DOI: 10.57755/ntsk-bh25
title: The Effects of Chemically Modified Biochar on Biomethane Production from Glucose and Sugar Beet Pulp

description: <p>The research aimed to study the effects of straw-derived biochar and two types of chemically modified biochar on biomethane production from glucose as a model substrate and sugar beet pulp as a real substrate.The biochar chemical modification with H3PO4 acid and KOH base resulted in a change in biochar surface area properties and its functional group&#39;s abundance and a decrease in biochar mass yield production. The anaerobic digestion process was performed in batch reactors kept at 37 °C for 20 days. The substrate-to-inoculum ratio by volatile solids was 0.5, while the mass of added biochar corresponded to 16 g×L⁻¹. The results showed that neither the addition of biochar nor the chemically modified biochar had any positive effects on biomethane production or its kinetics in the case of both substrates. The highest methane production was found in reactors without biochar added, respectively 385 and 324 mL gVS⁻¹ for glucose and sugar beet pulp. It is hypothesized that an anaerobic digestion process was performed at optimal conditions therefore, biochar could not help to enhance methane production, while biochar could adsorb some volatile fatty acids, making them less available to anaerobic microorganisms.<br /></p>
description: <p class="MDPI22heading2" style="margin:4px 0px;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-style:italic;font-size:13px"><span lang="EN-US" style="font-size:12px">Materials</span></p>

<p class="MsoNormal" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:13px"><span lang="EN-US" style="font-size:12px">In the research, six materials were used,
        including inoculum to start AD, two substrates for AD, one biochar, and two
        chemically modified biochars as additives to the AD process.</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">The
        inoculum (I) was made from the digestate collected from a 1 MW<sub>el</sub>
        commercial agricultural biogas plant (Bio-Wat Sp. z o.o., Świdnica, Poland).
        The digestate was mechanically sifted through a fabric filtration bag to clean
        it of various solids: untreated substrate, plastics, stones, etc. The purified
        liquid digest (inoculum) was stored in a plastic container at room temperature
        for 24 hours before being used as an inoculum to start the AD process.</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">The
        substrates were glucose and sugar beet pulp. The glucose (G), chemical formula
        C<sub>6</sub>H<sub>12</sub>O<sub>6 </sub>(pure chemical). The glucose was kept
        in a sealed plastic container at room temperature. The sugar beet pulp (BP) in
        pellet form was purchased at a local market. The pellets were ground using a
        knife mill (LMN100, Testchem, Pszów, Poland). The material was ground until it
        reached a homogeneous consistency. The ground BP was stored in a plastic bag at
        room temperature.</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">The
        biochar (BC) was made from wheat straw collected at a farm in Wrocław (Lower
        Silesia, Poland). The straw was ground to a size &lt; 1 mm using the laboratory
        knife mill. The shredded wheat straw was stored in a plastic bag at room
        temperature until conversion to biochar. The conversion of wheat straw to
        biochar was as follows. The wheat straw was placed into four metal containers,
        (sizes of container 30 cm x 9 cm x 7 cm), then closed with lids. Sealed
        containers with materials were placed into a muffle furnace (Snol 8.1/1100,
        Utena, Lithuania). After the two containers were placed in the furnace chamber,
        the chamber was closed and filled with CO<sub>2 </sub>gas to provide an inert
        atmosphere. The inert gas flow rate was 10 mL<sup>.</sup>min<sup>-1</sup>. The
        furnace programmer was then set and turned on. The process temperature was 600
        °C, and the pyrolysis duration was 4 h. After that, the heating was turned off
        and the chamber was left to self-cooling to room temperature.</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">Biochar
        was chemically modified using two methods: with a 30% H<sub>3</sub>PO<sub>4</sub>
        and with a 30% KOH solution. The chemically modified biochar was labeled BC_A
        (biochar modified with acid, H<sub>3</sub>PO<sub>4</sub>) and BC_B (biochar
        modified with base, KOH). The chemical modification procedure was as follows:
        200 g of BC was placed into a 2000 ml beaker, then poured with 300 g of both
        acid and base 30% solution, and stirred with a glass dipstick until the
        solution was evenly distributed in the beaker. For 1 g of BC, 1.5 g of solution
        was used. After the mixing process, the beaker was set aside under the fume
        hood for 1 h. After a time of 1 h, the wet chemically impregnated biochar was
        lined on trays and placed in a drying oven at 105 °C for 24 h, until dry. After
        24 h, the dry chemically impregnated BC were removed from the dryer and then
        weighed. After this step, dry chemically impregnated BC were placed in a muffle
        furnace and heated at 500 °C for 1 h, under an inert atmosphere (CO<sub>2</sub>).
        The inert gas flow rate was 10 mL<sup>.</sup>min<sup>-1</sup>. Chemically
        modified biochars were washed with water to remove the chemicals used for
        modification. Chemically modified biochars were placed on a 63 µm mesh sieve
        and then rinsed with warm tap water (~50 °C) for 25 min, with a break every 5
        min to check pH. The pH was measured using a pH meter (Elmetron, CPC-411,
        Zabrze, Poland). The rinsing process was continued until the pH compactness
        stopped changing significantly. The flow rate of tap water was approximately
        0.216 m<sup>3</sup>×h<sup>-1</sup>, and to wash 1 g of biochar used around 0.82
        L of water.</span></p>

<p class="MDPI22heading2" style="margin:4px 0px;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-style:italic;font-size:13px"><span lang="EN-US" style="font-size:12px">Methods</span></p>

<p class="MDPI23heading3" style="margin:4px 0px;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:13px"><span lang="EN-US" style="font-size:12px">Substrate
        analysis</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">The
        materials were analyzed to determine total solids (TS), moisture content (MC),
        volatile solids (VS), and ash content (AC,). Moisture content and total solids
        were determined using a laboratory dryer (WAMED, model KBC-65W, Warsaw,
        Poland), according to PN-EN 14346:2011. Materials were placed into crucibles,
        in triplicate for each type of material, and weighed. Next, samples were placed
        in the laboratory dryer for 24 hours at 105 °C. Once the drying process was
        complete, the samples were weighed. The TS and MC were calculated following
        Equations 1 and 2: </span></p>

<p class="MDPI39equation" style="margin:8px 0px 8px 47px;text-align:center;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:12px"><img src="data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAn8AAABsCAIAAACU8UOwAAARX0lEQVR4Ae2d3bmjLBeGacEaaMEeKMEaaMEOPJ4TO7ACK7ABG7ADe&#43;C7vqx31rBVjNsYY5LbgxmCwII7ZD8u/jSBCwIQgAAEIACBcwmYc81hDQIQgAAEIACBgPrSCSAAAQhAAAJnE0B9zyaOPQhAAAIQgADqSx&#43;AAAQgAAEInE0A9T2bOPYgAAEIQAACP9T3z&#43;0KIUjgz58/hGFCH6AP0AfoA/SBR/rA4qPGVH0XExEJAQhAAAIQgMAOAuLHzjP&#43;UN/5bWIgAAEIQAACH0kgpYvHNjZl5Yf6phIdWxVKgwAEIAABCLycwGslD/V9eQegAhCAAAQg8AIC56hvygrq&#43;4KvHJMQgAAEIPAlBDap75ewoJkQgAAEIACBlC4eSyZlBd/3WM6UBgEIQAAC70EgpYvn1B71PYczViAAAQhA4FoEzlHflBXU91q9gdpAAAIQgMAnEdikvp/UYNoCAQhAAAIQWCGQ0sWVLDtupazg&#43;&#43;6ASRYIQAACEHh7AildPKdhqO85nLECAQhAAALXInCO&#43;qasoL7X6g3U5gsJNE2TZVme5yGEsiyNMdbacRwlPsuytm2/EAtNhsBnENikvp/RVFoBgTciMAyD977rOmNMVVVd143jaIwpy7Ku6xCCtbaqqjdqEVWFwLsQSOnisfVPWcH3PZYzpUFgDwFxc7uuCyGI&#43;qriWmubptlTKHkgAIFVAildXM102E3U9zCUFASB3QS890VRSPa2bbMsk/AwDMaYcRx3l0xGCEAgReAc9U1ZQX1T3wvxEDiPgLVWxpll6tc5J7brupb54H1VKYpC/Ol92ckFAQg8TmCT&#43;j5uhhIgAIHfEhAHt&#43;97yZjnuQ47F0Xhve/7vq5r770x/z0uO&#43;dEVo0x3nvJWFVVlmXGGBHyPM/N7bLW/rZKpIfANxBI6eKxbU9Zwfc9ljOlQeDXBGTSV7LJpK86rFVVyQosnQ8ebpdKbFmWwzCEEKqq8t4Pw1CWpQxi933/iN/862aQAQLvRiCli&#43;e0A/U9hzNWIHAAAWtt13Xe&#43;7Isq6oahkG9ZPF6syzz3ss8cVVVOpp9gG2KgMDHEThHfVNWUN&#43;P61A06HMJOOdkJ1LXdc45FdoQgizOGsdRPOAQgnOuvV06pv25YGgZBK5LYJP6Xrf61AwCpxCQkV6ZLo3/1QXJIm/iaBpjnHNnLkiuqkpHlbMskzFnAaPHdOiotcwTl2V5FLm2bXU5mJYpQIRVURQxjb7vnXPGmCzL1qvR9721VsfYtfCu6&#43;TgEY0hAIEDCaR08UATIYSUFXzfYzlT2nsTKG7XOI5t2xpjRN6cczrA65zL81zi&#43;77XrUFv1&#43;yyLOfbiJumWZTJcRyLopCnjUlLBcg4jsMwWGtVnsdxlGHwEIKeJTLJqx&#43;ttW3bjuM42dwsI&#43;2ajAAEjiWQ0sVjraRKQ31TZIj/RgJ5nov3VpalLhWWI6gEhzFmLlrvSEoeHeK2yOKvxWHqoij6vq&#43;qSsVVmiyrtfUgzLqu9ZGlaZp4p3JRFMpzgksKkcjqdmlYhxwmWfgIgUMIPKK&#43;cV&#43;V3q5bEuSJUzcjpKygvod8iRTyaQTyPNcfT9w2a636vnF8HE4NX8vwbJzyteFYgFekVys5V9&#43;JxIqPK0Pf3vt4xbUw0aLigMxhS4z&#43;RZO6xePYcRbCEHgtgaZp9O&#43;DtdZ7L7&#43;FuFZN08iY2Sb1jXMShsDXEpBtP&#43;rSxRx0hjL2GuME7xUWkfPeZ1m26PXGzZmr70RT4xFmd7s0u6TUOWmNDyEs&#43;r7OueZ2xTuY41yEIfA4gZQu3i05dnMl8Vx95ZD2YRhSVvB973ImwdcRkEnflOM1jqMscTpKgOPlXc8Lp75F0UWd2E4lky3Fk5HnRfWVbU6L6psSeHmPk8z7dl1X17UsZ7PW9rdrssRspZLcgsB2AildXC9B9tZP0iyqb9M0k59MnAv1jWkQhsD/CUxGTRehTNQlTiOalNLROOXLw33fy2lZ&#43;3xfmejVx5R45Lkoio0jzyEEOdpa1jzLcq1hGOIRaXGFX46LCnwYgX3qu7gYcFF9ZVwnZQX1/bDuRHMOIGCtXVz6GxftvV95qo1TXjb8&#43;LyviLcO0csfIBHjiTBveaARUEVRiCM&#43;Ud8t3vllUVOxjyEgghpv9pOmLaqvDD5vUt&#43;PAURDILCbQDx5GRdS3i6JEV9NVSdO9i7hWHqlzncXXsXrwLWZ&#43;e2SHUd5nutTyzAM2e3gLVn/KWPLkmtl2KBtW/WYJ&#43;r71rQVF4FLEUjp4kol5e/DPMGj6rujKvNKEAOBtyagI8aTJUKiPXI3z/N3F4Nf7fctyzJ&#43;Z0Ps9E&#43;O1NBRaBlPljM04jc4ySFci1uJdN5X&#43;o84GX3fi5DPvY237mZU/goEdkheSmVT8amNdv8/ny5GsKMqcXbCEIAABO4SSD27xKMLUoi441mWHbXA7W7dSPBVBHZIXsr3naxAVIzW2pQV1FcpEYAABJ5O4LaNqHm6GQxA4DkEUvO&#43;KfU1xmxS3&#43;fUllIhAAEIQAAClyOQ0sX1ik7OQ5UxZ52xioeaWfO8TpK7EIAABCDwjQT2qe/ift9FfOz3XcRCJASuQkBmN2Wtr76qaBxHnfV89xVeVwFNPSDwk8A&#43;9ZUXev4safmT7AxOWWHed5kasRA4h8AwDN573ebUdZ2ccynv8ZX9gmx1Pee7wAoENhKIz3lOZeGc5xQZ4iFwIQLi5soeJ1FfVdzJJNOFKk1VIPDmBFJe6ZZmVbcrlbLrOn0NQ8oKvm&#43;KHvEQOI&#43;A9163wMpRHmJbVm3Em2glXhZYMiJ93jeEpU8kkNLFc9qK&#43;p7DGSsQWCMQn0dRlqUeZ1HXtZ79NMmfZdlclSdp&#43;AgBCKwQOEd9U1ZQ35WvhlsQOIOAHuokxvI812Hnoii8933fy4uDxnEsisIYI68ZDiGM4&#43;ick3ckWGvl1UB5nqs2y6sLzmgGNiAAgSUCm9R3KSNxEIDAcwnIpK/YkElfPeRSRpj18GTnnEwmVVUlkV3XiXiXZSlKLOu29DUs3nvOaHzu90fpb0sgpYvHNihlBd/3WM6UBoFnEej7XjfyyzJpsRTHq&#43;2yLKuqGsdR3Wi9RQACEBACKV08hw/qew5nrEDgUQJd18kccF3XMrAsLnJd1&#43;ocqw3ZESEOsUYSgAAEYgLnqG/KCuobfxeEIXBdAvIKIHnlgHMuz3NZdVUUxXzxs7wXqO/767aHmkHgOwhsUt/vQEErIfDJBIqiGMfRe897gT75a6ZtRxBI6eIRZf8rI2UF3/cfI0IQ&#43;AACzjldcvUBzaEJEHgegZQuPs9iXDLqG9MgDAEIQAAC30LgHPVNWUF9v6Wf0U4IQAACEDifwCb1Pb9aWIQABCAAAQi8hEBKF4&#43;tTMoKvu&#43;xnCkNAhCAAATeg0BKF8&#43;pPep7DmesQAACEIDAtQico74pK6jvtXoDtYEABCAAgU8isEl9P6nBtAUCEIAABCCwQiCliytZdtxKWcH33QGTLBCAAAQg8PYEUrp4TsNQ33M4YwUCEIAABK5F4Bz1TVlZUN8/f68Qwt/gH8KCAg5woA/QB&#43;gD9IFf9YHFh44f6ruYgkgIQAACEIDA5xFIeaXHtjRl5Yf6phIdWxVKgwAEIAABCLycwGslD/V9eQegAhCAAAQg8AIC56hvygrq&#43;4KvHJMQgAAEIPAlBDap75ewoJkQgAAEIACBlC4eSyZlBd/3WM6UBgEIQAAC70EgpYvn1B71PYczViAAAQhA4FoEzlHflBXU91q9gdpAAAIQgMAnEdikvp/UYNoCAQhAAAIQWCGQ0sWVLDtupazg&#43;&#43;6ASRYIQAACEHh7AildPKdhqO85nLECAQhAAALXInCO&#43;qasoL7X6g3UBgIQgAAEPonAJvX9pAbTFghAAAIQgMAKgZQurmTZcStlBd93B0yyQAACEIDA2xNI6eI5DUN9z&#43;GMFQhAAAIQuBaBc9Q3ZQX1vVZvoDYQgAAEIHB9AtXtCiE0TWP&#43;Xt57qXnXdRrepL7XbzA1hAAEIAABCBxCIKWLdwtvmkbF1Tmn6Y0xGt80TVVVIYSUFXxf5UYAAssE5Lk2/o2FEIZhyG5X27ZxtqZp8jyXLNbapmniuw&#43;Gq6oqy3JSSNu21lpjzNzcyq1JIVs&#43;dl339xH/x/9Zlmn2qqqkMsaYPM&#43;7rtNbcaDve0k2aU7XddbacRzjxIQh8CQCKV28a86Y/6Sz67r4Ny5&#43;sGa31g7DoB8nAdR3AoSPEJgSaNtW1Ca&#43;4ZzLsmwiHt77LMvk19j3vYbjjPvCfd&#43;LqMvTtBYiiigWq6oyxqjgrdzS7L8K1HUtf03GcTTGqFF9LqmqKssyqcA4jtbavu8XTVhr27aVNPEfL2ut1n8xI5EQOJDAPvWtqkod3EllJurbNI1zLmUF9Z3Q4yMEpgSqqiqKQp92ZaZHYmLHV354sXg451LyM7Wx&#43;nkYhqIoxnF0zk3U13uf57nmzrJM/y6s3NL0vwp476V18jgiD/Vd12mVnHNqfaXkYRgUpk6ehRCE80pGbkHgCgRWnhGrqtKHURkhM8ZsUt8rNIw6QOBqBJxzsbKKxyYKFI&#43;RWmu3aE8I4ce47c8PqmSLEObqa62N/W93uyTvyq3FwrdHlmVprZ2nF9c/fv6YpwkhdF2nf6FUfWWoIOa5mJdICBxIIKWLKybk2XFxPHnxl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/></p><table style="width:522px;margin-left:35px"></table>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">where:</span></p>

<p class="MDPI31text" style="text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;margin:0px 0px 16px;text-indent:0px;font-size:12px"><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">TS</span></em><span lang="EN-US"> – total solids, %, </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">m</span></em><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif"><sub>wet</sub></span></em><span lang="EN-US"> – a mass of the material before the drying
process, g, </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">m</span></em><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif"><sub>dry</sub></span></em><span lang="EN-US"> – a mass of material after drying process at
105 °C, g, MC – moisture content, %.</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">Samples
after drying were placed in a muffle furnace (Snol 8.1/1100, Utena, Lithuania)
for the burning process. The process temperature was 550 °C, and the process
time was 3 h. After this step was completed, the crucibles were removed from
the furnace, and the samples were allowed to cool to ambient temperature in a
desiccator. The samples were weighed to determine the ash content (Equation 3)
and volatile solids (Equation 4).</span></p>

<p class="MDPI39equation" style="margin:8px 0px 8px 47px;text-align:center;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:12px"><img 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/></p><table style="width:522px;margin-left:50px"></table>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">where:</span></p>

<p class="MDPI31text" style="text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;margin:0px 0px 16px;text-indent:0px;font-size:12px"><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">AC</span></em><span style="line-height:107%;font-family:&#39;calibri&#39; , sans-serif;font-size:15px">  </span><span lang="EN-US">– ash content, %, </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">m</span></em><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif"><sub>ash</sub></span></em><span lang="EN-US"> – a mass of material after the combustion
process at 550 °C, g, </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">m</span></em><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif"><sub>dry</sub></span></em><span lang="EN-US"> – a mass of material after the drying process
at 105 °C g, </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">VS</span></em><span lang="EN-US"> – organic matter content in dry matter, %.</span></p>

<p class="MDPI23heading3" style="margin:4px 0px;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:13px"><span lang="EN-US" style="font-size:12px">Determination
of carbonaceous additives surface properties</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">Functional
groups present on the surface of wheat straw, BC, BC_A, and BC_B were
determined using a Nicolet iN10 integrated infrared microscope with a Nicolet
iZ10 external FTIR module (Thermo Fischer Scientific, Waltham, MA, USA). For
each spectrum, 32 scans were averaged over an average infrared range of
400-4000 cm<sup>-1</sup>. The results of specific materials were merged to show
the difference in functional groups due to pyrolysis and chemical modification.
Next, functional group peaks were quantified as an integrated peak area. To
quantify functional groups from FTIR data, the baseline for each functional
group was determined as the minimum absorbance value within its wavenumber
range to ensure that peak intensities are measured relative to the lowest
background absorbance in the region. The following wavenumber ranges were
considered: OH 3200–3600 cm⁻¹; C&#61;O 1650–1750 cm⁻¹; C&#61;C 1550–1650 cm⁻¹; C-O
1000–1300 cm⁻¹, and C-H Bending 850–950 cm⁻¹. The integrated peak area was
calculated using Equation 5.</span></p>

<p class="MDPI39equation" style="margin:8px 0px 8px 47px;text-align:center;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:12px"><img src="data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAnYAAABACAIAAAAYiq03AAAK9UlEQVR4Ae2dwZWjOhBFlQIxkAI5EAIxkAIZsJ4NGRABEZAACZABOeif6Xem0MduGmOjtuGymKGxUFVdZD9KEsJ57//8&#43;aN//3xt7MOBNkAboA3QBmgDT7YB773zbBCAAAQgAIHLEFBWeXS4svJXYuPYOzoe6ocABCAAAQj8SCCm5CGxP14OCkAAAhCAwHkIxJFYstjztBgigQAEIACBtyIwS&#43;xbuYUzEIAABCAAgeMIkMUex5aaIQABCEDg0gTiSKwQMxZ76aZG8BCAAASuRiCOxM4dxXHsXe0qEi8EIAABCFyWwCyxl0VA4BCAAAQgcDUCcbLKWWLj2LvaVSReCEAAAhB4QwIxJY&#43;x2DdsALgEAQhAAAJHEYgjsWSxR10/6oUABCAAgYsTmCX24iAIHwIvIdC2bZIkTdMMw5CmaVmWL6mWSiAAgdcSIIt9LU9qg8DrCUzTVJalcy5JkmmavPdd11VV1TRN3/fa7/s&#43;yzKV6bru9U5QIwQg8DiBOBIrvxiLffz6cAYEvC/LsmmaaZqc&#43;/sl0pbned/3/uvTcRzrulYuW1VVXdf/SvE/BCDwmwTiSOzcURzH3m8SxTYEXkpgHMckSW6rVEZbluUiZy3LchiG2/IcgQAEzkpgltizRkhcEDiIQF3XeZ7fVp4kSZqmSmT16TRNVVWhr7esOAKB3yIQJ6ucJTaOvd&#43;iiV0IvJxAnucbO35NX5n99PKrQIUQ2EcgpuQxFrvvGnHWpQkkSbJFYtu2df&#43;2u1nvpSESPAR&#43;iUAciSWL/aXLi9kPJ6ApTm3bfngcuA8BCBxIYJbYA41QNQROR6Dve&#43;dcOOB6uhAJCAJnJkAWe&#43;arS2yfTqDrOiT20y8i/l&#43;ZQByJFWHGYq/c0oh9D4G6rpHYPeA4BwLvQSCOxM4dxXHsvQdbvIDAswQksVrR6dm6OB8CEDgpgVliTxogYUHgEAJVVYUrOr3QhkZ5nXPHPUerCc5N07zQ7UerGsexLMv1&#43;WJaGGscx5XKDdd6sZUa&#43;OiaBOJklbPExrF3zWtJ1L9CYJqmjc/V7HAvz/ODJFYLHd9dN2qHn3dP0XTo4yT8rtHwoN6RsMWBvu/TNF0vWdd1mqZh/exD4EcCMSWPsdgfLwcFPo9AVVVJkhy02sOhElvXdVEUxxHv&#43;/5QCV/3XLc&#43;2ydjS2VX&#43;uRtFeh1u3wKgZDAMxJbf23eew0Y/Xvu/e//3vu&#43;7&#43;1nhyw2ZM7&#43;eQj0fa/Vlw5a7eFQiS2KYsuiFruvVtM0B2HZ4tKOG4gsy1aAFEXxu53eW6KmzGkItG1rClrX9d2bxbZt1WJniT1N/AQCAe99lmV6y813WmJjeOEdqO3f/dqEYLMsezIRHIahKAq95G6heUmS9H2v4d40TS2BG4ZB78VzzoXLIHddZ&#43;/Lq6pKfur0qqqmaZIhUylJuF5tK1sWmjpdnXN5nldVpTcZqGSWZd578yoc&#43;2yaJk1T0bPhVb3pL0kS1WYm0jS1MgtXvffqQ06S5LZ&#43;q0HZgyyWZRn2JFvU31UVVsL&#43;lQnszmLDEaK6rsOGGvJM03Qcx1lid9sLK2UfAu9AIOzG&#43;U5in/RzIRuP1qa39JRlOX1tSZKYNA7D4JzTS90Xw8l5nuv2eZomfYE1cKvyEhXnXNd17ddW13WWZbKS57m9&#43;Ufm9Gf&#43;tcl/da3r9sKmTGteku5IdM&#43;uoVy7CwnPMivyXAKviFR&#43;HMfFw05d1w3DYKHJW&#43;dc&#43;Msl63arEVrUvvzfUtWjV4ryZyWwT/IWoxIrEtu2rf34MBZ71lZ0xbjGcbSWXdf1k7nmdwSflNiiKMxJ732SJKZ/TdMkSWITfMKXDeisUHt0rsmz9945Z9mqMjxTJsUiwbM80t4XFAqh975pGqWtOkuJbCiTcuNWMlW&#43;LEsNJw/DIDd0XGKp/fDfsCt4mqYQjga3TJgXft52O69XFRpl/8oE9kls2Huk3hSNGakLx&#43;47vff6apDFXrmNnTP2sMWr3d&#43;N88mO4icl1jlnIifNMCEsimIhmTbQeNvlqyhMjxeCl6apnWsQFtqZ57m0cDEvd&#43;FGWZYme23b2gzexVlmxbrcsyxTLquPvpNYU1D1RYc/VQuJXVi8Hbder8o8ZAcCjxLQ9yu8xw0fPCvLctH7kqbpLLGPGqM8BN6QgEYlzTEtc2h/vnDnGYmVzJiKrGS0yh1NfeW/JFlZb9M0i5Ehy9r1c2Dqa7GH2hlKcthjvMgUvfehWpdlaTcB4VlmYhGgHTexvBuRfrlsnsjtWSIWWrz1U0dWqgqrZf/KBHZksd/dIBrGsA9J35pZYnfYs3rZgcA7END4n/W4qrdzcV/5Kj&#43;fkVgJW9M0elV7VVVlWQ7DoPFXG1jVozWWhtqkHp2uMEONkdxacix5vo03SRKlreM4ZllmTwdZr3Lf96pqHEcloAu11tPGbdtqTpYmZE3T1DSNrGuwVp4vlpgIRd180y/XNE2Lga7bAt77FT9DCf&#43;uKquTnYsT2CF5ejHlCrc0TW2YRhKrwozFrkDjo88gIMFwzlmCpR9u55ypyKsikYSE36VHa9bE1zzPh2FQqi23m69Nfd1pmoa3C9YBvlhPQ/N9nHNZloXlbTQ09E0jo7K&#43;qMem8iquLMtM1BdqHc5P1ripZjiHQHTK7XH97th9gHyT7i78D90OO4dX/LQBsJWqwmrZvzKBHRK7JYsNu5H/11G8w96VLw&#43;xX5mAvmmholyZxqOx2wSr7SeGM5i2n0VJCLyWgO4FTUTrug5vFvM8D//U3EMJ698slg0CENhIAIndCOpusUdXd&#43;q6Lnw4&#43;G6dHITAowT2ZZWLp7qtbymcZCdPpMezxO6z92hUlIfACQhIYsNe2RMEFTOE7WsUby8Z039snYDAPsnbPsbPc7EnaCSE8DsEJLE2Jfh3nPhwq4tpUHejaZqmLEvrl7tbhoMQ2Edgn8Sq&#43;3eLRT1BSxa7hRVlIPA/Alr56PZ5mP8V4g8IQOCMBMI1ir&#43;Lz549myX2u6IchwAEFgQksYuD/AkBCHwQgd1ZrBZ1WpntyJt2PqgZ4Oo7EgifIXlH//AJAhD4icAzEvtT3cvPeS52SYS/IbBCwBYdXCnDRxCAwDsTiCOxc0dxHHvvTBzfILCRQJZltujSxlMoBgEIXJDALLEXDJ6QIbCPgHOOuU770HEWBN6EQJyscpbYOPbeBC5uQGA3gfBNA3pqsygKrSNo75/ZXTknQgACcQjElDzGYuNcU6x8MAG9BV1Sak/E9n3ftm1RFOM4DsOgF6zqic&#43;VCYcfTAHXIXAWAnEkliz2LO2FOA4mMAxD8rUtliGtvza91afrunEctWACEnvwBaF6CHwAgVliP8BZXITA&#43;xEoikJvf7M3/Pz42Nz7BYFHELgcAbLYy11yAv5EAkVRJEmymGBsqe0nRoTPELgCgTgSK5KMxV6hRRFjPALV1xbPHpYgAIEHCcSR2LmjOI69ByFQHAKfR8AF2&#43;d5j8cQgMDrCMwS&#43;7o6qQkCEIAABCDw1gTiZJWzxMax99bIcQ4CEIAABK5BIKbkMRZ7jTZFlBCAAAQg8EUgjsSSxdLcIAABCEAAAocQmCX2kOqpFAIQgAAEIPB&#43;BMhi3&#43;&#43;a4BEEIAABCJyCQByJFSrGYk/RZAgCAhCAAAS2EYgjsXNHcRx722KnFAQgAAEIQODjCcwS&#43;/GhEAAEIAABCEBgG4E4WeUssX/&#43;bd77f7t/2BcKOMCBNkAboA3QBna0Ae/937FYNghAAAIQgAAEXk7gPwH8VQnrpqoYAAAAAElFTkSuQmCC" /></p><br /><p><span lang="EN-US">where: </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">A</span></em><span lang="EN-US">– integrated peak area, </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">v</span></em><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif"><sub>1</sub></span></em><span lang="EN-US"> – lower wavenumber limits for a functional
group, </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">v</span></em><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif"><sub>2</sub></span></em><span lang="EN-US"> – upper wavenumber limits for a functional
group, </span><em><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">absorbance(v)</span></em><span lang="EN-US"> – absorbance value at wavenumber.</span></p><table style="width:522px;margin-left:34px"></table>



<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">The
functional group regions identification and visualization, including FTIR peak
baseline determination and integration, were supported by AI-based computational
tools (ChatGPT, OpenAI, 2024). The methodology followed AI-assisted numerical
integration using Simpson’s Rule and graphical representation using Matplotlib
in Python.</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">Next,
the specific surface area and pore size were determined using an adsorption
analyzer (ASAP 2020, Micromeritics, Atlanta, USA). The analysis was performed
using the volumetric method. Before measurement, the materials were degassed at
300 °C. The degassing process lasted for 6 hours. Then nitrogen (N<sub>2</sub>)
sorption was measured at 77K. Based on the obtained isotherms, the following
parameters were determined: <em>SSA<sub>BET</sub></em> – specific surface area
calculated by BET method, <em>V<sub>T</sub></em> – total pore volume &lt; 50
nm,  <em>L</em> – average pore dimension
&lt;50 nm, <em>SSA<sub>DFT</sub></em> – specific surface area calculated by Q<sub>SDFT</sub>
method, <em>V<sub>DFT</sub></em> – pore volume by the Q<sub>SDFT</sub> method, <em>L<sub>DFT</sub></em>
– dominant pore dimension by Q<sub>SDFT</sub> method, <em>V<sub>0</sub></em> – a
volume of micropores by Dubinin-Radushkevich method, <em>L<sub>0</sub></em> –
average dimension of micropores by the Stoecklie method [14].</span></p>

<p class="MDPI23heading3" style="margin:4px 0px;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:13px"><span lang="EN-US" style="font-size:12px">The process of
anaerobic digestion</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">The
experiment was conducted using two automated systems for measuring biomethane
production potential (AMPTS® II, BPC Instruments, Lund, Sweden). Three
replications were carried out for each variant, and all variants and
replications were started at the same time. Materials in the reactors were
designated as follows: I – inoculum, G – glucose, BP – beet pulp, BC – biochar,
BC_A – biochar modified with H<sub>3</sub>PO<sub>4</sub> solution, BC_B –
biochar modified with KOH solution. Each reactor was filled with inoculum to a
working volume of 400 mL, the total volume of one reactor was 500 mL with 100
mL of the headspace. The masses of the substrates going into each reactor were
measured with a laboratory spatula and then placed on plastic trays. The
substrate masses were selected so that the ratio of substrate volatile solids
to inoculum volatile solids (SIR) was 0.5 [15]. The BC and chemically modified biochars were
measured similarly, and 6.4 g of additive material was added to each reactor.
The mass of added additives corresponded to 16 g<sub>BC</sub>×L<sup>-1</sup>.
The weighted substrates and additives were then placed in reactors filled with
inoculum. The next step of the experiment before starting the AD process was to
mix the contents of the reactors using a glass dipstick. The reactors were then
sealed tightly and placed in a water bath. The process was carried out under
mesophilic conditions (37 °C) for 20 days. During the process, the mixing of
the materials in the digester took place every hour and lasted for 2 minutes.
The distribution of all variants is described in Table 1.</span></p>

<p class="MDPI41tablecaption" style="margin:16px 0px 8px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:12px"><strong><span lang="EN-US">Table
1.</span></strong><span lang="EN-US"> Configuration of the anaerobic digestion
experiment, the mass of material placed in the reactors</span></p>

<p class="MDPI42tablebody" style="margin:0px;text-align:center;font-family:&#39;palatino linotype&#39; , serif;color:black;line-height:normal;font-size:13px"><img 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style="margin-left:auto;margin-right:auto" /></p><table style="width:100%"></table>

<p class="MDPI23heading3" style="margin:4px 0px;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;font-size:13px"><span lang="EN-US" style="font-size:12px">Analysis of
volatile fatty acids content and pH</span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:13px"><span lang="EN-US" style="font-size:12px">During
the process, samples were taken for analysis of VFA and pH. Samples were
collected on days 1, 3, 7, 14, 20. Samples from the reactors of about 0.5 ml
were collected with a one-way pipette into centrifuge tubes of 10 ml and then
stored in a fridge at 4 °C. During sampling, the pipet tips were changed
between variants to avoid mixing of materials. </span></p>

<p class="MDPI31text" style="margin:0px;text-align:justify;line-height:19px;font-family:&#39;palatino linotype&#39; , serif;color:black;text-indent:0px;font-size:12px"><span lang="EN-US">Before
VFA analysis, the samples were centrifuged at 10 </span><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">℃</span><span lang="EN-US"> at 4500 rpm for 10 minutes, then diluted in Milli-Q water, and
re-centrifuged at 10 </span><span lang="EN-US" style="font-family:&#39;cambria math&#39; , serif">℃</span><span lang="EN-US"> for 5 minutes at
15000 rpm. Quantitative and qualitative analyses were determined using
Ultra-Performance Liquid Chromatography (U-HPLC) with the use of the UltiMate
3000 System (ThermoFisher Scientific, UK). VFA standard solution (Volatile Free
Acid Mix, CRM46975, Merck, Poland) in a final concentration of 10 mM was stored
in the fridge at 4 </span><span lang="EN-US">°</span><span lang="EN-US">C, according to the manufacturer&#39;s guideline. The standard curve was
prepared based on the standard solutions by appropriate dilution of the
analytes in Milli-Q water. The concentration of VFA (and other components such
as ethanol) was studied using a HyperREZ XPCarbohydrateH&#43;8 μm column (Thermo
Scientific, Waltham, MA). Identification of analytes was performed with the
UV/VIS-DAD detector at 208 ± 1 nm. To determine the possible presence of
ethanol in the samples, a refractive index (RI) detector (Shodex, Ogimachi,
Japan) was used simultaneously. As the eluent 0.25 mM trifluoroacetic acid was
used, the flow rate was 1.1 mL×min<sup>-1</sup> (isocratic elution) and the
column temperature was 35 °C. Data analysis was performed using Chromeleon 7.1
Software. All samples were analyzed in triplicates. The pH was measured using a
pH meter (Elmetron, CPC-411, Zabrze, Poland)</span></p>