The use of continuous‐flow electrophoresis to remove pectin in the purification of the minor pectinesterases in lemon fruits (citrus limon)

Macdonald, Helen; Evans, Roger; Spencer, Wendy J

doi:10.1002/jsfa.2740640119

Cited by 15 publications

(22 citation statements)

References 5 publications

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“…The low K m value calculated for carrot PE indicates that this enzyme has a high level of activity in its substrate. This K m value was very close to those of PE isoforms of lemon,31 Vigna sinensis 32 and persimmon 33. However, the K m value calculated here was much lower than the value of ∼0.4 mg ml −1 calculated previously for carrot PE,21 probably owing to the different experimental conditions and substrate employed.…”

Section: Resultssupporting

confidence: 73%

Purification and characterisation of carrot (Daucus carota L) pectinesterase

et al. 2003

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A pectinesterase isoform with an alkaline isoelectric point of over 8.66 was detected in crude extracts of carrot. The enzyme was purified by ion exchange and molecular exclusion chromatography. The molecular weight of the isoform was 25 kDa, determined in native conditions by filtration through Sephadex G-75 SF. The enzyme showed a high affinity for its substrate, with K m and V max values of 0.031 mg ml −1 and 6.77 units respectively for apple pectin. The pectinesterase activity exhibited an optimum around pH 7.4 and was activated by metallic ions, with optimum activities at NaCl concentrations between 130 and 330 mM and at CaCl 2 concentrations between 15 and 50 mM. The enzyme was activated most by Ca 2+ and exhibited a greater tolerance of high concentrations of Na + . Comparison of its heat stability with other pectinesterases of vegetable origin indicated that the purified isoform was very thermolabile, being rendered inactive by heating for 5 min at 70 • C. The enzyme was inhibited by high concentrations of polygalacturonic acid and competitively inhibited by D-galacturonic acid, with a K i value of 1 mM.

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Section: Resultssupporting

confidence: 73%

Purification and characterisation of carrot (Daucus carota L) pectinesterase

et al. 2003

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“…One form, purified from a commercial 'Fresh Frozen Orange Juice' was relatively thermally stable, retaining ϳ50% activity after 60 sec incubation in a 90ЊC water bath (Cameron and Grohmann, 1996). Thermally tolerant forms of PME (TT-PME) also have been identified from navel oranges (Versteeg et al, 1980), white grapefruit (Seymour et al, 1991), citrus tissue culture cells (Cameron et al, 1994), lemon fruit (Macdonald et al, 1994), and red grapefruit (Cameron and Grohmann, 1995). Versteeg et al (1980) reported that at 5ЊC only a TT-PME was capable of destabilizing juice cloud in Ͻ4 wk (see Guyer et al, 1956and Carroll et al, 1957 for discussion and definition of juice cloud clarification).…”

Section: Introductionmentioning

confidence: 99%

“…Multiple forms of PME have been identified in various citrus fruits (Versteeg et al, 1980;Seymour et al, 1991;Macdonald et al, 1993Macdonald et al, , 1994Cameron et al, 1994;Grohmann, 1995, 1996). One form, purified from a commercial 'Fresh Frozen Orange Juice' was relatively thermally stable, retaining ϳ50% activity after 60 sec incubation in a 90ЊC water bath (Cameron and Grohmann, 1996).…”

Section: Introductionmentioning

confidence: 99%

Citrus Tissue Extracts Affect Juice Cloud Stability

Cameron

Baker

Grohmann

1997

Journal of Food Science

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Salt extractable proteins were isolated from hand expressed juice, rag and peel of Valencia oranges. Each tissue extract was divided into dialysis supernatant (DS) and precipitate (DP) (forms during dialysis). DP, DS and DS heated for 2 min at 80ЊC (HDS) were added to pasteurized, reconstituted frozen concentrated orange juice at 5 U ⅐ mL Ϫ1 of pectinmethylesterase. Samples were incubated either at 25ЊC for 14 days or 4ЊC for 28 days and periodically sampled to determine the effects of tissue extracts on juice cloud stability. Tissue specific differences were observed for the rate of juice cloud precipitation and among the HDS, DS, and DP fractions of a given tissue. HDS fractions destabilized juice cloud more rapidly than DS or DP at both 25 and 4ЊC.

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“…One thermally sensitive form precipitated the juice cloud at 4°C but took -4 wk to produce this effect. However, Macdonald et al (1993) reported that both forms of PME from lemon fruits clarified orange juice, and that a minor lemon PME (Macdonald et al, 1994), which retained activity up to 88"C, precipitated lemon.juice cloud. Seymour et al (1991a,b) reported on thermal properties of PMEs from Marsh white grapefruit finisher pulp.…”

Section: Introductionmentioning

confidence: 99%

Partial Purification and Thermal Characterization of Pectinmethylesterase from Red Grapefruit Finisher Pulp

Cameron¹,

Grohmann²

1995

Journal of Food Science

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Four forms of pectinmethylesterase have been partially purified from red grapefruit finisher pulp by chromatography on Heparin-Sepharose CL-6B. Three forms bound to the heparin-sepharose and could be eluted with a sodium chloride gradient. One form (RGPME3) exhibited high thermal stability retaining 66.7% relative activity after 2 min incubation in an 80°C water bath, and 45.2% of its relative activity after 60 set incubation in a 95°C water bath. RGPME3 had a K,,, of 0.106 mg * mL-> citrus pectin and a pH optimum between 6 and 7. The most abundant form (RGPMEZ) had a K,,, of 0.079 mg * mL-' citrus pectin. Denaturing electrophoresis and gel filtration chromatography suggested molecular weights between 20 and 40 kDa for the various forms.

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The use of continuous‐flow electrophoresis to remove pectin in the purification of the minor pectinesterases in lemon fruits (citrus limon)

Cited by 15 publications

References 5 publications

Purification and characterisation of carrot (Daucus carota L) pectinesterase

Purification and characterisation of carrot (Daucus carota L) pectinesterase

Citrus Tissue Extracts Affect Juice Cloud Stability

Partial Purification and Thermal Characterization of Pectinmethylesterase from Red Grapefruit Finisher Pulp

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