CN102212594A - Method for producing xylose with straw - Google Patents

Abstract

The invention discloses a method for degrading the hemicellulose prepared from straw by utilizing the xylanase which is fermented by utilizing actinobacteria. The technical scheme in the invention comprises four main working procedures, namely the preparation of xylanase, the preparation of hemicellulose, the enzymolysis of hemicellulose and the purification and refinement of xylose. The enzymatic method is adopted to produce xylose; and the method has the following advantages: the specificity is high, the conditions are mild, the impurities of the intermediate are fewer, the intermediate is easy to purify and extract, the conversion rate of hemicellulose and the yield of xylose crystals are higher, clean and environmentally-friendly production can be maintained in industry, and the like. The total raw material conversion rate is more than 15% and the xylose content of the finished product is more than 98.5%.

Description

A kind of method of utilizing producing xylose with stalks

Technical field:

The invention belongs to the production method of wood sugar, be specifically related to a kind of method of utilizing producing xylose with stalks.

Background technology:

Wood sugar is a kind of white, needle-shaped crystals powder, and it is sweet to distinguish the flavor of, and sugariness is equivalent to 40% of sucrose.Soluble in water, 147～151 ℃ of fusing points have dextrorotation photosensitiveness and mutarotation photosensitiveness.

Wood sugar has following major function as a kind of base-material of functional foodstuff: (1) is not is not digested and assimilated, and does not have Energy value, can satisfy to greatest extent and like to eat the demand that sweets is worried the person of getting fat again; (2) bifidus bacillus in the activating human body enteron aisle and promote its growth; (3) do not utilized by Institute of Micro-biology in the oral cavity, but prevention of dental caries; (4) possess the part physiological function of food fibre, reducing blood-fat is arranged, the effect of reducing cholesterol and prevention intestinal cancer; (5) compatibleness of wood sugar and food is fine, adds a small amount of wood sugar in the food, just can embody good health-care effect; (6) wood sugar and calcium are taken in simultaneously, can improve specific absorption and the retention rate of human body to calcium, can also prevent constipation.

Present industrial production wood sugar mainly is to adopt mineral acid hydrolysis to be rich in the agricultural waste of hemicellulose and produce.Under certain pressure, temperature, mineral acid decompose to produce wood sugar with raw material, again through depickling, decolouring, ion-exchange, concentrate, operation such as crystallization, obtain wood sugar.Acid system is produced wood sugar and is mainly had following shortcoming:

(1) contain assorted sugar such as a certain amount of glucose, pectinose in the hydrolyzed solution, the viscosity that concentrates the back liquid glucose is bigger, influences the crystallization yield of wood sugar.

(2) gypsum (CaSO that forms in and in the deacidifying process ₄) finally understand on some tube wall that is deposited on vaporizer, be difficult to remove, equipment also there are in various degree damage, the work-ing life of reducing equipment.Bring calcium ion simultaneously into, increased the burden of Zeo-karb.The ion-exchange deacidifying process, though overcome the shortcoming of neutralisation, its technology more complicated, the ion exchange resin consumption is more, and equipment is more, and investment is big, has increased acid and alkali consumption, has strengthened cost.

(3) acid system is produced in the process of wood sugar, has used a large amount of strong acid and strong bases, has produced a large amount of waste liquids, and environment is caused huge pollution.

Summary of the invention

The present invention will solve traditional xylose production process complexity, and the ion exchange resin consumption is many, and the equipment that needs is many, and cost is big, and the liquid glucose viscosity that environment is easily polluted and produces is bigger, the problem that crystallization yield of xylose is low.

The present invention is achieved through the following technical solutions:

A kind of method of utilizing producing xylose with stalks, its production stage is as follows: (1) actinomycetes strain of wanting required for the present invention obtains in the following manner: collected specimens from the corn field compost of suburb, Yuci, Shanxi, adopt conventional microorganism separation screening technology, the sample of getting 1～10g places 50～100ml sterilized water to carry out gradient dilution, and the diluent of drawing 0.1ml～0.5ml is coated on the hemicellulose decomposer screening flat board, the pH value 6.5 of hemicellulose decomposer screening culture medium, hemicellulose decomposer screening culture medium is made up of the component of following weight content: (NH ₄) ₂SO ₄0.5 part, K ₂HPO ₄1.0 part, MgSO ₄7H ₂0.3 part of O, CaCl ₂2H ₂O0.2 part, K ₂SO ₄0.1 part, 0.2 part of NaCl, 15 parts of hemicelluloses, 1000 parts in tap water, in 36 ℃ of cultivation 72h, the strain fermentation degradation of hemicellulose that the choosing colony transparent circle is big is chosen the high a plurality of bacterial strains mutagenesis repeatedly of Xylanase activity, this bacterial strain of screening acquisition;

(2) earlier the actinomycetes bacterial classification is activated on the synthetic medium slant of Gao Shi, 35～40 ℃, cultivate 24h～48h; Get an inoculation hexamethylene then and grow good lawn access liquid seed culture medium, 35～40 ℃, 200 rev/mins, shake-flask culture 24～36h, form seed culture fluid, liquid seed culture medium pH value is 7.0～8.0, and liquid seed culture medium is made up of the component of following weight content: 20～50 parts of glucose, KNO ₃1～3 part, K ₂HPO ₄0.2～1 part, MgSO ₄7H ₂0.2～1 part of O, 0.2～1 part of NaCL, FeSO ₄0.01～0.1 part, 1000 parts in tap water inserts fermention medium with the ratio of 10%～20% (V/V) with seed culture fluid, 35～40 ℃, 100～200 rev/mins, shake-flask culture 48～96h, fermention medium pH value is 5.0～8.0, and fermention medium is made up of the component of following weight content: K ₂HPO ₄1～3 part, NH ₄NO ₃1～3 part, MgSO ₄7H ₂0.1～1 part of O, 20～50 parts of corn steep liquors, 50～100 parts of stalks, 1000 parts in tap water; Treat fermentation ends, filter and remove thalline, stalk residue that promptly get zytase liquid, enzyme activity reaches 2100U/mL;

(3) be 2～5mm with crushed stalk to particle diameter, in stalk (Kg): the ratio of NaOH solution (L)=1: 6～8, add 3～6%NaOH solution, 80～100 ℃, keep 3～5h, with dense HCL accent pH to 3.5～5.5, promptly get the hemicellulose feed liquid;

(4) the hemicellulose feed liquid is cooled to 40～50 ℃, adds zytase liquid, make that final enzyme activity reaches 200～400U/mL in the feed liquid, 45～55 ℃, enzyme digestion reaction 2～10h obtains crude xylose solution;

(5) with the ratio adding gac of crude xylose solution, be warming up to 60～70 ℃, keep 20～30min, filter, filtrate is evaporated to wood sugar content 20～40% at 60 ℃ in 0.2～0.5% (W/V); Select for use 732 type Zeo-karbs and 301 type anionite-exchange resin to carry out ion-exchange; Ion exchange liquid is concentrated into not water outlet, adds raw spirit, make that wood sugar content reaches 20～40% (W/V) in the feed liquid, filter, filtrate can be evaporated to alcohol at 60 ℃; With the cooling of dense syrup, be cooled to 5～10 ℃, crystalline xylose is separated out, and 4000 rev/mins, 10min is centrifugal to get rid of filter, with cold alcohol washed twice, 50～70 ℃ of oven dry of crystalline xylose of will wetting, pulverizing is finished product.

The present invention compared with prior art has following beneficial effect: the zytase that this law adopts a kind of actinomycete fermentation to produce is degraded by the hemicellulose of stalk preparation, generates thick wood sugar, obtains crystalline xylose through purification again.Assorted sugared content such as glucose, seminose, pectinose are few in the xylose crystallization mother liquor, and dense syrupy viscosity is less, thereby can obtain higher crystallization yield of xylose.This method technology is simple, working condition is gentle, the burden of aftertreatment alleviates in the purification process of wood sugar, the load of ion exchange resin diminishes, thereby reduced the consumption of the consumption and the soda acid of resin, the quantity discharged of acidic and alkaline waste water significantly reduces in the production process, has reduced the pollution to environment, is a kind of production method of clean.

Embodiment:

A kind of method of utilizing producing xylose with stalks, its production stage is as follows:

(1) actinomycetes strain of wanting required for the present invention obtains in the following manner: collected specimens from the corn field compost of suburb, Yuci, Shanxi, adopt conventional microorganism separation screening technology, the sample of getting 1g places the 99ml sterilized water to carry out gradient dilution, and the diluent of drawing 0.1ml～0.5ml is coated on the hemicellulose decomposer screening flat board, the pH value 6.5 of hemicellulose decomposer screening culture medium, hemicellulose decomposer screening culture medium is made up of the component of following weight content: (NH ₄) ₂SO ₄0.5g, K ₂HPO ₄1.0g, MgSO ₄7H ₂O 0.3g, CaCl ₂2H ₂O 0.2g, K ₂SO ₄0.1g, NaCl 0.2g, hemicellulose 15g, tap water 1000ml, in 36 ℃ of cultivation 72h, the strain fermentation degradation of hemicellulose that the choosing colony transparent circle is big is chosen the high a plurality of bacterial strains mutagenesis repeatedly of Xylanase activity, this bacterial strain of screening acquisition.

(2) elder generation is connected to bacterial classification on the synthetic medium slant of Gao Shi and activates, and 36 ℃, cultivates 24h.

Press glucose 20g, KNO ₃1g, K ₂HPO ₄0.5g, MgSO ₄7H ₂O 0.5g, NaCL 0.5g, FeSO ₄0.01g, tap water 1000ml, pH 7.0～8.0 obtaining liq seed culture mediums, 121 ℃ of sterilization 20min.An inoculation hexamethylene activated spawn is inserted in the cooling back, substratum loading amount 50mL/250mL, and 36 ℃, 200 rev/mins, shake-flask culture 24h is seed culture fluid.

Press K ₂HPO ₄2g, NH ₄NO ₃2g, MgSO ₄7H ₂O 0.2g, corn steep liquor 50g, stalk 100g, tap water 1000ml, pH 7.2 preparation fermention mediums were sterilized 20 minutes for 121 ℃.After the cooling, the ratio with 10% inserts seed culture fluid, substratum loading amount 50mL/250mL, 36 ℃, 200 rev/mins, shake-flask culture 72h.Treat fermentation ends, filter solid substances such as removing thalline, stalk residue, promptly get required zytase liquid, enzyme activity can reach more than the 2100U/mL.

(3) preparation hemicellulose

It is 2～5mm that the 2000g maize straw is crushed to particle diameter, adds 4%NaOH solution 14000mL, 90 ℃, and insulation 4h.Transfer pH to 4.5 with dense HCL, get the hemicellulose feed liquid.

(4) enzymolysis hemicellulose

The hemicellulose feed liquid is cooled to 50 ℃, keeps constant temperature, add 1500mL zytase liquid, enzyme digestion reaction 10h gets crude xylose solution.

(5) with the ratio adding gac of crude xylose solution, be warming up to 60 ℃, keep 30min in 0.5% (W/V).Filter, filtrate is evaporated to wood sugar content 20% at 60 ℃.Select for use 732 type Zeo-karbs and 301 type anionite-exchange resin to carry out ion-exchange, ion exchange liquid is concentrated into not water outlet.Add raw spirit, make wood sugar content reach 20% (W/V).Filter, filtrate can be evaporated to alcohol at 60 ℃.With dense syrup cooling, be cooled to 10 ℃, crystalline xylose is separated out.4000 rev/mins, 10min is centrifugal to get rid of filter.With cold alcohol washed twice.The 60 ℃ of oven dry of crystalline xylose of will wetting are pulverized.Packing is finished product after the assay was approved, gets 301g.Wood sugar purity is more than 98.5%, total conversion rate 15%.

Claims (1)

1. method of utilizing producing xylose with stalks, it is characterized in that: production stage is as follows:

(1) actinomycetes strain of wanting required for the present invention obtains in the following manner: collected specimens from the corn field compost of suburb, Yuci, Shanxi, adopt conventional microorganism separation screening technology, the sample of getting 1～10g places 50～100ml sterilized water to carry out gradient dilution, and the diluent of drawing 0.1ml～0.5ml is coated on the hemicellulose decomposer screening flat board, the pH value 6.5 of hemicellulose decomposer screening culture medium, hemicellulose decomposer screening culture medium is made up of the component of following weight content: (NH ₄) ₂SO ₄0.5 part, K ₂HPO ₄1.0 part, MgSO ₄7H ₂0.3 part of O, CaCl ₂2H ₂0.2 part of O, K ₂SO ₄0.1 part, 0.2 part of NaCl, 15 parts of hemicelluloses, 1000 parts in tap water, in 36 ℃ of cultivation 72h, the strain fermentation degradation of hemicellulose that the choosing colony transparent circle is big is chosen the high a plurality of bacterial strains mutagenesis repeatedly of Xylanase activity, this bacterial strain of screening acquisition;

(2) actinomycetes strain that obtains is activated on the synthetic medium slant of Gao Shi, 35～40 ℃, cultivate 24h～48h, get the long good lawn of a transfering loop then and insert liquid seed culture medium, 35～40 ℃, 200 rev/mins, shake-flask culture 24～36h forms seed culture fluid, and liquid seed culture medium pH value is 7.0～8.0, liquid seed culture medium is made up of the component of following weight content: 20～50 parts of glucose, KNO ₃1～3 part, K ₂HPO ₄0.2～1 part, MgSO ₄7H ₂0.2～1 part of O, 0.2～1 part of NaCL, FeSO ₄0.01～0.1 part, 1000 parts in tap water inserts fermention medium with the ratio of 10%～20% (V/V) with seed culture fluid, 35～40 ℃, 100～200 rev/mins, shake-flask culture 48～96h, fermention medium pH value is 5.0～8.0, and fermention medium is made up of the component of following weight content: K ₂HPO ₄1～3 part, NH ₄NO ₃1～3 part, MgSO ₄7H ₂0.1～1 part of O, 20～50 parts of corn steep liquors, 50～100 parts of stalks, 1000 parts in tap water; Treat fermentation ends, filter and remove thalline, stalk residue that promptly get zytase liquid, enzyme activity reaches 2100U/mL;

(3) be 2～5mm with crushed stalk to particle diameter, in stalk (Kg): the ratio of NaOH solution (L)=1: 6～8, add 3～6%NaOH solution, 80～100 ℃, keep 3～5h, with dense HCL accent pH to 3.5～5.5, promptly get the hemicellulose feed liquid;

(4) the hemicellulose feed liquid is cooled to 40～50 ℃, adds zytase liquid, make that final enzyme activity reaches 200～400U/mL in the feed liquid, 45～55 ℃, enzyme digestion reaction 2～10h obtains crude xylose solution;

(5) with the ratio adding gac of crude xylose solution, be warming up to 60～70 ℃, keep 20～30min, filter, filtrate is evaporated to wood sugar content 20～40% at 60 ℃ in 0.2～0.5% (W/V); Select for use 732 type Zeo-karbs and 301 type anionite-exchange resin to carry out ion-exchange; Ion exchange liquid is concentrated into not water outlet, adds raw spirit, make that wood sugar content reaches 20～40% (W/V) in the feed liquid, filter, filtrate can be evaporated to alcohol at 60 ℃; With the cooling of dense syrup, be cooled to 5～10 ℃, crystalline xylose is separated out, and 4000 rev/mins, 10min is centrifugal to get rid of filter, with cold alcohol washed twice, 50～70 ℃ of oven dry of crystalline xylose of will wetting, pulverizing is finished product.