Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 1-3 and 6 are rejected under 35 U.S.C. 103 as being unpatentable over Xu (CN106517340A: An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below) in view of Zhao "Synergy of Fe and biogenic Mn oxide components mediated by a newly isolated indigenous bacterium to enhance As(III/V) immobilization in groundwater" and Schaefer "Manganese, Arsenic, and Carbonate Interactions in Model Oxic Groundwater Systems".
Regarding claim 1, Xu discloses a method for preparing a supported manganese carbonate material, comprising: dissolving manganese sulfate with water to obtain a manganese sulfate solution, and separately dissolving ammonium bicarbonate with water to obtain an ammonium bicarbonate solution as recited in step (S1) (Xu p. 2-4: “deionized water dissolves manganese sulfate” and “with deionized water at room temperature dissolve … ammonium hydrogen carbonate”); Xu further teaches adding a solid particulate seed to the manganese sulfate solution and adding the ammonium bicarbonate solution to the manganese sulfate solution while reacting under magnetic stirring to obtain a manganese carbonate precipitate substantially recited as step (S2)(Xu p. 4 items 4 and 6 defines “positive addition” as adding the precipitating reagent into the manganese-salt solution and expressly states that NH4HCO3 solution is added into MnSO4 solution; Xu, p.7 embodiment 9 places a pretreated MnCO3 seed directly into the MnSO4 solution, adds the NH4HCO3 solution, and reacts the mixture under a magnetic stirrer for ten minutes; Xu p. 4 item 8 additionally teaches that the rate of introducing the solutions may be controlled using rapid or slow pumping, thereby teaching controlled addition of the ammonium bicarbonate solution to the manganese sulfate solution); Xu further teaches washing the first precipitate with deionized water followed by drying in a vacuum drying chamber to obtain the supported manganese carbonate material substantially recited as step (S3)(Xu p. 4-7 e.g. embodiment 9 steps 3-4 repeatedly washing the precipitate with deionized water and vacuum drying it at 60° C for six hours).
Xu does not expressly teach that the particulate seed added to the MnSO4 solution is ferrihydrite, such that the resulting product is specifically a manganese carbonate-supported ferrihydrite material. Xu also does not expressly characterize its controlled introduction of NH4HCO3 by the particular term “dropwise”.
Zhao teaches that iron and manganese oxide components were known to form a combined remediation material in which the iron oxide component was specifically two-line ferrihydrite. Zhao reports that the resulting biogenic Fe-Mn oxide was a bacteria-mineral complex composed of two-line ferrihydrite and biogenic δ-MnO2 and exhibited removal efficiencies of approximately 83% and 82% for As(III) and As(V), respectively (Zhao abstract and Fig. 5). Zhao further determines that the combined ferrihydrite/manganese oxide material removed substantially more arsenic than either component individually and expressly attributes that result to a synergistic effect between the Fe and Mn oxide components (Zhao p. 8 Fig. 7-8). Zhao thus teaches that combining a manganese-containing phase with ferrihydrite provides a useful composite in which the manganese component contributes arsenic oxidation and ferrihydrite provides strong adsorption and immobilization or arsenic.
Schaefer further teaches that ferrihydrite provides a reactive surface for retaining and transforming manganese in a bicarbonate-containing aqueous system. Schaefer conducted column experiments using ferrihydrite-coated sand and solutions containing Mn(II), bicarbonate, and arsenic, and reports that addition of bicarbonate increased manganese retention and that ferrihydrite surface sites were involved in catalyzing Mn(II) oxidation (Schaefer abstract, p. 10621-10623, Table 1). Schaefer additionally explains that dissolved manganese concentrations may be controlled through MnCO3 precipitation in bicarbonate-containing systems and reports substantially greater solid-phase manganese retention when bicarbonate was present (Schaefer p. 10622-10623, Table 1), teaching that ferrihydrite is a suitable reactive surface for association and retention of manganese under carbonate/bicarbonate conditions of the type used by Xu to precipitate MnCO3.
It would have been obvious to one of ordinary skill in the art prior to the effective
filing date of the claimed invention to use ferrihydrite as the particulate seed in Xu’s seeded MnCO3 precipitation process because Zhao teaches that combining ferrihydrite with a manganese-containing phase provides synergistically enhanced arsenic remediation through complementary manganese-mediated oxidation and ferrihydrite-mediated adsorption, while Schaefer teaches that ferrihydrite provide reactive surface sites for manganese retention and transformation in bicarbonate-containing aqueous systems. A person of ordinary skill in the art would have reasonably expected that precipitating Xu’s MnCO3 in the presence of ferrihydrite would associate the MnCO3 with the ferrihydrite surface and produce a manganese carbonate-supported ferrihydrite composite having the complementary remediation properties taught by Zhao. It further would have been obvious to introduce Xu’s NH4HCO3 solution dropwise because Xu expressly recognizes the feeding manner and charging rate as controllable precipitation parameters, including slow pumping, and a person of ordinary skill would have routinely selected a gradual or dropwise addition rate to control local supersaturation, nucleation, particle growth, and uniform deposition of MnCO3 on the ferrihydrite particles, with a reasonable expectation of obtaining the supported composite.
Regarding claim 2, Xu in view of Zhao and Schaefer discloses or renders obvious the method of claim 1, wherein in step (S1), a concentration of the manganese sulfate solution is 0.5-1.2 mol/L (Xu p. 2 claim 2, 6 “0.3 ~ 2mol/L” encompasses and includes this range); and a concentration of the ammonium bicarbonate solution is 1.5-2.3 mol/L (Xu p. 2 claims 3,7 “0.1 ~ 2mol/L” encompasses and includes the significant portion of this range).
Regarding claim 3, Xu in view of Zhao and Schaefer discloses or renders obvious the method of claim 1, wherein in step (S2), an addition amount of the ferrihydrite is 50-120 g/L (Xu p. 2 claim 5; p. 4 step 2 teaches that the amount of particulate seed is a controllable parameter and expressly discloses using seed in an amount of 0.1-5% of the final product; Xu p. 3-4 further explains that providing a particulate seed controls MnCO3 nucleation and growth and influences particle size, particle-size distribution, morphology, and product properties; as explained above in the rejection of claim 1 Zhao and Schaefer would have suggested selecting ferrihydrite as Xu’s particulate seed. Once ferrihydrite was selected it’s concentration would have been a result-effective variable because the amount of ferrihydrite determines the available nucleation and support surface, while also affecting dispersion, mixing precipitation, and recovery of the resulting composite. It would have therefore been obvious to determine an effective ferrihydrite concentration through routine optimization and to select an amount within 50-120 g/L to provide sufficient ferrihydrite surface for MnCO3 deposition while maintaining a stirrable and recoverable suspension. No criticality or unexpected result is apparent from the claimed range itself).
Regarding claim 6, Xu in view of Zhao and Schaefer discloses or renders obvious a manganese carbonate-supported ferrihydrite material prepared by the method of claim 1 (as explained above in the rejection of claim 1).
Claim 4 is rejected under 35 U.S.C. 103 as being unpatentable over Xu (CN106517340A: An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below) in view of Zhao "Synergy of Fe and biogenic Mn oxide components mediated by a newly isolated indigenous bacterium to enhance As(III/V) immobilization in groundwater" and Schaefer "Manganese, Arsenic, and Carbonate Interactions in Model Oxic Groundwater Systems" as applied to claim 1 above, and further in view of Wei (CN114797779A:An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below).
Regarding claim 4, Xu in view of Zhao and Schaefer discloses or renders obvious the method of claim 1, including preparing a manganese carbonate-supported ferrihydrite material by precipitating MnCO3 from manganese sulfate and ammonium bicarbonate in the presence of ferrihydrite, followed by washing and drying the resulting precipitate, for the reasons set forth above.
Xu in view of Zhao and Schaefer does not expressly teach the particular method recited in claim 4 for preparing that ferrihydrite, including the claimed quantities, addition conditions, washing, centrifugation, and drying conditions.
Wei teaches preparing ferrihydrite from ferric nitrate and potassium hydroxide under stirring conditions. More particularly, Wei teaches preparing 500 mL of a 0.2 mol/L ferric nitrate solution, which contains approximately 0.1 mol, or approximately 40.4g, of ferric nitrate nonahydrate and thus substantially corresponds to dissolving 40g of ferric nitrate nonahydrate in 500mL of water followed by stirring to obtain a ferric nitrate solution, as recited in step (S2.1) (Wei p. 5-7, examples 1-3). Wei further teaches using a 1 mol/L potassium hydroxide solution and adding approximately 330 mL of that solution to 500 mL of the ferric nitrate solution under stirring conditions, with the final 20mL being added dropwise, substantially corresponding to adding 310mL of the potassium hydroxide solution and thereafter adding additional potassium hydroxide to complete pH adjustment, as recited in steps (S2.2)-(S2.3) (Wei p. 3-6; claims 5-7; example 1). Wei teaches continuously stirring during KOH addition, adjusting the reaction mixture to pH 7-8, and then continuing to stir for 30 minutes to form the ferrihydrite-containing mixture (Wei p. 2, 4 and 6). Wei additionally teaches centrifuging the resulting mixture at 3000-5000 rpm for ten minutes, repeated washing and centrifuging the precipitate with deionized water until electrolyte is removed, and drying the washed precipitate to obtain ferrihydrite (Wei, p 2 and 4-7). Wei therefore expressly teaches the claimed ferric-nitrate/KOH ferrihydrite synthesis framework, including essentially the claimed ferric-nitrate quantity, 1 mol/L KOH, approximately the claimed KOH volume, pH 7-8, stirring, washing, centrifugation and drying.
Wei does not expressly state that the initial KOH portion is added at exactly 100mL/min, that stirring after pH adjustment continues for exactly one hour, that the precipitate is washed specifically two or three times with 0.1 mol/L sodium chloride, or that drying is performed specifically at 80°C for 10-12 hours. Wei nevertheless identifies KOH addition rate, post-addition stirring time, washing conditions, centrifugation conditions, and drying conditions as process parameters used to complete precipitation, remove residual electrolyte, separate the fine ferrihydrite precipitate, and recover a dry product.
It would have been obvious to one of ordinary skill in the art prior to the effective
filing date of the claimed invention to prepare the ferrihydrite used in the Xu-Zhao-Schaefer process according to Wei because Wei expressly teaches producing ferrihydrite from essentially 40 g ferric nitrate nonahydrate in 500 mL water using approximately 330 mL of 1 mol/L KOH, adjustment to pH 7-8, stirring, washing, centrifugation, and drying. A person of ordinary skill in the art would have selected the claimed 100 mL/min KOH addition rate and one-hour stirring period through routine optimization to control neutralization, avoid excessive localized pH, and permit completion and equilibration of ferrihydrite precipitation. Such a person likewise would have selected two to three washes with dilute NaCl, centrifugation at 3000 rpm for 8 to 12 minutes, and drying at 80°C for 10-12 hours as ordinary solid-recovery parameters to remove soluble nitrate and potassium residues, maintain controlled ionic strength during handling of the fine precipitate, provide effective solid-liquid separation, and remove retained water without unnecessarily converting ferrihydrite to a more crystalline iron-oxide phase, with a reasonable expectation of successfully obtaining the ferrihydrite required by the modified Xu process.
Claim 5 is rejected under 35 U.S.C. 103 as being unpatentable over Xu (CN106517340A: An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below) in view of Zhao "Synergy of Fe and biogenic Mn oxide components mediated by a newly isolated indigenous bacterium to enhance As(III/V) immobilization in groundwater" and Schaefer "Manganese, Arsenic, and Carbonate Interactions in Model Oxic Groundwater Systems" as applied to claim 1 above, and further in view of Jiang (WO2011009351: An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below).
Regarding claim 5, Xu in view of Zhao and Schaefer discloses or renders obvious the method of claim 1, including repeatedly washing the precipitate with deionized water at room temperature, and drying the washed precipitate (Xu p. 3-7; p. 4-7 embodiments 1-10).
Xu in view of Zhao and Schaefer does not explicitly disclose wherein in step (S3), the first precipitate is washed 2-4 times successively with 90-100 ℃ distilled water and 20-30 ℃ distilled water; and the drying is performed at 80-90 ℃ in the drying oven for 14-20 h.
Jiang teaches a method for preparing and purifying manganese carbonate using the same MnSO4/NH4HCO3 precipitation chemistry employed by Xu. Jiang teaches adding ammonium bicarbonate to a stirred manganese sulfate solution at 40-80°C, separating the resulting MnCO3 solid, and washing the separated manganese carbonate with hot water at 80-100°C to remove entrained soluble impurities (Jiang abstract and claim 1; also, description Step A). The disclosed 80-100°C range overlaps the claimed 90-100°C range. Jiang further teaches repeatedly washing the MnCO3 precipitate with hot water, including washing the solid twice at 80°C in Example 1 and repeatedly washing the solid with water at approximately 100°C in Example 2 (Jiang Examples 1-2). Jiang additionally teaches drying the washed MnCO3 precipitate in a vacuum oven at 85°C for 16 hours, which falls squarely within the claimed drying temperature of 80-90°C and drying time of 14-20 hours (Jiang Example 2). Jiang therefore teaches that MnCO3 produced from MnSO4 and NH4HCO3 should be repeatedly washed with hot water within a range overlapping the claimed range and then dried under conditions expressly encompassed by claim 5.
Jiang does not expressly state that its repeated hot-water washes are followed by a separate distilled-water wash at 20-30°C. Xu, however, already teaches repeated washing its MnCO3 precipitate with deionized water at room temperature until sulfate is no longer detected. The combined teachings thus provide both the hot water purification wash taught by Jiang and the ambient-temperature purified-water wash taught by Xu.
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to apply Jiang’s hot-water washing and drying procedure to the manganese carbonate-supported ferrihydrite precipitate produced by the Xu-Zhao-Schaefer process because Jiang uses the same MnSO4/NH4HCO3 precipitation chemistry and teaches hot-water washing to purify the resulting MnCO3 by removing entrained soluble reaction products and impurities. A person of ordinary skill in the art would have retained Xu’s subsequent room-temperature deionized water washing as a final rinse to continue removing residual sulfate and other soluble species while cooling the precipitate before drying. Such a person would have selected two to four successive hot and room-temperature washing cycles through routine optimization based on the purity of the wash effluent, as exemplified by Xu’s sulfate testing and Jiang’s repeated washing, and would have used distilled water interchangeably with deionized water as a conventional purified wash liquid. Finally, a person of ordinary skill would have dried the washed precipitate at 85°C for 16 hours because Jiang expressly teaches those conditions, with a reasonable expectation of obtaining a purified, dry manganese carbonate-supported ferrihydrite material.
Claims 7 and 9 are rejected under 35 U.S.C. 103 as being unpatentable over Xu (CN106517340A: An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below) in view of Zhao "Synergy of Fe and biogenic Mn oxide components mediated by a newly isolated indigenous bacterium to enhance As(III/V) immobilization in groundwater" and Schaefer "Manganese, Arsenic, and Carbonate Interactions in Model Oxic Groundwater Systems" as applied to claim 6 above, and further in view of Wang '335 (CN107470335A) and Wang 2015 ("Simultaneous removal of tetracycline hydrochloride and As(III) using poorly-crystalline manganese dioxide").
Regarding claim 7, Xu in view of Zhao and Schaefer discloses or renders obvious the manganese carbonate-supported ferrihydrite material of claim 6 as explained in the rejections of claim 1 and 6 above.
However, Xu in view of Zhao and Schaefer as explained in the rejections of claim 1 and 6 above, does not explicitly disclose a method for treating a wastewater sample polluted by arsenic (As) and an antibiotic by (a) inoculating Pseudomonas putida strain MnBl with an accession number of ATCC 23483 into a culture medium A for culture to obtain a Pseudomonas putida suspension; and (b) adding the Pseudomonas putida suspension into a culture medium B containing the manganese carbonate-supported ferrihydrite material followed by addition of 4-(2-hydroxyethyl)piperazine-l-ethanesulfonic acid (HEPES) and the wastewater sample in a clean bench to realize removal of As and the antibiotic.
Wang ‘335 teaches a method for treating arsenic contamination using Pseudomonas putida strain MnB1 having accession number 23483, as recited in step (a) (Wang ‘335 p. 3-5). Wang ‘335 inoculates MnB1 into a first nutrient-broth culture medium and cultures the strain under shaking conditions to obtain an enriched MnB1 bacterial suspension (Wang ‘335 p. 3-5; e.g., inoculating MnB1 at 1-5% by volume into 100-500 mL of nutrient-broth medium and culturing it in a horizontal shaker at 20-30°C). Wang ‘335 further teaches adding the resulting MnB1 suspension to a second culture medium containing a manganese carbonate material, substantially as recited in step (b). Specifically, Wang ‘335 prepares a biological iron-manganese culture medium containing ammonium ferrous sulfate, yeast extract, sodium citrate, and sodium pyrophosphate; inoculates the medium with the enriched MnB1 suspension; adds 0.1-10g of manganese carbonate; and cultures the mixture aerobically to obtain a manganese-carbonate-supported biological iron-manganese remediation material (Wang ‘335 p. 3-5). Wang ‘335 then applies the resulting suspension to arsenic-contaminated material to immobilize phenylarsenic compounds and secondary products including arsenite and arsenate (Wang ‘335 p. 3-5). Wang ‘335 thus teaches the claimed MnB1 strain, cultivation in a first medium to obtain a bacterial suspension, addition of that suspension to a second culture medium containing manganese carbonate, and use of the resulting manganese-containing biological system to remove or immobilize arsenic.
Wang ‘335 does not expressly teach substituting the manganese carbonate-supported ferrihydrite material of claim 6 for its ordinary manganese carbonate, adding HEPES and an arsenic-and-antibiotic-polluted wastewater sample in a clean bench, or treating arsenic and an antibiotic simultaneously. The claimed manganese carbonate-supported ferrihydrite material is supplied by Xu in view of Zhao and Schaefer, as explained above.
Zhao further teaches using HEPES in a culture medium for generating a combined ferrihydrite/biogenic manganese oxide remediation material. Specifically, Zhao uses a modified minimal-salt medium containing yeast extract and 15 mM HEPES at pH 7.20, sterilizes the medium before use, and separately sterilizes the HEPES solution by filtration before adding it to the biological system (Zhao §2.2). Zhao further teaches using the resulting ferrihydrite/biogenic manganese oxide material to treat arsenic-contaminated artificial groundwater and reports synergistically enhanced arsenic immobilization by the combined iron and manganese phases (Zhao abstract §§2.5 and 3.5; Figs. 7-8). Zhao therefore teaches adding HEPES under aseptic conditions to a biological ferrihydrite/manganese treatment medium and applying that material to arsenic-contaminated water.
Wang 2015 teaches treating wastewater-like aqueous contamination containing both arsenic and an antibiotic using a poorly crystalline manganese oxide material. In particular, Wang investigates the simultaneous removal of tetracycline hydrochloride and As(III) from water using poorly crystalline MnO2 and reports that both contaminants were effectively oxidized and removed by the manganese oxide (Wang 2015 p. 103-107, Figs. 1-6). Wang therefore teaches that manganese-oxide treatment systems are suitable for combined aqueous pollution containing both arsenic and an antibiotic.
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to use the manganese carbonate-supported ferrihydrite material supplied by the Xu-Zhao-Schaefer combination in Wang ‘335’s MnB1 treatment method because Zhao teaches that combining ferrihydrite with a manganese-containing phase provides synergistically improved arsenic treatment, while the claimed material would continue to provide the manganese carbonate source used by Wang ‘335 for biological manganese-oxide formation. It further would have been obvious to add HEPES under aseptic conditions, including in a clean bench, because Zhao teaches HEPES in a sterilized biological Fe-Mn treatment medium, and use of a clean bench would have been a conventional aseptic practice for preventing contamination while combining sterile bacterial cultures, media, buffers, and wastewater samples. Finally, a person of ordinary skill in the art would have applied the resulting MnB1/ferrihydrite/manganese system to wastewater containing both arsenic and an antibiotic because Wang expressly demonstrates that poorly crystalline manganese oxide can simultaneously oxidize and remove arsenic and tetracycline from water, with a reasonable expectation that the biological manganese oxide generated by Wang ‘335’s MnB1 system, together with the arsenic-adsorbing ferrihydrite taught by Zhao, would remove both contaminants.
Regarding claim 9, Xu in view of Zhao and Schaefer and further in view of Wang ‘335 and Wang 2015 discloses or renders obvious the method of claim 7, wherein in step (b), an addition amount of the Pseudomonas putida suspension is 3-5% by volume of the culture medium B (Wang ‘335 claim 1 and p. 4-5 inoculates the biological iron-manganese culture medium with the enriched MnB1 suspension at 1-5% by volume); an addition amount of the manganese carbonate-supported ferrihydrite material is 0.1-2.0 g/L (Zhao §2.5 adds collected ferrihydrite/biogenic manganese oxide material to artificial groundwater at 0.2g/L, which falls within the claimed range); the culture medium B comprises 0.1-0.4 g/L of ammonium ferrous sulfate (Wang ‘335 p. 4-6 Examples 1-2 “0.15g” in “1L of deionized water”), 0.1-0.4 g/L of sodium citrate (Wang ‘335 p. 4-6 Examples 1-2 “0.15g” in “1L of deionized water”), 0.05-0.2 g/L of yeast extract powder (Wang ‘335 p. 4-6 Examples 1-2 “0.075g”) and 0.02-0.1 g/L of sodium pyrophosphate (Wang ‘335 p. 4-6 Examples 1-2 “0.05g”), and a pH of the culture medium B is 6-8 (Wang ‘335 claim 1 step 3 “pH value is 6.5-7.5”); an initial concentration of As in the wastewater sample is 0.5-3 mg/L (Zhao §2.5 “1000 µg/L” or 1 mg/L); and an initial concentration of the antibiotic in the wastewater sample is 10-40 mg/L (Wang 2015 p. 103 and 106-107, Table 1 and Fig. 6 uses TC or Tetracycline hydrochloride as “50µM” which corresponds to 24 mg/L).
Claim 8 is rejected under 35 U.S.C. 103 as being unpatentable over Xu (CN106517340A: An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below) in view of Zhao "Synergy of Fe and biogenic Mn oxide components mediated by a newly isolated indigenous bacterium to enhance As(III/V) immobilization in groundwater" and Schaefer "Manganese, Arsenic, and Carbonate Interactions in Model Oxic Groundwater Systems", and further in view of Wang '335 (CN107470335A:An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below) and Wang 2015 ("Simultaneous removal of tetracycline hydrochloride and As(III) using poorly-crystalline manganese dioxide") as applied to claim 7 above, and further in view of Wang '380 (US11738380B2:An English machine translation is provided with this office action and is used for claim mapping in the prior art rejection below).
Regarding claim 8, Xu in view of Zhao and Schaefer and further in view of Wang ‘335 and Wang 2015 discloses or renders obvious the method of claim 7.
Xu in view of Zhao and Schaefer and further in view of Wang ‘335 and Wang 2015 does not explicitly disclose wherein in step (a), the culture medium A comprises 0.3-0.8 g/L of yeast extract, 0.2-0.8 g/L of casein acid hydrolysate, 0.3-0.8 g/L of glucose, 0.1-0.4 g/L of calcium chloride, 0.1-0.6 g/L of magnesium sulfate, 1-5 mL/L of a trace element and 1-3 mL/L of ferric chloride, and a pH of the culture medium A is 7;and the culture is performed through steps of:
inoculating the Pseudomonas putida strain MnB1 into the culture medium A with an inoculation amount of 3-5% by volume followed by culture under shaking and aerobic conditions at 25-35 °C for 1-2 days.
Wang ‘380 teaches culturing Pseudomonas putida MnB1 having accession number ATCC 23483 in a culture medium comprising, based on one liter of deionized water, 0.03-0.08 wt.% yeast extract, 0.02-0.08wt.% hydrolyzed casein, 0.03-0.08 wt.% glucose, 0.01-0.04 wt.% calcium chloride, 0.01-0.06 wt.% magnesium sulfate, and 0.01-0.05% by volume trace elements (Wang ‘380 claim 1). In an aqueous medium, these disclosed quantities correspond to approximately 0.3-0.8 g/L yeast extract, 0.2-0.8 g/L casein hydrolysate, 0.3-0.8 g/L glucose, 0.1-0.4 g/L calcium chloride, 0.1-0.6 g/L magnesium sulfate, and 1-5 mL trace-element solution, respectively, thereby teaching the corresponding compositional ranges recited in claim 8. Wang ‘380 further teaches inoculating MnB1 into the culture medium at 2-10% by volume and culturing it under aerobic conditions at 15-35°C for 1-5 days (Wang ‘380 claim 1). More particularly, Examples 3 teaches inoculating MnB1 at 3% by volume, followed by shaking and aerobic culture at 35°C for two days, thereby expressly falling within the claimed inoculation amount of 3-5%, temperature of 25-35°C, and culture duration of one to two days (Wang ‘380 col. 4 Example 3). Wang ‘380 therefore teaches the claimed culture-medium components and substantially all of the claimed MnB1 culture conditions. Wang ‘335, already included in the combination, teaches preparing its MnB1 nutrient-broth medium at pH 7-7.4, thereby encompassing the claimed pH of 7 (Wang ‘335 p. 3-5).
Wang ‘380 does not expressly teach adding 1-3 mL/L of a ferric-chloride solution to the MnB1 enrichment medium. Nevertheless, Wang ‘380 expressly uses an iron-containing reagent, ammonium ferrous sulfate, in its MnB1-based biological iron-manganese treatment system (Wang ‘380 abstract col. 3-4 examples 1-3), and Zhao likewise teaches supplying Fe(II) to a bacterial medium to generate a ferrihydrite-containing biogenic Fe-Mn material (Zhao §§ 2.2-2.3). The collective art therefore teaches intentionally supplying an iron source to Mn-oxidizing bacterial systems to promote formation of an iron-manganese treatment material.
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to use Wang ‘380’s expressly disclosed MnB1 enrichment medium and culture conditions in the method of claim 7 because Wang ‘380 employs the same Pseudomonas putida MnB1 strain for biological Fe-Mn formation and arsenic remediation and identifies medium composition, inoculum amount, temperature, duration, shaking, and aerobic conditions suitable for propagating that strain. A person of ordinary skill in the art would have adjusted the medium to pH 7 as taught by Wang ‘335 and supplied an iron-containing nutrient stock, including ferric chloride, in an amount of 1-3 mL/L because Wang ‘380 and Zhao teach providing iron in MnB1 or other manganese-oxidizing bacterial systems to form combined iron-manganese remediation materials. Selection of ferric chloride as a soluble iron source and adjustment of the stock-solution volume would have involved routine culture-medium optimization to provide sufficient bioavailable iron while maintaining approximately neutral pH and viable bacterial growth. Moreover, because claim 8 does not specify the concentration of the ferric-chloride stock solution, the recited 1-3 mL/L volume would represent an ordinary medium-preparation parameter that a skilled person would have adjusted according to the selected stock concentration, with a reasonable expectation of successfully culturing MnB1 and forming the desired biological iron-manganese treatment system.
Conclusion
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/W.A.G./Examiner, Art Unit 1779
/Bobby Ramdhanie/Supervisory Patent Examiner, Art Unit 1779
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