VERC - A Preliminary Study of Industrial Enzyme Preparations for Color Extraction/Stability in Red Wines

- Research Note -

A Preliminary Study of Industrial Enzyme Preparations for Color Extraction/Stability in Red Wines
by
Barry H. Gump and Katherine G. Halght
CATI Publication #950901
© Copyright September 1995, all rights reserved

Introduction

      The color of young red wines is almost entirely due to the monomeric grape anthocyanins. Due to condensation reactions, there is a steady shift towards predominantly polymeric pigment forms in aged red wines, along with a progressive decline in the amount of anthocyanins. These changes can be detected as wine ages, so the color can be regarded as an integration of contributions from monomeric and polymeric pigment forms (12).
      During the 1940s experiments were already being run on the use of enzyme preparations for increased color extraction-stability in red wine (2). Due to anthocyanase activity the treated wines of this time had problems with the red color deteriorating to brown. Some enzyme preparations produced during the mid 1950s had decreased anthocyanase content, and wineries were producing enzyme treated red wines with very satisfactory results (2).
      The technology of enzyme preparations and the benefits of their use are moving at a fast pace. The benefits of enzyme usage are partially dependent on the specific variety of grapes - and the growing conditions - on which the enzyme is used (4). Today's enzyme preparations can promote improved color extraction in the processing of red grapes (8,9,11,13).
      Most commercial pectinase preparations are derived from cultures of Aspergillus, mainly Aspergillus niger (1,3). These preparations are accepted as "generally recognized as safe" (GRAS) formulation and by the federal Bureau of Alcohol, Tobacco, and Firearms (BATF). Macerating type enzyme preparations are typically blends of pectinases, cellulases, hemicellulases, and other carbohydrase activities (5). Cellulases, either as a single preparation or as part of the pectinase preparation, are effective tools for increasing the rate of color extraction from the berry (7). Macerating type preparations are commonly used in world-wide winemaking operations (15), but are not allowed by the BATF for winemaking in the USA. Presently, these types of preparations are GRAS formulations allowed in U.S. fruit juice processing (10,13,14).
      Our objective was to find a pectinase counterpart derived from Aspergillus niger, with equal benefits to the multi-fermentation products used for fruit juice processing in the USA and for winemaking in other countries. Experiments were designed to evaluate the efficacy of four commercial enzyme preparations for color extraction and stability with one of the leading red wine varieties produced in the San Joaquin Valley (Barbera). Our thrust was to evaluate several pectinases with "novel side activities," against a macerating enzyme preparation commonly used in juice production (6,15), and a no-enzyme control. For statistical purposes experiments were designed to run in triplicate with relatively large (22.7 Kg) laboratory scale grape lots. Each lot would be fermented on the skins for one week prior to pressing and malolactic fermentation. Pre- and post-bottling measurements of A520, A420, Hue, and color Intensity were to be monitored to evaluate color levels and stability over time (16).

Materials and Methods

      Grape material: The variety used throughout this trial was Barbera grown in the San Joaquin Valley. These grapes were hand-picked and delivered to the California State University, Fresno winery and processed during September, 1994.
      Enzymes: In this trial Cytolase® PLC5 (Gist-brocades); Pectinex® BE 3-L (Novo Nordisk Ferment); Pectinex® 5XL (Novo Nordisk Ferment); and Vinozym® EC (Novo Nordisk Ferment) were evaluated along with a "no enzyme addition" control.
      Experimental procedures: Each lot was individually weighed and crushed. The experiments were run in triplicate, using 22.7kg replications. The enzyme preparations were first diluted to a 10% solution using cool, clean water, and added [25 ppm enzyme] to the freshly crushed grapes. The "no-enzyme addition" lots received an equal amount of deionized water as a replacement for the enzyme additions. K-1 yeast was prepared and added per package instructions. Each lot was fermented on the skins for one week prior to pressing and malolactic fermentation. No temperature control was used other than the cellar temperature was consistently kept between 19 and 20°C. All lots of wine were treated the same during processing and were bottled on December 3, 1994. Color measurements [A420, A520, Hue (A420//A520), and Intensity (A420 + A520)] were made monthly on all lots and treatments.

Results & Discussion

      In this trial several pectinases derived from Aspergillus niger but containing various "novel side activities" were used. In the first year of this study the enzyme dosage rate was 25 ppm, a rate we consider to be high. This high dosage rate was used to help determine the enzyme preparations that would be tested in future "enzyme dosage rate optimization trials."
      Cytolase® PLC5 (Gist-brocades) was selected because of our previous experience using it for juice processing. This enzyme would not normally be used in red wine production due to some side anthocyanase activity. Pectinex® BE 3-L (Novo Nordisk Ferment) is currently used in current and berry production; it is not advertised for winemaking at this time. Pectinex® 5XL (Novo Nordisk Ferment) is presently being used by the U.S. wine industry at lower dosage rates for its pectinase activity. The macerating enzyme, Vinozym® EC (Novo Nordisk Ferment), was included in the trial, again due to our experience with it in juice processing and as another processing control. These enzyme preparations were evaluated against a "no enzyme addition" in order to provide a "minimal treatment" type of control.
      The color measurements taken over time are representing the time series from November 1994 to June 1995. The color data (A520) "red," (A420) "brown," and Intensity are statistical evaluations [February through June 1995] at significance levels of p = 0.01. During the same period, the Hue data is statistically different at p = 0.05 . The averages of each set of measurements were plotted and are presented as a series of line graphs (Figures 1 to 4). With regard to the values for Hue over time there is obvious consistency. Except for some individual up-and-down variation due to uncertainties in the measurement process, overall Hue values have not changed over the time period from December to June. Following bottling and one month to stabilize Intensity, values have also remained relatively constant. The June 1995 data is the most current data available at publishing [Table I]. We plan to analyze these wines for color again in December 1995.

      The preliminary results of this study demonstrated significantly increased color in bottled wines over the time period studied. These wines will continue to be followed for longer term color stability even as this project continues with additional enzyme preparations and grape varieties. Based on results to date, the most promising enzyme preparation evaluated for color extraction and stability, Pectinex® 5XL, is a pectinase preparation currently available to the U.S. wine industry. Another enzyme preparation that produced significant color increases is Pectinex® BE3L, a pectinase produced by a selected strain of Aspergillus niger and labeled for currants and other berries. Although this product is not advertised for grapes at this time it should be further evaluated.
      In the fall of 1995 we plan to continue this enzyme research using two different grape varieties, Zinfandel and Merlot. In these trials we also plan to evaluate additional enzyme preparations: Rapidase® EX Color (Gist-brocades), and a new enzyme preparation (Valley Research).

LITERATURE CITED:

      1. Canal-LLauberes, R.-M. Enzymes in winemaking. In: Wine Microbiology and Biotechnology. G. H. Fleet (Ed.). pp. 477-506. Harwood Academic Publishers, Philadelphia (1993).

2. Cruess, W. V., R. Quacchia, and K. Ericson. Pectic enzymes in wine making. Food Technology. pp. 601-607 (1955).

3. Grampp, E., and R. A. Urlaub. The application of enzymes in grape juice and wine processing. The International Symposium on Cool Climate Viticulture and Enology. Oregon State Univ. (1984).

4. Haight, K. G. Grape yield and color extraction in response to various enzyme preparations. Thesis: California State University, Fresno (1992).

5. Haight, K. G. Utilization of enzymes in juice and wine production. In: Wine Analysis and Production. Zoecklein et al. Chapman & Hall, New York (1995).

6. Haight, K. G., and B. H. Gump. The use of macerating enzymes in grape juice processing. Am. J. Enol. Vitic. 45:113-116 (1994).

7. Janda, W. Fruit Juice. In: Industrial Enzymology. T. Godfrey and J. Reichelt (Eds.). pp. 315-320. The Nature Press, New York (1983).

8. Ough, C. S., and H. W. Berg. The effect of two commercial pectic enzymes on grape musts and wines. Am. J. Enol. Vitic. 50:208-11 (1974).

9. Ough, C. S., A. C. Noble, and D. Temple. Pectic enzyme effects on red grapes. Am. J. Enol. Vitic. 26:195-200 (1975).

10. Pilnik, W., and A. G. J. Voragen. Pectic substances and other uronides. In: The Biochemistry of Fruits and Their Products. Vol. 1. A.C. Hulme (Ed.). pp.63-87. Academic Press: New York (1970).

11. Schwimmer, S. Source Book of Food Enzymology. pp.535-551. The AVI Publishing Company, Inc. Westport, CT (1981).

12. Somers, T. C. The polymeric nature of wine pigments. Phytochemistry 10:2175-86 (1971).

13. Urlaub, R. Enzymes for Wine Making. Rohm GmbH, Enzyme Technology. Darmstadt, Germany.

14. Rombouts, F. M., and W. Pilnik. Enzymes in fruit and vegetable juice technology. Process Biochemistry 13:9-13 (1978).

15. Zent, J. B., and S. Inama. Influence of macerating enzymes on the quality and composition of wines obtained from Red Valpolicella wine grapes. Am. J. Enol. Vitic. 43:311 (1992).

16. Zoecklein, B. W., K. C. Fugelsang, B. H. Gump, and F. S. Nury. Wine Analysis Production. Chapman and Hall, New York (1995).

Acknowledgements

      We would like to thank the California Agricultural Technology Institute [CATI] for the financial support for this research, and a special thanks to the students who helped with this project [Jeff Poe, Tim Henley, and Chris Lake].

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Copyright © 1995. All rights reserved.
CALIFORNIA AGRICULTURAL TECHNOLOGY INSTITUTE - CATI
College of Agricultural Sciences and Technology
California State University, Fresno