 9.55 ± 0.42 a | |
7 | 0.41 ± 0.01 p | 0.33 ± 0.00 q | 8.03 ± 0.10 cde | 8.77 ± 0.11 bcd | 9.48 ± 0.40 a | |
10 | 0.93 ± 0.03 s | 0.55 ± 0.00 t | 6.12 ± 0.10 a | 7.60 ± 0.13 a | 9.02 ± 0.38 a | |
L4-75 | 0 | 0.02 ± 0.00 a | 0.02 ± 0.00 a | 8.77 ± 0.31 ef | 8.81 ± 0.32 bcd | 9.56 ± 0.36 a |
1 | 0.02 ± 0.00 a | 0.07 ± 0.00 d | 8.87 ± 0.10 ef | 8.96 ± 0.26 cd | 9.56 ± 0.35 a | |
3 | 0.06 ± 0.00 c | 0.09 ± 0.00 f | 8.89 ± 0.14 ef | 9.04 ± 0.35 cd | 9.56 ± 0.41 a | |
5 | 0.18 ± 0.01 j | 0.13 ± 0.00 j | 8.51 ± 0.31 def | 8.82 ± 0.31 bcd | 9.51 ± 0.40 a | |
7 | 0.33 ± 0.01 n | 0.24 ± 0.00 o | 8.15 ± 0.21 de | 8.72 ± 0.31 bcd | 9.40 ± 0.38 a | |
10 | 0.58 ± 0.01 q | 0.37 ± 0.00 r | 7.19 ± 0.11 bc | 8.14 ± 0.36 abc | 9.20 ± 0.39 a | |
L4-50 | 0 | 0.02 ± 0.00 a | 0.02 ± 0.00 a | 9.20 ± 0.31 f | 9.24 ± 0.31 d | 9.53 ± 0.39 a |
1 | 0.02 ± 0.00 a | 0.07 ± 0.00 d | 8.84 ± 0.29 ef | 8.93 ± 0.29 cd | 9.53 ± 0.41 a | |
3 | 0.07 ± 0.00 d | 0.11 ± 0.00 h | 8.79 ± 0.32 ef | 8.97 ± 0.32 cd | 9.55 ± 0.40 a | |
5 | 0.15 ± 0.01 i | 0.12 ± 0.00 i | 8.44 ± 0.26 def | 8.71 ± 0.27 bcd | 9.46 ± 0.40 a | |
7 | 0.29 ± 0.01 m | 0.22 ± 0.00 n | 8.22 ± 0.19 de | 8.73 ± 0.20 bcd | 9.38 ± 0.38 a | |
10 | 0.91 ± 0.02 s | 0.42 ± 0.00 s | 6.08 ± 0.18 a | 7.41 ± 0.20 a | 8.79 ± 0.39 a | |
L4-25 | 0 | 0.08 ± 0.00 e | 0.02 ± 0.00 a | 8.60 ± 0.31 def | 8.70 ± 0.31 bcd | 9.53 ± 0.40 a |
1 | 0.02 ± 0.00 a | 0.08 ± 0.00 e | 8.84 ± 0.29 ef | 8.94 ± 0.29 cd | 9.53 ± 0.41 a | |
3 | 0.09 ± 0.00 f | 0.10 ± 0.00 g | 8.85 ± 0.28 ef | 9.04 ± 0.28 cd | 9.50 ± 0.40 a | |
5 | 0.21 ± 0.00 k | 0.16 ± 0.00 l | 8.45 ± 0.25 def | 8.82 ± 0.25 bcd | 9.55 ± 0.37 a | |
7 | 0.41 ± 0.01 p | 0.33 ± 0.00 p | 8.03 ± 0.35 cde | 8.77 ± 0.36 bcd | 9.48 ± 0.41 a | |
10 | 0.67 ± 0.01 r | 0.37 ± 0.00 p | 6.90 ± 0.31 ab | 7.94 ± 0.32 ab | 9.08 ± 0.39 a | |
G-100 | 0 | 0.05 ± 0.00 b | 0.02 ± 0.00 a | 8.74 ± 0.41 ef | 8.81 ± 0.41 bcd | 9.57 ± 0.42 a |
1 | 0.02 ± 0.00 a | 0.05 ± 0.00 b | 8.87 ± 0.39 ef | 8.94 ± 0.39 cd | 9.47 ± 0.38 a | |
3 | 0.02 ± 0.00 a | 0.07 ± 0.00 d | 8.81 ± 0.38 ef | 8.90 ± 0.38 bcd | 9.51 ± 0.37 a | |
5 | 0.14 ± 0.00 h | 0.10 ± 0.00 g | 8.74 ± 0.35 ef | 8.98 ± 0.35 cd | 9.54 ± 0.43 a | |
7 | 0.35 ± 0.00 o | 0.24 ± 0.00 o | 8.08 ± 0.29 de | 8.67 ± 0.29 bcd | 9.43 ± 0.41 a | |
10 | 0.25 ± 0.00 l | 0.20 ± 0.00 m | 7.83 ± 0.28 cd | 8.28 ± 0.28 abcd | 9.21 ± 0.39 a |
Sample | Fermentation, Days | Inhibition Zone Diameter, mm | |||||
---|---|---|---|---|---|---|---|
Microorganisms | |||||||
E. coli ATCC 8739 | S. aureus ATCC 25923 | E. faecalis ATCC 19433 | S. enteritidis ATCC 13076 | P. aeruginosa ATCC 27853 | S. saprophyticus ATCC 15305 | ||
L4-100 | 0 | 0.00 ± 0.00 a | 16.30 ± 0.50 c | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 14.50 ± 1.00 c |
10 | 17.50 ± 0.60 b | 17.30 ± 1.00 c | 16.30 ± 0.50 c | 18.80 ± 1.00 cd | 16.00 ± 0.80 d | 19.80 ± 1.50 d | |
L4-75 | 0 | 0.00 ± 0.00 a | 13.30 ± 0.50 b | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 10.50 ± 0.60 b |
10 | 16.80 ± 1.50 b | 13.50 ± 0.60 b | 16.30 ± 0.50 c | 20.0 ± 0.00 d | 13.90 ± 0.30 bc | 17.00 ± 0.00 d | |
L4-50 | 0 | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a |
10 | 17.80 ± 1.30 b | 12.30 ± 0.50 b | 13.00 ± 1.20 b | 20.50 ± 1.30 d | 14.40 ± 0.90 c | 17.20 ± 1.30 d | |
L4-25 | 0 | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a |
10 | 16.8 ± 0.50 b | 16.00 ± 0.00 c | 11.30 ± 1.00 b | 17.00 ± 0.00 c | 16.00 ± 0.80 d | 13.00 ± 1.40 bc | |
G-100 | 0 | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a |
10 | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 0.00 ± 0.00 a | 14.50 ± 0.60 b | 12.50 ± 0.60 b | 12.50 ± 0.60 c |
Kingdom | Species in M. gisevii MI-2 | Species in Kombucha L4-100 | ||
---|---|---|---|---|
Number | % | Number | % | |
Bacteria | 227 | 81.07 | 318 | 82.60 |
Eucaryote | 44 | 15.72 | 38 | 9.87 |
Virus | 9 | 3.21 | 28 | 7.28 |
Archaea | 0 | 0 | 1 | 0.26 |
Total | 280 * | 385 * |
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Jagelavičiūtė, J.; Girtas, J.; Mažeikienė, I.; Šarkinas, A.; Almonaitytė, K. Pressurized Cyclic Solid–Liquid (PCSL) Extraction of Sea Buckthorn Leaves for Microbiologically Safe, Value-Added Kombucha Production. Appl. Sci. 2025, 15, 6608. https://doi.org/10.3390/app15126608
Jagelavičiūtė J, Girtas J, Mažeikienė I, Šarkinas A, Almonaitytė K. Pressurized Cyclic Solid–Liquid (PCSL) Extraction of Sea Buckthorn Leaves for Microbiologically Safe, Value-Added Kombucha Production. Applied Sciences. 2025; 15(12):6608. https://doi.org/10.3390/app15126608
Chicago/Turabian StyleJagelavičiūtė, Jolita, Juozas Girtas, Ingrida Mažeikienė, Antanas Šarkinas, and Karolina Almonaitytė. 2025. "Pressurized Cyclic Solid–Liquid (PCSL) Extraction of Sea Buckthorn Leaves for Microbiologically Safe, Value-Added Kombucha Production" Applied Sciences 15, no. 12: 6608. https://doi.org/10.3390/app15126608
APA StyleJagelavičiūtė, J., Girtas, J., Mažeikienė, I., Šarkinas, A., & Almonaitytė, K. (2025). Pressurized Cyclic Solid–Liquid (PCSL) Extraction of Sea Buckthorn Leaves for Microbiologically Safe, Value-Added Kombucha Production. Applied Sciences, 15(12), 6608. https://doi.org/10.3390/app15126608