PHYSIOLOGICAL AND CHEMICAL STUDIES QN THE KEEPING QUALITY OF BETTER TIMES ROSES

DISSERTATION

Pr©eontad In Partial Fulfillment of the RequiromentB for the Degree Doctor of Philosophy In the Graduate School of The Ohio State University

By

MARVIN CLINTON TWIGG, B.S., M.S. The Ohio State University 1952

Approved hy:

Adviser -7- condltions.

Reduced Temperature. Die uee of reduced temperatures during

storage periods has long been an accepted practice for roses & b well as for other florlcultural and hortloitural crops. Perret (39)

In 1904 advocated the use of low temperature storage and stressed the Importance of the relative humidity In cut flower storage.

Hitchcock and Zimmerman (lQ) reported that Briarcllff roses kept

two to three times longer at a temperature of 10* C. than they did at room temperature. In their work they did not find that

increased humidity during storage was of any benefit to roses though

it was for carnations. Bancroft (3 ) found that a humidity of 80

percent during storage was best for both carnations and roses.

Light. Neff (3*0 reported on Investigations dealing with the effect of light during storage. He found that Illumination of red

roses Increased keeping quality and reduced or eliminated bluing.

Roses which had blued when stored in the dark regained normal color when exposed to light.

Carbon Dioxide Storage. Thornton (37) subjected roses with

their stems wrapped In moist paper to carbon dioxide concentrations

of 5 to 80 percent. His results showed that when the storage period was longer than one day roses were benefited by carbon dioxide concentrations in the range of 5 to 15 percent. Higher concentrations were injurious resulting in blue spots or browning of petals depending on the concentration of the gas. Reported beneficial effects of the carbon dioxide storage were improved -97-

AUTOBIOqRAFHT

I, Marvin Clinton Twigg, vaa born in Oldtown, Allegany County,

Maryland, September 20, 1921. I received ray secondary school edu­ cation at the Oldtown High School. first two years of under­ graduate training were received at Bridgewater College, Bridgewater,

Virginia. Following a period of 29 months spent with the armed services of the United States I enrolled at the University of Mary­ land where I received the degree Bachelor of Science In Horticulture in 19^8* I was enrolled in the Graduate School of the University of Maryland from 19^8 to 195°• During this time I had an appoint­ ment as graduate assistant in the Department of Horticulture. I received the degree Master of Science in 1950* ln 1950 I received an appointment to a fellowship sponsored by Roses Incorporated at

The Ohio State University. I held this appointment for two years while completing the requirements for the degree Doctor of Philosophy. -8- lastlng qualities, delayed opening of buds, and delayed petal drop.

Hose buds held in a 15 percent concentration of carbon dioxide at

30* F. remained in good condition for seven days and lasted as long at room temperature as untreated buds held three days. In prepackaging work conducted at The Ohio State University, Sir Ingen

(5 5 ) and Koon (2 3 ) found that the enclosed atmosphere of the packages contained approximately 3 to 7 percent carbon dioxide due to the respiration of the flowers. Koon (2 3 ) reported that removing the carbon dioxide from the package as it forma: resulted in the bluing and opening of Better Times rose buds and reduced keeping quality. Longley (3 1 ) reported that 8 to 10 percent carbon dioxide kept roses from opening as rapidly in storage but did not improve their keeping qualities over that of the checks. In experiments conducted by Neff (35) it was found that additions of carbon dioxide caused bluing of red roses at all concentrations tested after 12 to 20 days storage.

Dry Storage. Further investigations by Neff (35) reported in the same paper showed that rose buds could be maintained in the desired stage of maturity during storage at low temperatures by not placing the stems in water or other solutions. Dry storage of carnations was later studied by Neff and Loomis (3 6 ). Ihey found that this was a preferred method of storage.

Post and Fischer (4l) have recommended low temperatures, 31° F., and dry packaging in an air tight container as a means of extending the storage life of a number of cut flowers, including roses. They -9- reported that Better Times roses stored for periods of 18 days or lees under the low temperature-dry storage conditions lasted for five days when placed in water at room temperature. Bluing during storage was eliminated when the cut roses were placed directly in storage rather than first in water.

C. Ihe Use of Chemicals to Prolong Keeping Quality.

The addition of chemicals to the water in which the stems are placed has been a favorite method of attempting to Increase the life and preserving the color and appearance of cut flowers.

Mary workers (3) (6) (15) (17) (35) have reported chemical treatments that were effective in prolonging the life of cut flowers. Hitchcock and Zimmerman (18), however, found no marked beneficial results from the use of 50 different chemical treatments, many of which had been favorably reported on by other investigators.

Two proprietary materials, Floralife and Bloomlife, neither of which were included in the above test, are now in common use as flower preservatives and others have recently been placed on the market. It has been shown by a number of workers (21) (23) (^5) that these materials as well as other preparations (3 ) (^5) are effective in increasing the time during which roses will remain turgid and in good condition. It has been observed (23) that after a few dayB the petals of chemically treated red roses lose their original color Just as do those with their stems In water but that the flower may maintain a highly satisfactory and pleasing color as contrasted with the dull violet or bluish tinges of the latter. -10-

Similar observations are reported by Koon (23). It has been found

(3) (21) that during periods of refrigeration too little absorption takes place to justify the use of preservatives.

According to Iaurie (26) the value of flcver preservatives depends on their effectiveness in reducing the rate of respiration, decreasing bacterial action, Increasing absorption and transpiration, and providing the proper hydrogen-ioii concentration and osmotic pressure. To these might be added the role which they play in stabilising the flower pigmentB or otherwise perform in maintaining an attractive petal color for a longer period of time. Neff (35) has pointed out that most of the natural plant colors are mordant dyes, that is, they must be adsorbed by a mordant or mordants if the color is to be stabilized. It may be that certain flower pre­ servatives stabilize flower color by this means.

In work done at the Ohio State University two formulas were developed that proved particularly effective in prolonging the life of cut flowers. These formulas as given by Laurie and

KIplinger (29) are:

1. Hydrazine sulfate* 2 teaspoonfuls

Manganese sulfate 2 grams

Sugar 1 tableepoonful

Water 1 quart

2. Potassium aluminum sulfate (Alum) l/i+ teaspoonful

Sodium hypochlorite (Chlorox) l/b teaspoonful

* Use stock solution - 1 ounce to 1 quart of water. -11-

Ferris oxide l/lb teaepoonful

Sugar 2 teaspoonfuls

Water 1 quart

Laurie (27) stated that it ie very probable that Floralife and

Bloomlife are very similar in composition to Formula No. 2. This is also indicated by Neff (35) who, in a report on the effect of storage conditions on cut roses, stated that two cornnercial preser­ vatives, United States Patent Numbers 2230931 aud 2166304, Flora- life and Bloomlife respectively, are both composed principally of sugar and aluminum salts.

From the literature that has been cited, it appears to be definitely established that certain flower preservatives are effective in prolonging the life of cut flowerB and in maintaining a more desirable petal color. More work needs to be done to fully determine the effect that these preservatives have on a cut flowering stem that results in better keeping quality.

III. Flower Pi^nent Studies

A. Nature of Anthocyanin Pigments

The anthocyanin pigments, water soluble and present primarily in tho vacuolar Bap of the cell, are chiefly responsible for the red, violet, and blue coloring matter of flowers and other plant parts. These pigments are present in the plant as glycosides and yield on acid, alkaline, or enzymatic hydrolysis, a sugar, or sugars, together with an aglycon or sugar free residue that is known as tlie anthocyanidin. The anthccyanid.1ne are all derivatives of 2-phenyl- benzopyrylium salts (flavylium salts). The ring oxygen atom of*

this structure ia known as an oxonium group ajid is the ionic group

of the flavylium ion. Though commonly written in the form of its

chloride, the flavylium ion is undoubtly combined with other anioib

in the plant, probably to a large extent with those of plant acids

thus producing axonium salts (5>).

In solution the oxonium salts of anthocyanine are red in

color. For example, cyanin is red in solution of pH 3 or less,

violet at pH 8.5 and blue at pH 11.0. The red form of cyanin is

the oxonium salt, the violet form is represented by the color base

and the blue form by the salt of the color base (^).

ort MO -OH

Oxonium bait Color Base ha Balt of Color Base

Seven naturally occur!ng anthocyanidins have been reported.

As stated above, the anthocyanine are glycosides of the antho-

cyanidine. Since a great variety of sugars may be involved and

since both mono- and di-glycosides as well as mixed diglycosides may occur, it is evident that a great variety of anthocyanine may be derived from each anthocyanidin.

B. Factors Affecting Colors of Anthocyanin Pigments

An excellent review concerning the chemistry and biochemistry, occurrence, morphology, physiology, and genetics of anthocyanins -13- ha b been written by Blank (4). This paper covers chiefly the research which has been conducted since 1925, i.e., since the appearance of the second edition of an earlier work by Onslow (37).

In this review Blank (4) discussed several factors capable of affecting the color of anthocyanin pigments in plants. These in­ clude the following: simultaneous presence of several anthocyanins, variability of anthocyanin content, the hydrogen-ion concentration of the cell sap, the ash content of the cell sap, co-pigmentation, and colloidal condition of the cell sap.

Changing Amounts and Mixtures of Anthocyanin Pigments ♦ There seems to be no question concerning the effect of changing amounts and mixtures of anthocyanine on the color of flower petals. The effect of some of the other factors, however, haB not been so well clarified.

Hydrogen-ion Concentration of Cell Sap. It has long been known that anthocyanins are natural indicators, changing color as the hydrogen-ion concentration is increased or decreased. Buxton and Darbishire (7) buffared pigment extracts from a number of flcwere at hydrogen-ion concentration varying from pH 3 to pH 11.

They found that a pigment extracted from a rose "more red than blue" was verraillion at pH 3, pink at pH 4.5 and 6, purple at pH 7*

8, and 9 an

Hcwever, investigations by Robinson (U6) and Scott-Moncrieff (50)

have shown that the differences in the pH between red and blue

petals of different varieties of the same plant are often too

smell to explain the different colors. Blank (4) pointed out that

despite the properties of anthocyanins as indicators, a much more

important role than that accorded to the cell sap reaction is played

by other factors in the color changes of anthocyanins in plant

tissues. According to Small (52) the different colors in different

flowers are due to different anthocyanins but different colors in

the same cells of petals at different times are due to different

pH values affecting the same anthocyanin.

As pointed out by Scott Moncrieff (50) it still remains to be

ascertained haw often color variations are directly affected by pH

differences and how often the differences found are merely the

result of changes in pigmentation or in concentration, which them­

selves result in a change in hydrogen-ion concentration.

Influence of Ash Content. Hie influence of the ash content

of the cell sap on the color of plant tissue has not been completely

settled. However, it has generally been found that blue petals have higher ash contents than red petals of closely related plants.

Karrer and co-workers (20) determined the ash content of various

red and blue petals and found that blue ones contained more ash than

red ones. This was regarded as evidence that the color of red flowers is determined by oxonium salts, whereas that of blue flowers

is determined by alkaline salts or by those of the alkaline earths -15- of the anthocyanin*. Mlhalleacu (32), however, In a alnilar investigation noticed no significant differences In the ash content of red and blue flower petals of closely related plants. In a later investigation (33) the same author determined the alkalinity of the ash contents and found a larger amount of alkaline substances

In the ash of blue flcwer petals than In those of red. Investiga­ tions by Allen (2) and Chenery (8 ) have indicated the great signi­ ficance of aluminum In blue flowers of hydrangea.

Co-pigmentation. Die phenomenon of co-pigmentation was

Investigated In detail for the first time by Robinson and Robinson

(47)* They found that certain substances, mainly anthozanthlns, the flavones and flavonols, and tannins had a bluing effeot on the color of anthocyanin solutions irrespective of the pH. It was their belief that this was the result of the formation of weak additive complexes rather than from salt formation. Uaey stated that these complexes occurred in the flowers themselves and played an Important part In producing variations of color. In Fuchsia flowers they observed that the violet Inner corolla gave a bluer red acid extract than that obtained from the outer bluish red petals. It was found that this was due to the tannin derived gallic acid contained in the first extract.

Scott-Moncrieff (5 0 ) reported that flavones modify flower color not only by their co-pi&nent effects but also by their tendency to vary the concentration of anthocyanins by their inter­ action effect. Thus the presence of ivory flavone is often -1 6 - accompanled by a bluing effect and alao by a decrease In antho- oyanln.

Iawrence (3 0 ) found that ivory flavone in the flovers of

Dahlia varlabills had the capacity to act as a co-pigment with a mixture of cyan in and pelargonln to change the color from a chocolate to a bluish purple.

Condition of Cell Sap. The colloidal condition of the cell sap has been shown by the Robinsons (47) to be especially important along with co-pigmentation as a factor affecting flower color. They observed that the cell sap of the cornflower colored blue litmus red and actually had a more acid reaction than the cell sap of a red rose, both of which contained cyanln. Further investi­ gations shewed that a lyophilic colloid was present in the cell sap. They concluded, therefore, that the cyanln must be present

In the cornflower in a complex form and not in the form of the potassium salt as assumed by Willstatter and Everest (59). On the basis of these observations it was suggested that cyanln in the blue cornflower cell sap was blue because it was absorbed on colloidal particles which stabilized the anions at a pH of about 4*9*

C. Rose Pigment Studies

Wlllstatter and Noland (60) Isolated a pigment from the red rose, Rosa sallca. in 1914 which they found to be identical to the pigment in the blue cornflower, Centaurea cyanus ♦ This pigment was

Cg.jH^O^gCl, a 3/5 diglucoside of cyanldln. Currey (11) in an examination of the red rose "George Dickson" found that the antho- I

ACKN OWLEDGEMENT

Bie writer wishes to express his sincere appreciation for the assistance which was so freely given him by students and facility of the Horticulture Department of The Ohio State University and by the staff of the Horticulture Department of The Ohio Agricul­ tural Experiment Station during the course of this study.

Special thanks are extended to Professor Alex Imirie for hia suggestions and inspiration and to Roses Incorporated, the spon- sorer of the Fellowship which made this work possible. -17-

cyanin pigment contained in the flowers of thle variety vaa alao

cyanln and that it ezieted aa an azonlua salt* Both Cana oner (9 ) and Thiaann and Edmondson (5 6 ) have found oryetalline cyanln to have a characteristic adsorption peak at 510 mu.

Curry (12) In a later paper reported on the cause of bluing

In red roses.He worked with the varieties Hadley, a bluing type, and lady Maureen Stewart, one which seldom blued. A summary of his work shews:

1 . The pigment, cyanln, was present in both varieties.

2 . The quantities of individual ash constituents were prac­

tically the same.

3* Flowers from the variety Hadley contained much leas tannin

than those from lady Maureen Stewart.

4. There was a direct relationship between the quantity of

anthocyanin pigments and tannin present In the two

varieties. (Blue Petals of the variety Hadley contained

6*33 percent tannin and 1.26 percent anthocyanin calculated

on a dry weight basis, lady Maureen Stewart contained

11.62 percent tannin and 2*39 percent anthocyanin.)

As In the variety George Dickson It was found that the cyanln was present as an oxonium salt. On the basis of these results Curry postulated that the tannin acted to stabilise the red color and that the bluing coesnon to the one variety was caused by a lack of acidity

In the sap due to a decreased content of tannin. His work Indicated that tannin was the only acid preeent In the cell sap of the petals. -1 8 -

While blue petals from the variety Hadley was round to contain.6.33 percent tannin on a dry weight baale, normal colored petals con­ tained 7 * 3 8 percent.

Robinson (ltd) determined the pH and the anthocyanin content of several rose varieties. He noted that the "Vlelohen" rambler rose had red buds (pH 6 .3 ) changing to dull violet flowers (pH 6 .3 ).

Bis anthocyanin ratio of red petals to violet was round to be

29*6 : 29*9* He also observed that In dry seasons the riowers of this variety became violet almost as soon as they opened but In a wet one they changed only Just before fading. It was found that the anthocyanin content was higher under the more moist conditions.

Hie flowers of "Madame Bdward Harriot", a salmon pink rose, was observed to change to a bluish pink with age. In this variety

Robinson also found that the pH was the same In both cases, 5*9*

The anthocyanin ratio of young to older flower petals was 4k : 20.

Flavones were present but shewed little change.

A change In color was noted by Robinson only In the rose varieties In which the flowers were characterised by a relatively high pH, about 6.0. Flowers of the rambler "Kxcelslor", containing the same pigment as "Vielchen", I.e. cyanln, had a pH of ^.6 and did not show any color change with age.

Hiornton (5 8 ) has shewn that the bluing of red roses from excess carbon dioxide Is accompanied by a reduction in the hydrogen- ion concentration of the cell sap of the petals.

Ratsek (43) (44) reported that the development of deep color 19

In outdoor roses v u dependent upon available carbohydrate supplies.

This indicated that tha palar color ocmmonly 00curing in a u m r was dua to insufficient carbohydrate s rathar than to a hi earthing of color pigments by sunlight.

Because, as was pointed out in tha literature review, there are a number of factors that can affect the color of anthocyanin pignenta, it is not likely that the change in petal color that usually occurs in cut Better Times roses and other red roae varie­ ties la always produced by the same factor or combination of factors. Likewise, varietal characteristics would also seem to be important. Even though the bluing that occurs in the variety

Hadley may noimlly be caused by a deficiency of tannins, as concluded by Curry (12), the bluing in Better Times roses may normally be caused by s o s m other factor.

It therefore seems important that further studies of a chemical nature be made In order to gain a better understanding of the physiological changes which occur and that may be responsible for the observed color changes. In this way it may be possible to develop methods or practices which will be effective, at least to a certain degree, in preventing the loss of quality in cut, red roses that Is associated with bluing. - 20 -

STUDY 1 1 W E EFFECT 0V SOIL POTASSIUM AND NITRATE CONCENTRATIONS

ON ZEE KEEPING QUALITY OP TEX HETZBt TIMES ROSE

HATKAIfl AMD METHODS

A H roses used In the study on the effeot of various levels and combinations of level* of potassium and nltratss In the sub­ strata on tha keeping qualities of tha out flowers vara grown In tha Floriculture Greenhouses of tha Department of Horticulture and

Forestry of Tha Ohio State University.

Started eye plants of tha variety Better Timas vara obtained through tha courtesy of Mr. X. Sohvertscharft, Valley Rosa Nurseries,

San Lorenzo, California. These plants vara benched on February 1^

1950 in a shredded bluegraes sod soil. Except for tha fertiliser treatments the plants vara grown according to standard ccnerolal practices.

Nina plots, each containing 2k plants and replicated one time, were set up in which it was proposed to maintain the nitrate and potassium levels, as shown by periodical soil tests, within certain definite ranges. The desired concentration of nitrate, potassium, phosphorus, and calcium for each plot is shown in Table 1. However,

it was found that these definite ranges of nitrate and potassium content were hard to maintain. For this reason the nitrate and potassium status of the plots will be given as lew, medium, or high.

The parts per million of nitrate and potassium In the substrate, as determined by the soil tests, that are associated with these

1 21

TABU I. Proposed concentration of mineral aalte In aubetrete

for roaa keeping quality plota.

______Concentration yf qineral Salta______Plot ______Nitrate Jhoanhorua Potaaalum Oaloiua

p.p.m. p.p.m. p.p.m. p.p.m

1 5-10 5 5-10 150

2 25 5 5-10 150

3 50-75 5 5-10 150

4 5-10 5 20 150

5 25 5 20 150

6 50-75 5 20 150

7 5-10 5 40-60 150

8 25 5 40-60 150

9 50-75 5 40-60 150 terms In this pspsr srs as folloirt: HO3 X

Hifih 50+ 40+

Medium 25-50 15-40

Low 0-25 O-15

Soil tssts for potaaalum and nltrats wars aada on representa­ tive aaoplaa from aaoh plot at bi-monthly Intervals by tha Spurvay method (5 3 ) of soil analysis. During tha first part of tha com­ parative keeping quality study, October 1950 to May 195l, additions of mineral salts to maintain tha desired levels In tha soil vara made on tha basis of tha soil test results. Starting in July,

1951* applications of mineral salts to tha "high” level plots vara made at veekly intervals.

Hie desired levels of nitrate and potassium vara easier to maintain toy tha latter procedure. However, in all Instances the concentration of nitrate and potassium as shewn toy soil analysis, rather than the desired level, Is taken as the true mineral salt status of a plot for any given period of time.

Hoses for the comparative keeping quality tests vere out during the morning and at a stage of maturity comparable to that at which roses are cosmonly cut In commercial ranges. After cut­ ting they vere placed in water in a refrigerator at 4o# F. for

24 hours. After this time the stems were cut to 12 to 15 Inches and tagged as to date of cut and plot of origin. The flowers vere then placed in containers of vater and kept at room temperature -23 until discarded. As nearly as possible only flowers of apparently

•qusl quality vsrs obossn to bs used in ths tssts.

Observations vers made daily from the time the roses vers placed at room temperature until discarded. Records vere kept as to longevity and the condition of flovers during the observation period. The flovers vere discarded vhen petals began to wilt, the stems became "soft", thus allovlng the flower head to droop, or vhenever they otherwise reached a stage at vhloh it vae assumed they would no longer be desirable to the average consumer.

Bach flover was rated on the basis of the number of days it remained in this so-oalled "desirable" condition from the time it vas placed at room temperature. The average keeping quality of roees cut from any one plot vas then determined by multiplying the number of roses falling in each keeping quality group by the number of days they vere kept at room temperature before being discarded, totaling the results, and dividing by the number of flowers tested from that particular plot.

Seven comparative keeping quality tests vere made over a period of lk months. The period of time covered by a test varied from one to four veeks. These periods were chosen in such a manner that the relative keeping quality of the roses from the various plots could be determined in relation to the season of the year.

An analysis for total nitrogen, potassium, and calcium vas made on representative leaf samples from each plot at a time correspon­ ding to three of the comparative tests. Ihese analyses served as a check on the mineral ealt statue of the plants at the time these particular comparative tests vere conducted.

Leaf samples o one is ted of leaflets taken from the lover tvo leaves of a out f love ring shoot. As the flovering shoots vere comonly out Just above the second 5-leaflet leaf, these samples represented, for the most part, the third and fourth 5 -leaflet leaves of a flovering shoot. Only the leaflet blade vas included in the sample.

After washing In distilled water to remove dust and other foreign material the leaf samples vere dried In a forced draft oven for 2k to 30 hours at a temperature of 95* C. Die dried material vas then ground in a Wiley Mill through a lt-0-mesh screen.

Nitrogen vas determined on one gram allquota of the dried material by the KJeldahl method (l). Potassium and calcium were determined by the use of the Beckman Flame Photometer. RJBPIJB i 9 m rtiulti of the comparative keeping quality studies a n shewn la Tables 2 to 8 . Oata la thaaa tables show tha average keeping quality la daya of roaaa out from tha dlffarant plots at various tlmse of tha yaar. The percentage of flowers out from aaoh plot that vara dlsoardad la lass than four days aftsr being plaosd at roosi taaparaturs haoausa of premature wilting of tha pa tala, softsnlag of tha steam, bluing, or for othar raasoas la also ah ova. Tha paroantaga flguras 11s tad uadar "Othar" rafar to flowers dlsoardad prematurely for raasoas othar than tha thraa llstad above* For tha most part this would Inoluda shattarlng of tha petals, extensive fading of tha petals, or rapid opening of tha bud.

In aost lnstanoes flovers exhibiting softening of tha stem or stem breakdown ware also characterised by wilted petals or buds that failed to open. However, these flowers ware included only under tha coluan head "Stem Breakdown".

Die results presented in these tables shew, that except for high potassium, tha mineral nutrition of tha rose plant has vary little effect on tha kaaplng quality of Better Times roses. Under conditions of low, medium, and high nitrate and low or medium potassium, tha season of tha yaar had more effect on the kaaplng quality of tha roses than did the concentration of potassium or nitrate in tha substrate. -26-

Bie poorest keeping quality was obtained from roses cut during the winter and early spring months. During this time the average keeping quality was approximately 3.3 days compared to an average of 4.3 to 4.5 days during other times of the year.

Neither was premature wilting of petals or steins correlated with any of the different nutrient treatments. However, bluing of the petals occurred more extensively and quicker on roses cut from plants growing In the high potassium plots.

The bluing of Better Times Hoses was observed to begin first on the margin of the innermost petals. It may then further develop, progressing outwardly, until all of the mid-petals and even the outer petals are bluish in appearance.

Tables 4, 7 , and 8 show, that In general, there was a corre­ lation between the soil test results and leaf analysis.

The roses characterized by the excessive and rapid bluing were cut from plants having a leaf potassium content equal to 3*4 to 4*0 percent of the dry weight. This greater tendency towards bluing was not apparent on roses cut from plants having a leaf potassium content up to 2 . 6 percent of the dry weight. ii

Table of Contents

Page

Introduction...... • . . . . 1

Review of Literature...... 3

Study I: Hie effect of Soil Potassium and Nitrate Concen­ trations on the Keeping Quality of the Better Times Rose

Materials and Methods...... 20

Results...... 25

Study II: Some Chemical Changes Associated with Bluing In Petals of Better Times Roses

Materials and Methods...... 34

Results...... 43

Study III: Some Chemical Changes In Petals of Better Times Roses Resulting from the Use of Flower Preserv­ atives,

Materials and Methods...... 57

Results...... 6l

Discussion...... 70

Summary and Conclusions...... 80

Appendix...... 83

Bibliography...... 92 27-

T A B U 2. Bffect of Soil Nitrate and Potassium lavs 1b on the Keeping Quality of Better Times Roses. (November 10 to December 14, 1950).

Avg. Percent of Flowers Shewing Inferior Quality* Plot No. FIs. Keeping Wilted Stem Tested Quality Petals Breakdown Blulxux Other Total (days) loir NO3 , low K II-l 25 3.8 20.0 4.0 4.0 8.0 36.0 II-2 33 4.2 3.0 ---- 6.0 6.0 15.2 II-4 30 It .2 3-3 3.3 --- 6.7 13.3 II-5 34 4.1 2.9 ---- 2.9 8.8 14.7 III-l 31 3.8 12.9 --- 9.7 6.5 29.0 III-2 31 4.3 3.2 --- 6.5 9-7 med. NO3 , low K II-3 25 3-9 6.0 4.0 20.0 32.0 III-3 38 4.2 10.5 2 .6 5.3 2.6 21.1 lew N0o, med. K n - a 23 4.3 4.4 4.4 8.7 XI-9 27 4.1 7.4 7.4 ------14.8 III-4 21* 4.1 12.5 1*.2 ---- —- 1 6 . 7 III-5 25 4.4 ------III-6 25 4.2 12.0 ---- 4.0 16.0 III-7 22 4.1* —***■ ---- i*.6 9.1 13-6 med. N0^, med. K II-6 21* 4.2 4.2 1*.2 8.3 II-7 34 4.2 ------2.9 5.7 8.8 III-8 29 1*.2 6-9 3.5 ---- 3*5 13-8 high NOo, med. K III-9 21* 4.0 25.0 _ _ _ — — — 2 5 .O

* Discarded in less than four days after being placed at room temperature• - 28-

TABUS 3* Nffeot of Soli Nitrate and Potaselun Levels on the Keeping (Quality of Better Tines Roses. (February 20 to March 18, 1951).

Plot No.FIs. Keeping Wilted Stan Tested Quality Petals Breakdown Bluing Other Total (days) loir NOo, low K II-l 11 3.3 36.4 9.1 9.1 54.5 11-2 6 2-5 62.5 2 5 .O ------87.5 H - 3 16 3.1 1*3.8 18.8 --- 6.3 68.8 mad. N0^, low K III-2 8 3.3 --- 62.5 62.5

low ned. K 11*5 17 3.* 23.5 17.7 47.1 IX-7 14 3.5 21.4 14.3 --- 7.1 42.7 II-6 11 3-5 27.3 18.2 --- 9.1 54.5 II-9 13 3.2 38.5 15.4 --- 15.8 69.3 III-l 17 3.* 11.8 5.9 --- 29.4 47.1 III-4 18 3.* 22.2 ------33.3 55.6 III-8 12 3-5 16.7 8.4 --- 16.7 41.7 ned. NO3 , ned. K II-4 17 3-5 17.7 11.8 — — — 23.5 52.9 II-6 17 3.1 1*1.2 11.8 17.7 70.6 III-3 11 3.4 9.1 9.1 27.3 45.5 III-6 17 3.1 23.5 17.7 --- 29.4 7 0 .6* III-9 12 2 . 8 16.7 41.7 16.7 75.0

low NOo, high K III-7 11 3*4 27.3 9*1 --- 9.1 45.5

* Discarded In less than four days after being placed at roan temperature. TABLB 4. Effect of Soil Nitrate and Potassium Levels on the Keeping Quality of Better Times Roses. (April 16 to May 3, 1951)*

Avg. Percent of Flowers Shewing Inferior duality* Plot Leaf Analysis No.Fls. Keeping Wilted Stem (ft of dry weight) Tested

low NCh, med. K II-4 3.3 1.8 16 3.6 6.3 18.8 6.3 37.5 68.8 II-7 3.8 1.8 14 3.0 21.4 35.7 21.4 7.1 85.7 II-8 3.6 2.1 9 2.6 33.3 55.6 ---- 11.1 100.0 II-9 4.0 1-9 16 3 A 18.8 18.8 6.3 18.8 62.5 III-4 3.6 1.8 15 3.2 13.3 46.7 6.7 6.7 73.3 III-7 3-5 1.9 10 3.3 10.0 30.0 — 20.0 60.0 III-8 3-9 2.2 15 3.1 26.7 46.7 — 6.7 80.0 med., NOoj med' . K 11-5 3-9 1.8 20 3.3 45.0 15.0 — — 25.0 85.0 III-6 4.0 1.7 13 3.5 30.8 30.8 7.7 — 69.2 III-9 3-9 2.1 9 3.6 33.3 22.2 —— 55.6

* Discarded in less than four days after being placed at room temperature. -30-

TAB I£ 5* Effect of Soil Mltrate and Potassium Levels on 'the Keeping Quality of Better Times Roses. (August 2 to August 6, 1951)•

Percent of Flowers Showing Inferior Quality* Plot Mo.FIs. Keeping Wilted Stem Tested Quality Petals Breakdown Bluing Other Total (days) low MO^, low K II-l 5 4.6 — ... -- _ __ -- II-2 10 4.6 ------II-3 10 *•7 --- ——— ------III-l 8 4.8 ------III-2 12 4.8 ------III-3 7 4.3 ------——-

low med. K II-4 6 ^.7 ------16.7 16.7 II-5 8 4.1 ------12.5 12.5 25.O II-7 15 4.4 ------6.7 6.7 13.3 II-8 11 4.6 ------9.1 9.1 II-9 12 4.7 ------III-4 10 4.7 — ------10.0 10.0 III-5 6 *•5 ------III-6 7 4.4 ------III-7 7 4.6

mod.. NO^, med. K II-6 12 4.5 ------

* Discarded in less than four days after being placed at room temperature. -31- tabus 6 . Effect of Soil Nitrate and Potassium levels on the Keeping Quality of Better Times Roses. (September 3 to September 8 , 1951).

Plot No.FIs. Keeping Wilted Stem Tested Quality Petals Breakdown Bluinc Other Total (days) low NO^, low K II-l 15 4.0 20.0 20.0 med. NO3 , low K II-2 22 3-9 9.1 18.2 27.3 III-2 30 4.1 -- — 20.0 20.0 high NO^, low K H - 3 33 3*9 3.0 12.1 15.2 30.3 low med. K II-4 17 4.2 11.8 11.8 III-3 37 4.0 2.7 2.7 2.7 10.6 18.9 III-4 22 4.1 ------18.2 18.2 med. NO^, med. K II-5 19 4.1 5.3 ... 15.8 21.0 III-5 33 4.1 ------6.1 12.1 18.2 high NO^, med. K II-6 14 3.8 7-1 7.1 35-7 50.0 III-6 40 4.0 2.5 10.0 7-5 20.0 low NO^, high K II-7 11 3-5 36.4 18.2 54.4 III-7 29 3-7 10.3 27.6 7.0 44.8 med. NO3 , high K II-a 14 3*4 42.9 1^-3 57.1 III-8 29 3*4 ------51.7 10.3 62.1 high NO^, high K H - 9 26 3-4 57.7 3.9 61.5

* Discarded In less than four days after being placed at room temperature. TABI£ 7* Effect of So:!l Nitrate and Potassium Levels on the Keeping Quality of Better Times Hoses. (October 8, to October 17, 1951)*

Plot Leaf Analysis ho .FIs. Keeping Wilted Stem (£ of diy weight) Tested Quality Petals Breakdown Bluing Other Total NK (days) low lfOn, low K II-l 2.9 1.5 23 4.5 — ... -- ^•3 III-l “ “ “ — mm 17 4.5 • — _ 5.9 5.9 med. NO?, lew K II-2 3.0 1.6 27 4.4 3 — — 3.6 — 3.6 III-2 — — 19 4.4 ... — 5.3 5.3 high N0V low K n - 3 3-3 1.7 33 4.4 J — —— 3.0 3.0 — — —— i IJI-l 36 4.5 2.8 2.8 u> to low NCb. med. K i —— ii 2.9 1.9 22 4.4 3 — ^ 5 *•5 III-U — mm mmm 15 4.6 — ——— m — — ——— * — — — med. NO3, med. K i i -5 3.3 1.9 25 4.2 — 20.0 — 20.0 III-5 —— 31 4.4 — 6.5 — O.jft * high med. K II-6 — 1.8 33 4.2 J — — 15.2 9.1 24.2 ... — — R ?-6 — 36 4.4 8.3 8.3 low MOn, high K II-7 3.6 3-4 13 3.4 — 61.5 7-7 69.2 III-7 mmm “ ™ — 25 3.8 36.C » —— 36.0 med. NOo, high K II-8 3-7 3.8 22 3-7 — 54.5 -- 5^.5 III-8 ... — 30 3.7 — 36.4 3.0 39.1 high NOq , high K II-9 3.8 3-4 22 3.3 — 66.2 18.2 86.4 — — — III-9 18 ,, 94.4 — 2 W * Discarded in lees than four days after being placed at room temperature. TABI£ 8. Effect of Soil Nitrate and Potassium Levels on the Keeping Quality of Better Times Roses. (November 28 to December 1951).

Plot Leaf Analysis No.Fla. Keeping Wilted Stem (% of dry weight) Tested duality Petals Breakdown Bluing Other Total N K (days) low MO3, lew K

.... II-l 3.0 1.7 25 4.9 — — III-l 3-5 1.7 10 4.4 4.8 • “ m m " • • m ™ 4.8 med. N0^, lew K II-2 It.2 1.8 29 4.7 — « V M 3.5 3.5 III-2 3.6 2.0 12 4.8 • • • * * * * “ “ * ••• high N0o, low K II-3 3.7 1-5 30 4.8 --- -- — .... 1 III-3 3.8_ 1.6 24 4.6 4.2 __ 8.3__ 1 u> to lew N0a, med. K l II-4 4.0 2.5 15 U .5 — — — ------III-4 3.9 2.6 22 U.5 — 4.5 — 4.5 med. NO?, med. K II “5 3.6 2.2 18 4.5 — 5.6 — 5.6 III-5 3.8 2.2 32 4.4 — 9.4 3.1 12.5 high N0o, med. K II-6 3.8 2.3 31 4.5 12.9 — 12.9 III-6 _ 3*9_ _ . 35 4.5 — 8.6 — 8.6 lew NO3, hlgi K II-7 4.3 3.8 26 3.3 — 69.2 — 69.2 III-7 4.1 3*^ 22 3.8 31.8 13.6 *5-5 med. NO.,, high K II-8 4.3 4.0 19 3.5 3 — — 63.2 — 63.2 III-8 4.4 4.0 21 3.5 — 57.1 — 57.1 high N0q, high K II-9 4.6 3.4 3.3 * — ... 78.3 — 78.3 III-9 4.6 lr.8., 17 _ 3,5 5.9 ... 38.1 -5 *9 _ 58.8 * Discarded In less than four days after being placed at roan temperature. -34-

STUDY II: SOME CHEMICAL CHANGES ASSOCIATED WITH BLUING IN PETALS

OF BETTER TIMES ROSES.

MATERIALS AND METHODS

A. GENERAL PROCEDURE

In the study previously reported it vas discovered that more rapid bluing was characteristic of Better Times roses grown with high concentrations of potassium. Another observation was that bluing begins first on the margins of the inner petals and then further develops to include greater petal area and more petals.

In this study a series of chemical and physical tests were conducted in an effort to determine some of the factors associated with bluing of Better Times Roses and the possible cause of the more rapid bluing that resulted when the roses were grown at high potassium levels.

Roses for these tests were cut from the same plots as those used in Study I and handled in a way similar to that described for the comparative keeping quality tests. However, in order to have easily removable petals for sampling, the blooms were not cut until at least four petals had begun to unfurl. The hardening period was reduced from twenty-four to four hours.

Quantitative measurements of anthocyanins and phenolic com­ pounds were made on inner and mid petals of bluing and non-bluing roses. Also two quantitative determinations of anthocyanln and phenolic compounds were made on each of ten representative roses -35-

from each of the fertilizer plots. This made a total of twenty

flowers for each differential treatment. The first test for antho-

cyanins was made on duplicate samples from the four outer petals of

freshly cut, hardened, roses. The second determination was made

four days later on duplicate samples from the 9th, 10th, 11th, and

12tb petals. Bie first tannin test was made on duplicate samples

taken from the 5 th, 6th, 7th, and 8th petals approximately 2k hours

from the 13th, lhth, 15th, and l6th petals four days after cutting.

The pE of the exx>ressed cell sap was determined for five blue,

five non-bluing, and five freshly cut roses from the high and the

lew potassium plots. In addition, pH measurements were made on the

expressed Bap of 10, five day old, representative flowers from

each of the differential nutrient treatments.

An analysis for potassium and calcium was made on freshly cut

flowers from each plot and on bluing and non-bluing five day old

flowers from the high and the low or medium potassium plots. In

the latter case samples consisted either of complete flowers or

only of inner and outer petals depending on the nature of the

sample. The inner and outer x»tals represented the bluing and

non-bluing portions of the flower.

The absorption spectra of the anthocyanin and anthoxanthin pigments were determined for bluing, non-bluing, and freshly cut

roses.

The sampling for this study was done during the month of March, 36

1952. Leaf s u p lea oollected at the beginning and end of this period were analyzed for total potassium, calcium, and nitrogen to

Indicate the mineral salt status of the plants during this time.

B. MEASUREMENT OF IBZNOXIC (TANNIN) COMPOUNDS.

In an investigation by Folln and Dennis (l4) on phosphotungstlc phosphomolybdlc compounds as color reagents, one of these compounds

(Folin-Dennls reagent) was found to give a positive reaction with all axyphenyl compounds and particularly with tannic and pyrogalllc acid. They reported that aliphatic compounds did not Interfere with the color reaction.

Rosenblatt and Peluso (49) developed a colorimetric method for measuring tannins In tea leaves that is based on measuring the blue color produced by the reaction of the Folin-Dennls reagent with the axyphenyl compounds. They stated that the presence of more than one oxyphenyl bond In tannins made the application of this reaction distinctly suitable for these compounds• It was found that Beer's law is valid for tannic acid concentration of 0.05 to O .25 milli­ grams per 100 milliliter of solution and that any filter from 640 to 850 mm. was satisfactory for the determination.

Ihe method used In this study for the determination of phenolic compounds, assumed to be mostly tannins, was essentially that des­ cribed by the above authors. A modification was made by using a

10 percent solution of sodium carbonate In place of a saturated solution as one of the reagents. This modification was also made by

Qrice (1 6 ) in using the method for measuring tannins in fresh and -1-

IM TR ODUCTI OI>i

Hoses have been and still remain the moat important of all cut flower crops. Records of Roses Incorporated show that approxi­ mately 15,0 0 0,000 rose plants were grown for cut flower production in the United States during 1951* It is estimated that about

^00,000,000 roses having a wholesale value of 1*0,000,000 dollars and a retail value of 100,000,000 dollars were produced by these plants. A recent market survey by Swingen (55) showed that the rose was the preferred flcwer of shoppers at all times of the year.

However, regardless of the popularity and economical importance of the rose, it is one of the poorest of cut flowers from the stand­ point of keeping quality. Under average home conditions carnations or chrysanthemums may be expected to retain a presentable condition for a period up to seven dayB from the time they are sold by the retailer. Roses, under the same conditions, are usually'' ready to discard after the second or third day.

Because of the unfavorable keeping quality of roses in com­ parison to some of the other cut flower cro£ns there ie a possibility that the rose may not be able to retain its present favored position.

For this reason it is of great importance to the rose industry that studies be made concerning this rapid loss of usefulness and means of prolonging the life of the rose bloom.

There are a number of factors to consider .in a study cuncemin; tne keeping quality of roses. Among these are "bluing" and fading

>1' the petals of red rose varieties, wilting or bending of the stem 37-

froten peaches.

The percentage of light transmission vaa measured toy — ana of a La it*, Rouy-Fhotrameter, using a 640 ms. filter. Die instrument vaa adjusted to read 100 percent transmission with the blank. The

transmission of the sample waa then calculated In terms of optical

density (Log l/T).

As the degree of color development follows Beer's lav within

the range of concentrations reported In this paper and alnce only

the relative quantity of tannins or other phenolic compounds present

in the petals waa of Interest In this study the results are reported

in terms of optical density. Concentrations of pure tannic acid of

0.05 and 0.25 milligrams per 100 milliliters of solution, the range

of concentration in which Beer’s lav is applicable, gave density

readings of .091 and .485 respectively.

Samples for anthocyanln and tannin determinations consisted of

four discs cut from four different petals of the same flower with a

cork borer having an Inside diameter of 14 millimeters. To obtain

duplicate samples two discs were cut from each petal. These discs were removed, one from each longitudinal half of the petal, as near

the tip as it was possible to cut and still obtain a complete disc without extending beyond the mid-line of the petal. Because of the

else of the petals it was possible to obtain only one disc from the

innermost petal. For analyses involving these petals, samples con­

sisted of single discs removed from the tip center of four petals. -38-

The following are the standard procedures adopted for the ex­ traction and measurement of the phenolic compounds:

1. Petal discs finely Macerated In mortar with snail quantity

of 70 percent ethyl alcohol.

2. 'Hie resulting slurry transferred quantitatively by the

use of the alcohol solution to a 50 milliliter volumetric

flask.

3* Enough of the JO percent alcohol added to bring the solu­

tion to volume at 50 milliliters.

4. After standing for approximately one hour the contents of

the flask thoroughly mixed and a portion of the solution

filtered through Vftxatman No. 1 filter paper.

5 . Two milliliters of the filtered axtraot added to a 50

milliliter flask containing approximately 25 milliliters

of distilled water.

6. One milliliter of the Folln-Oennls reagent added to flask.

7 . Ten milliliters of 10 percent sodium carbonate added to

flask.

8 . Distilled water added to bring the volume to 50 milliliters.

9 . After standing for two hours the solution was thoroughly

mixed the percentage of light transmission measured by

the Rouy Photroneter. A blank prepared in the same way

used for the reference solution.

10. Transmission readings expressed In terms of optical

density. -39

C. MEASURJMXNT OV AflTHOCIABIM CQMCBmiATION

A lurvijr of the literature Indicated that a dilute eolution of hydrochloric acid waa one of the solvents noet commonly used for the extraction of water soluble plant pigments. A .1 N solution of hydrochloric acid, prepared by bringing 8*2 milliliters of 36 per* cent hydrochloric acid to a Yolume of 1 liter with distilled water, was used in this study,

Die relative amount of pigment extracted was measured by ob­ taining the percentage of light transmission with a Lsits, Rouy

Ihotrcawter using a 220 mu filter* The instrument was adjusted to read 100 percent transmission with the .IN hydrochloric acid sol* vent*

Preliminary work showed that with concentrations giving transmission readings of from 20 to 70 the adsorption definitely follows Beer's law. Similar results are reported by TtMmann and

Bdmondson (2 6 ). The 220 mu filter waa chosen on the basis of the absorption spectrum of an acid extract of Better Times rose petals*

The results of these measurements are reported in terms of optical density*

Bke following standard procedure was adopted for the extrac­ tion and measurement of the anthocyanln pigment*

1. Fetal discs finely macerated in mortor with small quantity

of .1 I hydrochloric acid*

2. The resulting slurry transferred quantitatively to a 2° -40-

mllllliter flask by ths uss of the hydrochloric acid

solution.

3* Ths flask brought to volume at 50 milliliters and allowed

to stand for approximately four hours.

1*. After four hours the solution transferred to a 5° milli-

llter pointed, centrifuge tube.

5* Solution centrifuged for 5 minutes at 2,000 r.p.m. In a

Size 1, Type SB, International Centrifuge.

6. Bie supernatant liquid transferred to an absorption cell

and the amount of light transmission measured by the Rouy

fhotrometer using the 520 mu filter. Bie .IN hydrochloric

acid solvent used as reference.

7* Transmission reading expressed In terms of optical density.

D. MSASURB4ENT OF pH

Bie measurement of the hydrogen ion concentration of the ex­ pressed sap was made with a Beckman, Model H, pH meter. 3he petals were first crushed in a mortor and the cell sap then expressed by the use of a small hydraulic pressing unit called a Succulometer (24).

E. MINERAL ANALISIS

Petals and leaves used for mineral analyses were dried in a forced draft oven at 95* C. for 24 to 30 hours and ground In a

Willey mill through a 40-mesh screen. Ihe leaf samples consisted of the leaflets from the lower two leaves of a cut flowering shoot as has been described in Study I. Before drying, the leaf samples were washed In distilled water. -41-

Ieaf nitrogen content was determined on one gram samples of the dried material by the KJeldahl method (l). Potassium and cal­ cium content of petals and leaves were determined on duplicate l/2 gram samples by the use of the Beckman flame photometer.

F. MEASUREMENT OF THE ABSORPTION SPECTRA OF ANTHOCYANIN

AND ANTHOXANTHIN PIGMENTS.

The extracts from ten 14 millimeter discs cut from blue, non­ bluing, and fresh petals were used for the measurement of the ab­

sorption spectra of the anthocyanin and anthoxanthin pigments

contained in each of the above types of petals. The discs were

crushed in a mortor with a small quantity of .1 K hydrochloric acid,

transferred quantitatively to a ^>0 ml. flask and brought to volume with more of the acid solution. After filtering, the extract was

shaken repeatedly in a separatory funnel with an equal volume of

ethyl acetate to remove the anthoxanthins. The two fractions were

then separated. The ethyl acetate fraction was saved and the acid extract was again shaken with on equal volume of the ethyl acetate.

After separation this second ethyl acetate fraction was discarded.

The absorption spectra of the anthocyanin extract and of the first ethyl acetate fraction were then measured in a Beckman Model DU

photoelectric spectrophotometer at wave lengths from 300 to 1 0 0 0 mu.

The respective reference solutions were prepared by thoroughly mix­

ing equal volumes of the acid and ethyl acetate solvents and sepa­

rating the two fractions.

To obtain an estimation of the relative concentration of antho- -42- xanthin pigments to a given concentration of anthocyanin pigments

obtained from each of the above types of petals, extracts were

prepared as described above except that larger quantities of petal

tissue were used and that before the anthoxanthlns were extracted

from the acid extract with the ethyl acetate each of the acid

solutions were diluted with .1 14 hydrochloric acid to give the

same transmission reading In the Rouy Riotrometer, uBlng the 520 mu

filter. -U3-

RBSUUS

As has been reported In Study I the Inner petals of the rose are the first to blue. Die data of Table 9 shew that though

there Is a quantitative difference In the anthocyanin content of

Inner and outer petals regardless of bluing, there la no relation­

ship between anthocyanin concentration and bluing. Die inner petals

of bluing roses contained as high a concentration of plgpnent as the

Inner petals of non-bluing roses.

Die concentration of tannins or other phenolic compounds, as

shewn in the same table, was found to be greater in the blue, inner

petals than In the non-bluing, outer petals of bluing roses. This

difference was not noted between Inner and outer petals of non­

bluing roses. Die analysis for phenolic compounds (tannins) In

petals of fresh cut and four day old roses from the various dif­

ferential nutrient plots did not show any relationship between the

concentration of potassium and nitrate in the substrate and the

concentration of tannins in the petals. Die petals analyzed In

these tests were the outer and mid petals and did not include the

Innermost petals which shew the greatest tendency towards bluing.

Diis information is presented in Ifeble 10.

A quantitative analysis for anthocyanin made on the same roses

indicates that lew potassium may adversely affect the formation of

anthocyanin pigment in roses. Diis trend was noticeable from the

aiialysis of the outer petals of fresh cut roses but not from the -kk- ana lysis of "the mid petals of the same rosea four days later* The complete resultB of the analyses summarized in Table 10 are pre­ sented In Appendix A to S.

Die hydrogen-ion concentration of the cell sap was found to decrease as roses aged after being cut. The decrease, however, was much greater in bluing than non-bluing flowers. The measurement of the hydrogen-ion concentration of roses from low, medium, and high potassium plots showed that the pH of the expressed cell sap varied directly with the concentration of potassium in the sub­ strate. This was true for fresh cut roses and also for bluing and non-bluing four day old roses. The cell sap of bluing roses was generally found to have a pH value in excess of 5*35* These results are shown in the data presented in Tables 11 and 12.

The results of the mineral analysis of rose petals presented

in Tables 13 and lV indicate a direct relationship between bluing of Better Times roses and the potassium content of the petals.

Bluing flowers were found to contain more potassium than non-bluing flowers. Likewise bluing petals were found to contain more po­ tassium than non-bluing petals from the same blooms. This dif­ ference did not exist between inner and outer petals of non-bluing flowers. Analysis of fresh cut roses showed that the potassium content of the petals varied directly with the concentration of potassium in the substrate.

Leaf analysis, Table 1^, showed an interaction between potassium and calcium with calcium being greatly decreased at high potassium -fc3- lovele. Diis was not evident from the analysis made on petal tissue.

Die absorption spectra of extracts from blue, non-blue, and fresh petals are shown in Figures 2, 3, and An anthocyanin pignent having a characteristic absorption peak at 515 mu was present in each of the three types of petals. An anthoxanthin pigment showing a characteristic absorption peak at 355 mu was also cannon to each. It is shown, that while on an area basis (Figure 3) freBh petals contained a greater quantity of this anthoxanthin pig­ ment than did the blue petals, for a given concentration of antho­ cyanin pigment (Figure 4) the blue petals contained proportionally a greater amount of the anthoxanthin pigment. Per unit area, older, non-blue petals, contained less of this anthoxanthin pigment than either fresh or blue petals.

An extract containing the anthocyanin pigments of Centaurea cyanus prepared in the same manner as has been described for similar extracts from roses, shewed an absorption peak at ^18 mu.

A comparison of the absorption spectrum of this extract with that of an extract from freshly cut Better Times roses (Figure l)

Indicated that the anthocyanin pigment in both was probably the same. Die slight difference in the location of the absorption peak of the two extracts could be due to impurities contained in the extracts. The anthocyanin pi^nent contained in flowers of Centaurea cyanus was identified by Willetatter and Everest (59) cyanln. TABLE 9 . Relative Concentration of Tannine and Anthocyanin* in Hid and Cuter Petals of Bluing and Non-Bluing Rosea. (5 days after cut).

Bluing Ploifers Non-Bluing Flcwers Rose Optical Density Rose Optical Density No. Plgnent Extract Tannin Test Solu. No. Plonant Extract Tannin Test Solu Mid Outer Hid Outer Mid Outer Mid Outer

1 0.264 0.357 0.367 0.284 1 0.284 0.276 0.284 0.319 2 0.319 0.357 0.431 0.319 2 0.284 0.301 0.347 0.301 3 0.337 0.328 0.357 0.268 3 0.337 0.310 0.284 0.268 4 0.268 0.284 0.337 0.252 4 O.292 0.292 0.292 0.292

5 0.319 0.301 0.409 0.292 5 0.284 0.244 0.292 0.284 6 0.301 0.337 0.398 0.292 6 0.301 0.284 O .328 0.319 7 0.310 0.367 0.337 0.252 7 0.244 0.301 0.319 0.276 8 0.328 0.301 0.357 0.292 8 0.284 0.284 0.292 0.260

9 0.260 0.301 0.310 0.301 9 0.252 0.284 0.284 0.292 10 0.268 0.292 0.319 0.292 10 0.276 0.3^7 0.319 0.310

Avg,, 0.299 0.324 0.362 0.285 0.284 0.292 0.304 0.292 -2- directly belcw the flcwer bud, wilting of petals and foliage, rapid

opening of bud, and failure of the bud to open. Any one of these

factors may limit the longevity or otherwise reduce the quality of

the flower.

It is known that the stage of maturity at the time of

cutting and the subsequent treatment of the bloom can influence

the keeping quality of most flowers. It iB also logical to assume

that the environmental conditions, including mineral nutrition, under which a flower developed, would likewise affect the keeping quality of the bloom.

Dae problem of ’’bluing" of the petals is one of the more

serious difficulties encountered from the standpoint of keeping quality of red roses. The Better Times rose is known to be a variety which blues very badly. Die petals of this variety will usually change from a normal rose pink to an unattractive magenta

or fuschia purple In a matter of two to three days or less after being exposed to ordinary room conditions. Bluing of this rose also often occurs during storage. Die flowers of this variety haie not been observed to blue when left to age on the plant.

The investigations reported in this paper are concerned chiefly with the effect of the potassium and nitrogen nutrition of the rose plant and of two commercial flower preservative preparations on the keeping quality and color changes in the Better Times Rose. -47-

TABIB 10* Effect of Nitrogen end Potassium Concentration In the Soli on the Relative Concentration of Anthocyanlns and Tfcnnlns In Petals of Better Times Roses (Average of ten roses from eaoh plot).

Optical Density Plot Salt Conct. Pipmsnt Extract Tannin Test Solu. MO3 K Fresh Petals Old. Petals Presh Petals Old Petals

(Bench II)

1 LL .396 .295 .262 .296 2 M L .421 .276 .296 .273 3 H L .427 .262 .296 .280 4 L M .446 .261 .304 .304 5 M M .456 .279 .304 .268 6 H M .426 .292 .299 .307

7 LH .442 .2 68 .329 .303 6 M H .430 .281 .303 .299 9 H H .497 .305 .310 .297

(Bench III)

1 LL .402 .267 .331 .296 2 ML .410 ,248 .300 .295 3 HL .385 .251 .320 .291

4 L H .490 .285 .322 .294 5 MM .412 .254 .320 .300 6 HM .449 .281 .335 .300 w 00 s 7 LE .446 -j .330 .307 8 ME .453 .302 .308 .307 9 HE .451 .322 .312 .302 48-

TABIZ 11, Hydrogen-ion Concentration of Sxpresaed Cell Sap of Blue, Non-Bluing, and Fresh Cut Roeea from Low and High Potassium Plots,*

Average of Roses Tested pH Range pH Readings

Blue, high £ 5-35 - 5.52 5.^5 Non-bluing, high K 5-28 - 5.35 5.32

Blue, low £ 5.30 - 5.50 5.40 Non-bluing, low K 5.17 - 5.35 5.26

Fresh, low K 5.00 - 5.31 5.13 Fresh, high £ 5.18 - 5*29 5.24

* Result of 5 determinations for each group. -U9-

TABIE 12. Relation of Soli Nitrate and Potassium Levels to the Hydrogen-ion Concentration of Expressed Cell Sap from Petals of Better Tines Roses.* (Four Days After Cut).

Salt Conct. In Soil Average of N 03 K pH Range pH Readings

LL 5.17 - 5-^5 5.30 M L 5-22 - 5.39 5.32 HL 5.18 " 5.39 5-27

L M 5*29 - 5.39 5.29 MM 5.12 - 5-50 5.32 HM 5.15 - 5*^0 5.29

L H 5.28 - 5*^5 5.35 MH 5.29 - 5.^9 5.33 HH 5.23 - 5-^9 5.39

* Result of 10 determinations per plot* a t oc. iea Aaye iea Analyeie Mineral Analyte Mineral Conct. Salt NO n oi lt C Po K Ca K Plot Ca K Plot Soliin

pret (ecn) pret (percent) (percent) mxt* w (percent) s (percent) e wst-* K 3 K www at s a: hhh C

W ft w WHH HHH HHH 0 0 # HHM HHH H H H III III III vo a>*^ a\vn u* ro h ■ 5 r r f .

e t* o HHH HHH HHH • •• it* • * • "j-jv© 00-TVJ 0\U>VJ1 • CO \Jl\OVO OdOVO 0)0)0 5 !f 0 ? 6 P I res Ul OOO OOO OOO 0 » « * It* ••• 1 & & & tste r & S HHH HHH HHH n HHH HHH HHH HHH HHH HHH 8 9 III III III vo od-j own u) ro h If SS. o 6 *WHH « • I HHH• • HHH H —3 Qp On-J-'I ro OnVJI ■tnjivd £-vovo o j ^-ui ?!

OOO OOO OOO O c* • •• ••• ••• C H

HHH HHW HHH

p - 'f r O -T -T - 51-

T A B U 14. Potassium and Calcium Content of Petals of Five Day Old Roses. (Dry Wt. Basis).

Mineral Analysis E Oa Source & Trpe of Sample (percent) (percent)

Low E, blue, all petals 2.63 0.18 Low E, non-blue, all petals 2.22 0 . 1 8

High E, blue, all petals 3.33 0.18 High K, non-blue, all petals 2 . 5 0 0.16

Lew E, blue, inner petals 2 . 3 8 0.16 Low K, blue, outer petals 2 . 1 7 0.16

Low E, non-blue, inner petals 1.79 0.16 Lew E, non-blue, outer petals 1.99 0.16

High E, blue, inner petals 3.28 0.16 High E, blue, outer petals 2 . 6 8 0.16

High E, non-blue, inner petals 2.54 0.16 High E, non-blue, outer petals 2. 2 5 0.16 TABUS 15. Nitrogen, Potassium, and Calcium content of Leaf Samples Collected from Rose Test Plots. (Dry Weight Basis).

Soli Leaf Analysis Plot Tests December 15, 1951_____ February 1. 1952______March 1. 1952 NO^EN E C a N E Ca N E Ca (percent) (percent) (percent) (percent) (percent) (percent) (percent) (percent) (percent)

II-l L L 2.95 1.68 0.94 3.61 1.89 I.56 3.76 1.64 1.13 2 M L 4.20 1.75 1.16 3.61 1.86 2.00 3.21 1.58 1.20 3 H L 3.73 1.50 1.23 3-59 1.71 I.56 3.36 1.50 1.04

4 I M 4.04 2.50 0.77 3.39 2.29 1.07 2.95 2.06 1.10 5 M M 3.57 2.17 1.13 3.58 2.20 1.27 3.61 2.06 1.23 6 E M 3.75 2.29 O.98 3.84 2.14 1.00 3.9^ 1.86 1-53

7 L H 4.25 3.78 O.56 4.00 2.92 0.77 3.77 2.82 0.80 8 M H 4.29 3.98 0.44 3.80 2.66 0.83 3.99 2.44 0.74 9 H H 4.62 3.42 0.58 3.99 2.82 0.98 4.08 2.66 0.56

III-l L 1 3.48 1.71 1.16 3.10 1.75 1.71 2.89 1.75 1.64 2 M L 3.64 1.99 I.27 3.56 1.46 1.53 3.35 1.79 1.38 3 H L 3.78 1.64 1.20 3.74 1.58 1.42 3.85 1.36 1.38

4 L M 3.89 2.59 0.80 3.36 2.63 I.56 3.45 2.29 1.27 5 MM 3.76 2.20 1.04 3.78 2.42 1.42 3.84 2.14 1.53 6 5 M 3.93 1.92 1.00 3.25 2.29 1.42 3.38 I.83 1.27

7 L E 4.08 3.42 0.42 3.76 3.00 0.63 3.30 2.56 0.63 8 M E ---- 3.98 0.42 4.14 3.16 O.56 3.97 2.60 0.77 9 H E 4.58 3.78 0.35 4.12 3.16 0.^6 4.26 2.50 0.77 By tract from retals of Central rea cyamiB. Extract from retals of Better Times mses.

0. 1-

00 WAVSLEKGTK - mu

FIGURE 1. ABSORPTION SPECTRA OF ANTHOCYAKIN EXTRACTS OPTICAL DE^*SI?K 0.0 •j • •j j . 300 FIGUHE ?. ABSORPTION S’~SCTRA Of ANTHOCYANIS EXTACIS. S’~SCTRA ANTHOCYANIS Of ABSORPTION ?. FIGUHE fcAVEUKGTH mu - fcAVEUKGTH xrcsfo e, l*iten, from mm. Extracts • * 1 • \ s / 500 ron-blue retals. ron-blue blue retals. blue tfisce. OPTICAL DEN3ITY 0.0 0.1 0.4- . o Ci 0.7- 1 ■ f w 1 . . 0 1 300 - - FIGURE FIGURE 3 ABSORPTION SPECTRA AKTHOXAKTHINOFEXTRACTS. . WAVELQiGTH -mu WAVELQiGTH Uoo xrcsfo tn 1*J ten, discs. mm. from Extracts — — blue petals. blue — — -•non-blue retals. -•non-blue ■freshretals. OPTICAL DENSITY 0.3 0 0.1 0.0 .? 300 FIGURE L. ABSORPTION SPECTRA OF AKTH0XAIJTR1N OF EXTRACTS. SPECTRA ABSORPTION L. FIGURE Extracts frmc anthocyar.infrmc Extracts solutions feul concentration.equalof ’ a 'AVELEWGTO Uoo fresh metals. fresh blue retals. blue - mu -

-3-

HEVIP.V OF LITERATURE

I. Relation of Mineral Salt Nutrition to Keeping Quality of Cut

Flowers.

Though a great deal of research has been done on the effect of mineral salt nutrition in regard to the growth and production of floriculturel crops, relatively little has been published on the relation of the mineral salt nutrition to the keeping quality of the product. However, in 193^> Darner (13) in reviewing work done at the University of Illinois, stated that over fertilization, overwatering, or excessively high temperatures during the growing period resulted in soft flowers and poor keeping quality. It is generally agreed that excess nitrogen in the substrate favors the production of carbohydrate deficient plants end plant parts. From a horticultural standpoint this condition is usually associated with a product of reduced quality.

Several workers have made recommendations concerning the level of the various mineral salts which should be maintained in green­ house rose soils for optimum growth and production. Iaurie and

Kiplinger (28) have suggested levels of 10 to 28 parte per million of nitrates, 8 parts per million of phosphorus, and 20 to 40 parts per million of potassium. Seeley and Poet (81) stated that soil nitrate levels should be maintained between 28 and 100 parte per million for optimum production and stem length. Though the figures are different these levels seem to be actually the same as was recommended by Iaurie and Kiplinger (28). In each case the levels - 57-

STUDY III: SONS CHEMICAL CHANGES IN PETALS OF BETTER TIMES ROSES

RESULTING FROM THE USE OF FLOWER PRESERVATIVES.

MATERIAL AND METHODS

A. GENERAL PROCEDURE

Aa has been reported In the review of literature it has been shown that certain preparations are effective in maintaining a desirable color In red roses for an extended period of time. This study was undertaken to determine some of the chemical changes occurring In the petals as a result of the use of several commercial flower preservatives.

These chemical studies were made in conjunction with tests being run to determine the relative effectiveness of two new proprietary preparations, "Aladdin" and "Survival 77" as flower preservatives. Roses used in the study were obtained from Walter

Engel, a local rose grower, partially with funds donated by the

General Floral Products Co., Inc., Winchester, tessachusetts, the manufacturer of Aladdin. All roses used In the tests were freshly cut and selected for uniform quality and maturity.

To obtain information concerning the best time for using the preservative solutions and to simulate conmercial conditions as nearly as possible the treatments were divided into three phases: a wholesale, a retail, and a consumer phase. For the wholesale phase the flowers were held with stems in water or a preservative solution in a refrigerator at ^+0* F. for 2k hours. After this -56- phaee the roees were removed from the original containers, stems shortened approximately one inch, placed in fresh preservative solutions or water, and kept in the refrigerator at 40* F. for another 2k hour period. Following completion of the retail phase the stems were shortened approximately another inch and again placed in fresh solutions for the consumer phase. During this phase the roses were kept at roam temperature.

Three flower preservatives "Aladdin;, manufactured by the

General Floral Products Co., Inc., Winchester, Massachusetts;

"Survival 77"> manufactured by Survival Products, Boston, Massa­ chusetts; and "Floralife", distributed by Floralife, Chicago,

Illinois were used in the keeping quality study. Each was used at the concentration recommended by the manufacturers. For Aladdin this was 2 l/k ounces per gallon of water; for Survival 77/ 3 ounces per gallon of water; and for Floralife, 2 l/k ounces per gallon of water. Tap water was used as a check treatment. A summary of the various treatments is given in Table l6. This keeping quality test was conducted three times. Only the results of the chemical analyses are reported in this paper.

B. CHEMICAL TESTS

The measurement of tannins, anthocyanin, hydrogen-ion concen­ tration, and mineral content of the petals and the absorption spectra of the pigments were made according to methods outlined in

Study II.

Analyses for potassium and calcium and the pH measurements -59-

TABLE 16. A Summary of Treatments Used In the Study Concerning the Effect of Certain Flower Preservatives on the Keeping Quality of Better Times Roses.

Treatment TREATMENT Wholesale Phase Retail Phase Consumer Phase No. (40*F.. 24 hours) (40*F.. 24 hours) Room Temperature

1 Water Water Water 2 Water Water Aladdin

3 Water Water Survival 4 Water Water Floralife

5 Water Aladdin Water 6 Water Aladdin Aladdin

7 Water Survival Water 8 Water Survival Survival

9 Water Floralife Water 10 Water Floralife Floralife

11 Aladdin Water Water 12 Aladdin Water Aladdin

13 Aladdin Aladdin Water 14 Aladdin Aladdin Aladdin

15 Survival Water Water 16 Survival Water Survival

17 Survival Survival Water 18 Survival Survival Survival

19 Floralife Water Water 20 Floralife Water Floralife

21 Floralife Floralife Water 22 Floralife Floralife Floralife - 60 - wore made on representative flowers of selected treatments of the first and second series of treatments. Sampling for these deter­ minations were made on the fourth day after the roses had been placed at room temperature. Tannin and anthocyanln measurements were made on representative flowers of selected treatments from each of the three series of treatments. These measurements were made on both the second and fourth day after the roses had been placed at room temperature. Extracts used in determining the ab­ sorption spectra of the pigments were obtained from clear, pink,

inner petals and from blue, outer petals of flowers of the Sur­

vival 77 and Aladdin test series and from magenta colored petals

occurring when no preservative was UBed. -61-

RESULTS

It was noted in the flower preservative testB that the pre­ servatives, Aladdin and Survival, were superior to Floralife in maintaining an attractive, bright, petal color and in prolonging the life of cut roeee. The use of these two flower preservatives, while not maintaining the original petal color, did prevent the inner and mid petals of Better Times roses from changing to a dull magenta color as occurred In the roses placed only in water. Hie

outer petals of roses placed in preservative solutions, however, developed a bluiBh hue. Beneficial effects from the use of the flower preservatives were observed only when they were used at the consumer phase, i.e. at room temperatures.

Die quantitative tests for anthocyanin indicated that the better petal color associated with the use of the flcwer preservatives was not due to a higher concentration of anthocyanins. Data presented

in Tables 17 and 16 show that in most instances the use of these preservatives resulted in a slight decrease in anthocyanin content.

Likewise there was no correlation between the tannin content and color of petals. Analysis of the outer bluing and Inner pink petals

of the Bams preservative test flowerb showed that the outer petals had a slightly smaller concentration of phenolic compounds but a higher concentration of anthocyanins. The results of the pH measurements, also presented in Table 17, show that lower pH values were correlated with the use of the flower preservatives and better -62- flower color.

The results of the mineral analysis (Table 19) of petals from roses of the different treatments showed that as In Study II there was a positive correlation between bluing and potassium content.

It was found that as a flower aged after being cut the concentra­ tion of potassium, calculated on a dry weight basis, Increased.

This Increase in potassium content was less in flowers that had been placed in the preservative solutions than in the check flowers that had been in water. Survival 77 depressed the potassium con­ tent more than aladdln. With one exception, the poorer appearing flowers contained a higher concentration of potassium than the better appearing flowers. A composite sample of badly blued petals showed a higher concentration of potassium than any other sample tested. Neither age or preservative treatment had any apparent effect on petal calcium concentrations.

A comparison of the absorption spectra of pigment extracts from the blue outer petals of rosea of the Survival treatments, clear pink inner petals of the same flowers, and of magenta petals

of roses from the water (check) treatment, shewed that the same anthocyanin and anthoxanthln pigments were present in each of the above types of petals. The anthocyanin extracts (Figure 5) showed a characteristic absorption peak at 515 mu. Hie absorption peak of

the anthoxanthln extracts (Figure 6) was at 355 • Per unit area,

the blue outer petals of Survival test flowers were found to con­

tain a higher concentration of the anthoxanthln pigment than the -63- plnk 1 nner petals of the sane flower or the magenta petals of roees from the check treatment, but lees than that contained In petals of freshly cut roses* In relation to a given concentration of anthocyanin pigment the pink petals again contained a smaller per­ centage of the anthoxanthln pigment than the magenta petals or petals from freshly cut roses* Fresh petals shewed a higher ooncen

tration of the anthoxanthln pigment than did the magenta petals. TABLE 17. Some Effect* of Flower Preservatives on the Chemical Constitution of Rose Petals*

Optical Density of Optical Density of Average Treatment Anthocyanin Brtract Tannin Test Solu. pH Range of pH da,y 4th day 2nd day 4th day Reading Condition of flo w n on 4th Day

Test #1 H20-fi20“H20 0.303 0.276 0,298 0.275 5.21-5.25 5.23 Poor - petals m&eenta in color. H20-Sur.Sur. 0.274 0.244 0.307 0.269 4.92-5.05 4.99 Fair - inner petals clear pink, bluing on outer petals. HgO-Alad .Alad. 0.246 0.282 0.310 0.328 5.12-5.19 5.15 Fair - inner petals clear pink, blulzut on outer petals. Test H^O-H^O-HgO 0.244 0.235 0.277 0.282 5.08-5.31 5.22 Poor - petals assents in color. HgO-HgO-Sur. 0.256 0.244 O.316 0.364 4.98-5.05 5.01 Fair - inner petals clear pink, bluins on outer petals. H^O-HgO-Alad. 0.261 0.258 0.339 0.326 5 .00-5.20 5.09 Fair - inner petals clear pink, bluin*z on outer petals.

Sur.-HgO-HgO 0.223 0.220 0.280 0.288 5.15-5 5.37 Poor - petals assents in color. Sur.Sur.Sur. 0.264 0.216 0.238 0.242 5.02-5.28 5.13 Fair - inner petals clear pink, hluins on outer petals. HgO-Sur.Sur. 0.276 0.264 0,288 0.297 5 .10-5.20 5.18 Fair - inner petals clear pink, blulxut on outer petals. HgO-Alad.Alad. 0.260 0.250 0.288 0.325 5 .10-5.20 5.15 Fair - inner petals clear pink, bluing on outer petals.

* Average of duplicate samples from three representative roses from each treatment. TABLE Id . Relation of the Relative Anthocyanin and Tannin Content of Inner and Outer Roee Petals to Plover Preservative Treatments.*

Optical Density of Optical Density of Anthocyanin Extract Tannin Test Solu. Treatment 2nd day 4th day 2nd day 4th day Condition of flowers inner outer inner outer inner outer inner outer on 4th day.

BgO-HgO-BgO 0.265 0.294 0.244 0.272 0.319 0.297 0.374 0.333 Generally poor, petals maaenta in color.

HgO-^O-Sur. 0.23* O .309 0.238 0.3 01 0.305 0.3 01 0.319 0.307 fair - inner petals mostly pink, outer bluing.

HgQ-HgO-Alad. 0.264 0.314 0.277 0.262 0.319 O .305 0.314 O .297 fair - Inner petals mostly pink, outer Maine.

I^O-HgO-riora. 0.238 0.314 0.252 0.2 9 8 0,314 0.2 90 0.335 0.314 fair - inner petals mostly pink, outer bluing.

* Average of duplicate samples from four representative roses from each treatment. -66-

ZAB12 19* Effect of Flavor Preservative Treatments on tho Potassium and Calcium Content of R o m Potalo.* (Dry ttelgbt Basis).

Mineral Analyses Treatment K Oa Condition at Sampling** (percent) (peroent)

(preservative Fresh petals 1.79 0.1k Hg O - ^ C ^ B g O 2.20 0.1? Potalv magenta, o o o m of Inner

HgO-Sur.-Sur. 1.68 0.12 inner pvtaiv mostly clear T'lrir. ****** ---- He 0-Alad.-Alad. 2.10 0.15 Innor pstala mostly clear ^ nV petals bluing.___

Sur.-HgO-HgO 1.99 0.1k Petals m a g e n t a , some of Inner bluHur rndit.------Alad. —HgO—HgO 2.20 0.15 Petals mostly magenta, seme nt ftlWi0; WV-

(Composite) 2.42 0.15 blued petals*

(preservative Fresh petals 1.66 0.16 HgO-HgO-HgO 2.38 0.18 Petals mostly magenta, of Inner petals bluing exten­ sively. _____ HgO-HgO-Sur. 2.10 0.1k Tnner petals mostly clear

"1nht a a ± m r ' rrt(*lr — HgO-HgO-Alad . 2.10 0.1^ Inner petals mostly clear rli^- r r W ^iwipg.__

Sur • -Sur. -Sur • 1.83 0.16 Inner petals mostly clear pink, outer petals ------HgO-Sur.-Sur. 2.1k 0.15 Inner petals mostly clear TiiBlr. oiitay ------HgO-Alad • -Alad. 2.17 0.16 Inner petals mostly clear o»tar ntfflir------

Sur.-HgO-I^O 2.50 0.16 Petals mostly igenrta nf * Average of duplicate aliquots from one composite sample. Sampled after four days at room temperature. -k- are based on the Spurway (53) aystem of soil analysis. Ihe apparent discrepancy In the two sets of figures is due to different methods of recording readings. TOila difference has led to much confusion among both research personnel and commercial growers.

Post and Fischer (^2) recommended that the concentration of potassium In rose soils be carried at 300 to 600 pounds per acre as determined by the Peech and English (36) method of soil analysis.

They state!that this Is equivalent to a concentration of 15 to 30 parts per million in terras of the Spurway system.

Relative to the effect of various nitrate levelB on the keeping quality of roses Seeley and Post (51) stated that there was no difference in the keeping quality of roses grown at levels of 50 to 1*00 parts per million. However, the production of unsalable flowers Increased as nitrate levels were raised above

10O parts per million. Biese levels were based on the Spurway (53) system of soil analysis. Due to the afore mentioned difference that exist at different institutions in methods of recording readings the interpretation of these results in the light of other work is somewhat difficult.

Culbert and Wilde (10) in a study concerning the effect of various amounts of potassium on the production and growth of Better

Times roses, In which the potassium levels were maintained at 3 to

15 parts per million, observed that the quality of the flowers, as determined by stem and flower length and weight, increased signifi­ cantly with each increment of potassium supplied. OPTICAL EEMSITY 0.5' u. 0 0.0 . 1 - \s 1 FIGURE FIGURE 5 . ABSORPTION SPECTRA OF ANTHOCTANIR EXTRACTS. ANTHOCTANIR OF SPECTRA ABSORPTION . 1)00 WAVELENGTH - mu - WAVELENGTH Extracts Extracts —. rn, ne eas-Sria treatment Survival - inner •rink,retals .— .— s * f / / blue, outer outer re blue, treatment Survival tale - ■amenta retals - water treatment. water - retals ■amenta from ten, 1 \\ m rtiecs.mm. ^ fco

Survival Survival treatment. Survlval t reatment. Survlval t water treatment, fresh retals fresh magenta petals petals - magenta oink, petals - oink, inner blue, - petals blue, outer WAVELENGTH - mu - WAVELENGTH Eytmct* (Uses mm. lU from ten, r S- r FIGURE f. FIGUREf. EXTRACTS. OF AETH0XANTHIN SPECTRA ABSORPTION 300 0.1 O.H 0.6 0.8 0.9 1.0 1.1 1.2

uisiea t o i w o OPTICAL DSKSITY 1 1.0 0 .1-0 0.0 j . . *+< . 2 7 - . FIGTHE 7- A?SORPTION SPECTRA. OF ATTHOXAFTHIN EXTRACTS. SPECTRA. ATTHOXAFTHIN OF A?SORPTION 7- FIGTHE feul concentration. equal solutions of anthocyanin from Extracts W W A.7ELENGTHmu - 400 fresh retals. fresh rink, inner retals - retals inner rink, rne^enta retnl - s

water treatment, water uvvl treatment. Survival

-70-

DISCU8SI0M

The results of the comparative keeping quality tests reported

In Study I would seem to Indicate that, except for bluing, factors

other than the nitrogen and potassium status of the rose plant

limit the keeping quality of cut roses.

That light Intensity Is one of these factors Is shown by the

poorer keeping quality exhibited by roses cut during the winter and early spring months. It was during this period also that the

greatest percentage of premature wilting of petals and stems

occurred. A commonly held opinion among rose growers la that

additional potassium added to the soil during the winter months will decrease the incidence of stem wilting during this period.

Because high potassium levels did not exist In any of the test

plots at the time of the observed Increase In wilting of the stems,

the validity of this assumption cannot be settled by these tests.

On the basis of the observation that the Incidence of stem

and petal wilting Increased during a period of lower light ln~

tensities It would seem that keeping quailV would to seme extent

be correlated with a carbohydrate deficiency In the plant. That

keeping quality of roses is often limited by the carbohydrate

supply of the plant Is also Indicated by the observation made by

Knapp (21) that In order to obtain maximum keeping quality of

roses during the winter months it was necessary to delay cutting

until three or four petals were unfurled. This delay In cutting - 71- vould h a w allowed for a greater accumulation of oarbohydrates In

tha •tarn, leaves, and bloc* of tha flowering shoots.

Since there la reason to believe that carbohydrate supply

can be a limiting faotor in regard to the keeping quality of roses

and as It is known that the nitrogen supply available to a plant

does affeot the carbohydrate supply of the plant and plant parts,

It Is logical to assume that soil nitrate levels would directly

affect the keeping quality of cut roses. It Is not clear why

such a correlation was not noticeable In this study. Undoubtedly,

the quality was affected to some extent,, but not to a degree where

the differences could be detected by subjective means of Judging

keeping quality.

During the first four comparative tests conducted the nitrate

levels, as shown by the results of the soil tests, were not main­

tained In any of the plots at the "high" concentration. Biis would

seem to be an explanation for the absence of any trend or correlation

between keeping quality and soil nitrate concentrations In these

tests. At the time when the last three comparative tests were con­

ducted high levels of soli nitrate existed in combination with low,

medium, and high levels of potassium and still there were no indi­

cations that the nitrogen supply had any effect on keeping quality*

However, the tine of the year during which the tests were conducted

could have directly affected the results that were obtained.

That an actual differential in the nitrogen status of the

plants did exist during this time was shown by the light colored -72- foliage and excesalve leaf fall observed on plants growing in the lew nitrogen plots during the above period. Hie results of leaf analysis mads on samples collected during this time are presented

In Thbles 7 end 8 . These analyses failed to shew that a nitrogen deficiency existed in any of the plants. The lowest nitrogen con­ tent reported was 2 . 8 6 percent of the dry weight of the leaves.

The "critical" level of leaf nitrogen content for most plants is usually thought to be around 2.00 percent. This apparent discre­ pancy was probably due to method of sampling. As explained in the section on methods, leaf samples consisted of leaflets of the basal

second and third 5 -leaflet leaves of a mature flowering shoot.

Apparently nitrogen was translocated from the lower portions of the plant to these upper leaves to such an extent that the analysis

of these leaves for nitrogen failed to reveal the true nitrogen

status of the plant. It is of Interest to note that the leaf nitrogen content of samples collected from plants growing in the

low nitrogen-high potassium plots were higher than that found in

any of the samples collected from the high nitrogen plots when in

combination with low or medium potassium.

Potassium content of leaf samples, on the other hand, corre­

lated directly with the results of the soil tests. That the potas­

sium content of the soil was high during the time that the excessive

bluing of the petals was observed was clearly shown by both the soil

and leaf analysis. Visual symptoms of potassium excess were also

apparent on plants of the high potassium plots during this time. -73-

Blulng of the roses cut from the high potassium plots was general regardless of the nitrogen level of the plot. It should he pointed out, however, that the potassium concentration in the soil during this time was excessively high. Concentrations of such intensity would not ordinarily be found under commercial practices. However, it is possible that such high potassium concentrations could exist for short periods of time following heavy applications of potassium fertilizers. This could be a possible explanation of occassional reports by rose growers of periods during which bluing becomes a more serious problem.

To determine the chemical effects which the high potassium concentrations had on the metabolism of the rose which resulted in the excessive bluing and to determine some of the chemical factors associated with bluing and non-bluing of Better Times roses was the object of the further investigations. As reported in the review of literature several factors are usually considered to be of impor­ tance in influencing petal color.

From this study it does not seem that changing amounts and mixtures of anthocyanins had any great effect on bluing of cut roses regardless of the potassium status of the plant from which the rose was cut or aiy subsequent treatment of the flower.

Spectrographic analysis of anthocyanin extracts from bluing, non-bluing, and fresh petals of both the fertilizer and flower preservative test series showed that all had a characteristic absorption peak at 515 mu Indicating that there was no variation -74- in the nature of the extracted anthocyanln pigment. Quantitative measurements of the anthocyanln pigment likewise failed to Indicate any relation between concentration of pigment in the petals and bluing or non-bluing of the petals. It was found that the concen­ tration of anthocyanln pigment in bluing petals was as great or greater as the concentration of the pigment In non-bluing petals of the same age.

It has been shewn that the pH of the cell sap does under some condition influence the color of flowers. Small (5 2 ) stated that it Is likely that different colors in the same cells of petals at different times are due to different pH values affecting the same anthocyanln. Robinson (46), however, reported that In the case of two different rose varieties that were characterized by a change in petal color as the bloom reached senescence there was no change In pH values as the flower color changed. The results reported in this paper shew that in the case of Better Times roses there was a decrease in the hydrojen-ion concentration of the cell sap as the flower blued. It was also shown that flcwers cut from plants growing in the high potassium plots had a higher pH value than did the ones cut from plants of the low or medium potassium plots. This would indicate that the more rapid bluing of roses found to be associated with high potassium levels might be caused by a decrease

in hydrogen-ion concentration brought about by a higher content of potassium. An Inverse relationship between potassium content and

pH was also found to exist in blooms from the flcwer preservative

tests. Here it was found that the blooms characterized by better -75- petal color had the lower pH values and also lower potassium con­ centrations. But the fact that the brighter, more attractive color resulting from the use of the flower preservatives, was not the normal color of freshly cut Better Times roses of approximately the same pH value indicated that some factor other than pH was involved.

It was found that the dried tissue of bluing roses and petals contained a larger percentage of potassium than non-bluing roses.

It was likewise noted that the potassium content, calculated on a dry weight basis, increased as the cut rose aged. Whether this was due to an actual increase in the content of potassium or due to a dilution effect was not determined. Analysis showed that flowers with stems in preservative solutions contained a smaller concentra­ tion of potassium after four days than those with BtemB in water during this period. This would indicate that in the latter case, at least, there was an actual increase in potassium content. If this were the case, then the flower preservative solutions apparently were effective in decreasing the translocation of potassium from "the stem or leaves to the petals.

Biis could be taken as an Indication that the ash content of the cells, specifically potassium, is a factor involved In the bluing of Better Times roses. However, again the fact that the pink color resulting from the use of flcver preservatives is not the normal color of Better Times roses indicates that some factor other than potassium content was involved. No apparent correlation between calcium content and bluing was found to exist. -76-

The role of co-pigments in modifying flover color he* been discussed In -the review of literature. Curry (12) reported that the cause of bluing in red roses was a deficiency of tannins which normally acted to stabilise the color of the anthocyanln pigment.

Other workers have shown that certain of the anthoxanthin pigment a also modify flower color. The results obtained in the present studies show conclusively that bluing of the petals of Better Times roses was not due to a lack of tannins. Conversely, it was found that bluing petals contained a higher concentration of tannina or other phenolic compounds than non-bluing petals. No consistent differences in tannin content was detected in inner or mid petals of roses cut from the various differential fertilizer plots. As it was observed that roses cut from plants of the high potassium plots

showed a greater tendency towards bluing and as it was found that blue petals contained a higher concentration of phenolic compounds

than non-blue petals, the above results indicate that the increase

of tannins or other phenolic compounds associated with bluing was a result of and not the cause of the bluing.

The same anthoxanthin pigment was found to be present in all

extracts tested. However, the concentration of the anthoxanthin

pigment extracted from blue, non-bluing, and fresh petals varied

considerably. The relative concentration of the pigment in these

various extracts can be calculated on the basis of the Beer-Lambert

laws of light absorption. These laws state that the amount of

light energy absorbed is a function of the number of molecules in

A -5-

Knapp (21) In a rather extenaive survey concerning factors

affecting keeping quality of roses observed that with roses grown

In soil there was a sharp up-awing In premature wilting as the

nitrate level was Increased and that the amount of wilting associ­ ated with the low and medium nitrate levels decreased as potassium was Increased. In general his results shewed that roses of better

keeping quality were obtained at low nitrate levels If potassium was also low, but at higher levels of potassium, 20 to 60 parts per million, the beat keeping quality was associated with Intermediate

levels of nitrogen.

With snapdragons, Howland (19) found that flower production and quality was not affected appreciably by relatively large varia­

tions In nitrogen and potassium fertilization, soil reaction, soil

type, soil moisture, day length, or light intensity. Laurie and

Kipllnger (29) stated that the softness of stem and burning of the

edges of petals in many soft varieties of carnations during the winter may be due to high nitrate and low calcium or potassium.

Hone of the work that has been cited proves conclusively that mineral salt nutrition in Itself plays a decisive role In determi­ ning or limiting the keeping quality of cut flowers. The varia­ bility in results that have been obtained Indicate a need for more work in this field.

II. Effect of Post-Harvest Treatments on Keeping Quality of Roses

A. Standard Handling and Storage Practices

The stage of maturity as which a rose Is cut Is known to -77- the path of the light beam.

On the baa 1b of unit area it was found that the concentration of the anthoxanthin pigment wae greatest in petals of freshly cut flowers but greater in blue than in non-blue petals. Their re­ spective absorption spectra shewed that bluing petals from roses produced in the high potassium plots, contained approximately k2 percent more of this pignent for a given petal area than non-bluiig petalB. By extracting the anthoxanthin pigments from acid extracts of petals containing an equal concentration of anthocyanin it was found that for a given concentration of anthocyanin, an extract from blue petals contained more of the anthoxanthin pignent than did the petals of freshly cut roses. This is an indication that bluing is accompanied by an increase in concentration of the antho­ xanthin pignent. However, the difference in the concentration of this pigment found in extracts from fresh petals and blue petals was relatively small. Extracts from the pink petals from roses of the preservative treatments was found to contain considerably less of the anthoxanthin pignent than any of the other extracts.

On the basis of these results it seems that the effect of tannins as a co-pigment involved in the bluing of Better Times roses is iiegligible. However, there are indications that antho- xanthins may be a factor.

Of the possible factors involved in the bluing of Better Times roses that were studied in this investigation it seems that the pH of the cell sap, the potassium content of the petals, and the conosn -78- trat ion of the anthoxanthin pigment found to occur in petals of

Better Times rosea are most likely to be of importance in this color change.

It is very probable that the color change is caused by a com­ bination of factors rather than by any one factor. Likewise, it seems that different factors are involved in the bluing of the petals that normally take place in Better Times roses and the retention of a lively pink color that results from the use of certain flower preservatives.

Studies reported in the review of literature show that antho- cyanin extracts of red roses, presumably cyanin, "in vitro" do not change from a pink to a purple or bluish color until a pH value of seven is reached. In this study It was found that the pH of even badly blued roses did not reach this value. It is realized, howevw; that the results obtained "in vitro" can not always be used to in­ terpret phenomena occurring in living cells.

A comparison of pH measurements of the cell sap from blue and non-bluing roses showed that bluing most often occurred at pH values above 5.39- However, there was no consistent line of demarkation.

Both bluing and non-bluing roses were tested that showed pH values above and below this value.

Higher potassium concentrations were consistently found to be associated with bluing regardless of the potassium status of the plant from which the rose was cut. Oiese data lead one to believe that the bluing observed to be associated with high levels of -79- potasslum la more a direct effect of the potassium than an Indirect

one associated vlth pH values. Just what this effect might be can­ not be determined from these studies.

Chemical studies carried on In conjunction with the flower pre­

servative tests showed that the use of flower preservatives helped

to maintain a more acceptable color, but not a natural one, In all but the outer petals of Better Times roses. This retention of color was associated with lower pH values, lower concentrations of potas­

sium in the petals, and lower concentrations of an anthoxanthin pig­

ment.

Any or all of these observed variations could be contributing

factors. However, as mentioned previously, since the color was not

a natural one, It seems likely that other factors were Involved.

The chemicals contained in Survival and Aladdin are not known.

It is known that some of the other preservatives that are effective

In maintaining petal color contain aluminum sulfate. Allen (2)

and Cherney (8) have shown that aluminum, by forming additive

complexes with the anthocyanln pigment influences the color of

hydrangea bracts. A similar reaction between aluminum or some

other chemical contained In the preservatives and the anthocyanln

pigment could be responsible for the change and stability of color

noted as a result of the use of Survival and Aladdin. -80-

SUMMARY AND CONCLUSIONS

In studies reported In this paper the effect of the soil

nitrogen and potassium levela on the keeping quality of cut roaea and acme of the chemical factors aaaociated with bluing and color

retention in petals of Better Times rosea were investigated.

From these studies the following summary and conclusions are made:

1. The nitrogen status of the plant had no apparent effect

on the keeping quality of cut Better Times roses.

2. Excessively high potassium concentrations in the substrate

resulted in more rapid and more extensive bluing of Better

Times roses. Bluing of rose petals was also correlated

with a higher potassium content in the petals. Ihis

indicated that the high potassium concentration played

a direct role in the observed bluing. This may have

been due to the formation of an additive complex between

the potassium and the anthocyanln pigment.

3. Premature wilting of petals and stems of cut roses were

more prevalent during the winter and spring months than

at other times of the year. This was apparently due to

the lower light intensities that commonly prevail during

these months.

k. Changing amounts and mixtures of anthocyanins were appa­

rently not responsible for color changes in petals of -81-

Better Tines roeee.

5 . Higgler pH values, higher potassium content of petals, and

higher concentrations of an anthoxanthin pigment were asso­

ciated with bluing.

6. Phenolic compounds (tannins) were found to be higgler in

blue petals than in non-blue petals. increase, however,

seemed to be a result of and not the cause of the bluing.

7* The retention of the more desirable color found to be

associated with the use of flower preservatives was ac­

companied by a reduction in pH values and in the concen­

tration of an anthoxanthin pigment.

8. The fact that the color of petals associated with the use

of flower preservatives is not the normal color of Better

Times roses is indicative of an additive complex being

formed In the petals with a material found in the preserva­

tive .

9. Better Times roses were found to contain an anthocyanln,

having an absorption peak at 5^5 rou in an acid extract,

and an anthoxanthin, having an absorption peak at 355 n*11

in an ethyl acetate extract. Because of the close

similarity of the absorption spectra of the anthocyanln

extracts with a like extract from flowers of Centaurea

cyanus, the anthocyanln pigment is presumed to be cyanin,

a 3,5 diglucoside of cyanidin. -82-

It 1b believed that these Investigations have furnished infor­ mation of a fundamental nature which Is of value In helping to understand some of the possible reasons for bluing In Better Times

roses* Hcwever, more work needs to be done to definitely establish whether or not more than one factor Is Involved In the color changes

that take place. Likewise, additional work is needed to determine

the role played by certain flever preservatives in maintaining a more desirable petal color in red roses.

Though the work on the effoct of mineral salt nutrition of the r rose plant on the keeping quality of cut roses indicated that the

nitrogen status of the plant does not play a significant role in

determining keeping quality, it Is felt that more work needs to be

done to definitely establish this fact. Because of the commonly

held opinion among commercial growers that higher potassium con­

centrations in the soil are effective in reducing the incidence of

stem wilting during the winter months and because no "high" potas­

sium levels existed in any of the test plots during the time when

a higher percentage of stem wilting was observed to occur, it

seems especially desirable to continue this phase of the nutritional

study. -83-

Appendlx A. Relative Concentration of Anthocyanln PI guest ■ end Tannins in Petal* of Better Tinea Roeee. Plot II-l. Low Nitrate, Low Potassium.

Date Q tPt 9f Q.D. of T ^ mia Teat Soln. Bluing Cut Freeh Petale Old Petal* Freeh Petal* Old Petals Tendency*

1-31 .426 • 357 .252 .293 B 2-2 .509 .289 .276 .252 N 2-2 .538 .301 •315 .276 N 2-4 .444 .347 .393 • 372 N

2-5 .328 .276 .264 •333 N 2-6 .280 .248 .237 .289 N 2-7 .310 .306 .237 .301 N 2-7 .306 .252 .248 .252 B

2-9 .398 .324 .293 .292 B 2-9 .420 .260 .306 .297 N .396 .295 .282 .296 * N - No noticeable bluing after four days at room temperature. B - Incipient bluing of inner petals after four daye at room temp. BB - inner petals definitely bluing after four days at roam temp.

Appendix B. Relative Concentration of Anthocyanln Pigments and Tannins in Petals of Better Times Roses. Plot II-2. Medium Nitrate, Low Potassium.

Date O.D. of Pigmejnt Sxtzact O.D. of Tannin1 Test Soln. Bluing Cut P*resh Petals Old Petals Fresh Petals Old Petals Tendency^

1-31 .620 .362 .276 .301 N 2-2 .523 .297 .284 .252 B 2-4 .362 .244 .301 .293 N 2-5 .248 .297 .260 .301 N

2-5 .421 .276 .296 • 273 N 2-8 .415 .289 .276 .248 N 2-9 .438 .297 .310 .289 N 2-11 .409 .211 .284 .237 N

2-13 .328 .226 .342 .260 N 0 e -p- .280 B 2-15 VJI .260 .337 Ay*. .421 .276 .896 ,.,•223.. * N - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at room temp. BB - inner petals definitely bluing after four days at room temp. -84-

Appendix C. Relative Concentration of Anthocyanln Pigments and Tannlna In Petals of Better Times Roses. Plot II-3* High Nitrate, Lew Potassium.

Date O.D. of Pigment Extract O.D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency*

1-31 .456 .328 .253 .260 B 2-2 .611 .260 .337 .310 N 2-5 .324 .244 .293 .337 N 2-5 .337 .233 .237 -315 N

2-6 .420 .324 • 303 .293 B 2-11 .523 .244 .3*3 .268 N 2-11 .415 .284 .293 .260 N 2-11 .426 .260 .276 .268 N

2-20 .289 .204 .248 .244 N 2-21 .372 .237 .301 .244 N Avfi* ■ -.,•*27 .262 .296 .280 * N - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at room temp. BB - Inner petals definitely bluing after four days at room temp.

Appendix D. Relative Concentration of Anthocyanln Pigments and Tannins in Petals of Better Times Rosea. Plot II-4. Lew Nitrate, Medium Potassium.

Date O.D. of Pigment Extract O.D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency*

1-31 .502 .284 .256 .289 N 2-2 •553 .293 .297 .252 N 2-5 .404 • 377 .315 •357 B 2-5 .328 .241 .306 .362 N

2-6 .333 .223 .264 .264 N 2-6 .393 .281 .293 .276 N 2-9 .502 .297 .310 .268 B 2-12 .420 .244 .382 .337 N

2-12 .648 .297 .362 .342 N 2-19 • 398 .276 .260 .297 N Alfi- . .448 .281 .304 .^04 * N - no noticeable bluing after four days at room temperature. B - Incipient bluing of Inner petals after four days at room temp. BB - Inner petals definitely bluing after four days at room temp. -85-

Appendix S. Relative Concentration of Anthocyanln Pigments and Tannins In Fata Is of Better Tins a Roses. Plot II-5. Medium Nitrate, Medium Potassium.

Date O.D. of Pisment Extract O.D. at Teiurin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency*

1-31 .393 .306 .272 *319 N 1-31 .569 *337 *337 *337 N 2-4 .426 .252 .252 .252 N 2-4 .367 .215 .324 .297 B

2-8 .450 .289 .347 .272 N 2-9 .456 .289 *293 .252 N 2-9 *538 *315 .306 .276 N 2-12 *531 .252 .284 .276 N

2-13 .476 .289 *319 *347 B 2-13 *362 .244 .306 .248 N Avg. .456 .279 .304 .288 * N - no noticeable bluing after four days at room temperature. B - Incipient bluing of inner petals after four days at room temp, BB - inner petals definitely bluing after four days at room temp.

Appendix F. Relative Concentration of Anthocyanln Pigmentb and Tannins in Petals of Better Times Roses. Plot II-6. High Nitrate, Medium Potassium. 3 0 £ s ■P Date O.D. of Pigment Extract 0.. D. of Tannin CD 0 Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency*

1-31 .404 .367 .233 .293 B 1-31 .668 .306 -357 •357 N 2-4 .426 .233 .268 .293 N 2-5 *377 .208 .241 .248 N

2-5 *357 .276 .319 .362 B 2-7 .426 .252 .306 .289 N 2-8 *357 .260 .301 .306 N 2-6 .475 .342 *372 .367 N

2-11 *337 .260 .276 .226 B 2-13 .456 •315 *319 • 333 N £?&*. .428 .292 . ..*229.. ... *322- _ * N - no noticeable bluing after four daye at room temperature. B - Incipient bluing of Inner petals after four days at room temp. BB - inner petale definitely bluing after four days at room temp. -86-

Appendix O. Relative Concentration of Anthocyanln Pigment* and Ifcnnins in Petals of Better Times Roses. Plot II-7 . Lew Nitrate, High Potassium.

Date O.D. of plament Extract Q.D._ of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Ratals Old Petals Tendency

1-31 .396 .293 .377 • 357 N 2-4 .V50 .260 • 333 .310 B 2-k .4 15 .233 •319 .252 B 2-k • 393 .215 .315 .301 B

2-8 .if 63 .306 • 342 • 293 B 2-13 .398 .252 .276 .284 B 2-20 .if63 .328 .301 .276 B 2-21 .516 .293 .367 •315 B

2-29 • 553 .264 .382 • 393 N 3 “1 •377 .241 .276 .248 N Avg. .442 .268 .329 .303 * h - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at room temp. BB - inner petals definitely bluing after four days at room temp.

Appendix H. Relative Concentration of Anthocyanln Pigments and Tannins in Petals of Better Times Roses. Plot II-8 . Medium Nitrate, High Potassium.

Date O.D. of Pigment Extract O.D. of Tannin1 Teat Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency*

1-31 .382 .244 .268 .264 B 1-31 .324 .268 .262 •337 N 2-5 .362 .260 .280 .328 B 2-6 .415 .289 .315 •315 N

2-11 .620 *297 • 337 .280 B 2-16 .438 .252 • 293 .276 N 2-20 .456 • 3^7 .315 • 357 N 2-20 .420 .272 .252 .284 B

2-29 .420 .268 .301 .256 B 2-26 .463 .310 .409 .247 N Ayg._ . ....r*2° .281 ...... -M * N - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at room temp. BB - inner petals definitely bluing after four days at room temp. -6- affoct ite keeping quality, Knapp (21) reported that during the winter at leaet 3 to ^ petals should be unfurled at the tine of cutting to obtain maximum keeping quality, Roses cut "tight" during this season of the year wilted and failed to open. In the same paper Knapp also stated that keeping quality was decreased and bluing Increased by prolonging the time the flowers were allowed to remain out of water after being cut and during grading opera­ tions .

Hie established practice for handling roses and other cut flowers following cutting is to subject them to a "hardening" period of several hours before shipping or otherwise offering them for sale.

During this hardening period the flowers are held at approximately

40* F. with steins in water. Iaurie and kiplinger (28) recommended a temperature of 45* F. and a relative humidity of 60 to 80 percent for this conditioning period. Knapp (21) found that a temperature of 38" F. was superior to 42* F. for hardening. The work of

Bancroft (3) indicated that roees would keep one day longer if hardened overnight rather than for only three hours.

B. Investigations on Handling and Storage Practices

It is sometimes necessary, and even desirable from the stand­ point of the grower, to hold cut flowers for rather extended periods of time before they are finally sold to the consumer. This storage period has a direct effect on the subsequent quality and longevity of the bloom. Several workers have conducted investiga­ tions In an attempt to discover or work out improved storage -87-

Appendix I. Relative Concentration of Anthocyanln Pignente and Tannine in Petals of Better Timas Roses. Plot II-9. High Nitrate, High Potassium.

Bate O.D. of Pignent Extract O.D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency

1-31 .546 .280 .387 .276 BB 2-2 • 577 .248 .293 .244 B 2-5 • 398 .233 .289 .315 B 2-7 .1+20 .284 .315 .280 BB

2-7 .432 .306 .367 .289 BB 2-9 .620 .362 .362 • 333 B 2-11 .611 .426 .342 .362 3 2-16 .509 .306 .306 .310 N

2-23 .516 .284 .284 .289 B 2-25 .342 .319 .237 .268 BB Avg. .497 __ .305 .318 .297 * h - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at room temp BB - inner petals definitely bluing after four days at room temp.

Appendix J. Relative Concentration of Anthocyanin Pigpnents and Tannins in Petals of Better Times Roses. Plot III-l. Low Nitrate, Low Potassium.

Date O.D. of Pigment Extract 0.D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Freeh Petals Old Petals 1Tendency*

1-31 .377 .297 .222 .284 N 2-4 .257 .241 .352 .306 N 2-5 .415 .289 .301 .310 h 2-6 .301 .272 .297 • 314 N

2-6 .306 .226 .284 .229 B 2-7 .362 .229 .329 .284 B 2-11 .516 .315 .347 .280 B 2-13 .450 .293 .420 .409 li

2-15 .531 .264 .409 .297 N 2-28 .404 .248 .337 .244 N .402 .267 .296 * N - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at roam tenp. BB - inner petals definitely bluing after four days at room temp. -88-

Appendix K. Relative Concentration of Anthocyanln Pi©nents and Tannine in Petals of Better Times Roses. Plot III-2 Medium Nitrate, Low Potassium.

Date 0.D. of Plraent Extract O.D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency*

1-31 .372 .252 .268 .276 N 2 -1+ .1*56 .260 .293 .319 N 2-5 • 372 .219 .260 .315 N 2-6 .310 .233 .280 .252 B

2-11 .1*38 .215 .301 .252 U 2-11 .289 .276 • 333 .301 N 2-11 .1*09 .2 M+ .21+8 .293 K 2-13 .372 •215 .333 .268 N

2-15 .502 .30 6 .352 .1+01+ B 2-22 .482 .261+ • 337 .276 li Avg. .1+10 .21+8 .300 .295 * D - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at room temp. BB - inner petals definitely bluing after four days at room temp.

Appendix L. Relative Concentration of Anthocyanln Pigments and Tannins in Petals of Better Times Roses. Plot III-3. High Nitrate, Low Potass ium.

Date O.D. of rlgraent Extract 0 .D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Freeh Petals 1Old Petals Tendency

1-31 .382 .237 .222 .276 II 2-5 .398 .21+1+ •393 • 333 li 206 .281+ .280 .333 .297 11 2-6 .293 .211 .281+ .293 li

2-7 .31+2 .256 -362 .306 B 2-7 .1+15 .237 .315 .284 if 2-8 .377 .21+8 .306 .268 N 2-9 .1+09 .259 .289 .27b II

2-11 .393 .229 .319 .2 68 II 2-11 .561 .310 .382 .310 II Avga.. J 85. .251 ^220 .291 * N - no noticeable bluing after four days at room temperature. B - incipient bluing of Inner petals after four days at room temp. BB - inner petals definitely bluing after four days at room temp. -89-

Appendix M. Relative Concentration of Anthocyanln Pigmente and Tannine in Petals of Better Times Roses. Plot III-4. Low Mitrate, Medium Potassium.

Date O.D. of Plraaent Extract O.D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency

1-31 • 396 .241 .284 .2^6 R 2-6 .3 Vf .268 .260 .276 h 2-7 .561 .382 .342 .297 li 2-7 • 531 .301 .372 .319 N

2-11 .585 .306 .280 .260 B 2-13 .404 .237 .306 .233 N 2-13 .482 .260 .367 .357 R 2-19 .561 .252 .310 .297 N

2-20 .444 .352 .362 •357 R 2-23 .489 .252 .337 .293 R Aye* .490 .28*5 .322 .294 * R - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at room temp BB - inner petals definitely bluing after four days at room temp.

Appendix M. Relative Concentration of Anthocyanln Pigments and Tannins In Petals of Better Times Roses, Plot III-5. Medium II it rate, Medium Potassium.

Date O.D. of Planent Extract O.D.. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals

2-5 .337 .219 .256 .315 R 2-5 .357 .215 .310 .347 R 2-6 .328 .208 .319 .268 R 2-6 .426 .284 •337 .297 R

2-7 .482 .264 .342 .293 B 2-7 .482 .301 .342 .324 BB 2-9 .393 .244 .293 .256 li 2-11 .469 .293 .347 .306 B

2-12 .432 .252 .293 .297 N 3-1 .420 .260 .362 .302 II Ayg. .412 .254 .320 .300 * R - no noticeable bluing after four days at room temperature. B - incipient bluing of inner petals after four days at room temp. HB - inner petals definitely bluing after four days at room temp. -90-

Appendlx 0, Relative Concentration of Anthocyanln Pifpnents and Ifennins in Petals of Better Times Roses. Plot III-6. High lilt rate, Medium Potassium.

Date O.D. of Pigment Extract O.D. of Tannin Teat Soln. Bluing Cut Fresh Petals Old Petals ]fresh Petals Old Petals Tendency*

2-5 .1*89 .306 .306 .357 B 2-6 .328 .252 .293 .276 N 2-6 .1*69 .301 .382 • 319 N 2-7 .352 .272 .272 .260 B

2-9 .1*50 .293 .357 .281* M 2-11 .495 .276 • 357 .252 B 2-11 .1*75 .272 .31*7 .289 N 2-12 .1*63 .281* .31*7 .280 a

2-12 .51*6 .319 .289 .31*7 N 2-15 .1*20 .237 .398 •333 H Av r . .1*1*9 . .280 .335 .300 * N - no noticeable bluing after four days at room temperature, B - incipient bluing of inner petals after four days at room temp, BB - inner petals definitely bluing after four days at room temp.

Appendix P. Relative Concentration of Anthocyanln Pigments and Tannins in Petals of Better Times Roses. Plot III-7. Lew Nitrate, High Potassium.

Date O.D. of Pifoaent Extract O.D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency*

2-6 • 377 .259 • :*15 .297 H 2-6 • 372 .337 • 315 .297 BB 2-12 • 1*95 .306 .393 .293 M 2-16 .1*15 .22 6 .297 • 337 M

2-19 .639 .31*7 .362 .372 B 2-27 .502 .321* .1*09 .357 B 2-27 .1*26 .226 .321* .276 B 2-29 .1*20 .259 .289 .252 B

2-29 .372 .252 .281* .260 B 3-1 .1*1*1* .21*1* .315 .328 N A .1*1*6 .278 .330 .3P3_. - * M - no noticeable bluing after four days at room temperature. B - Incipient bluing of Inner petals after four days at room temp. HB - inner petals definitely bluing after four days at room temp. -91-

Appendlx Q. Relative Concentration of Anthocyanln Pigments and Tannins In Petals of Better Times Roses. Plot III-8 . Medium Nitrate, High Potassium.

Date O.D. of Pigment Extract O.D. of Tannin Test Soln. Bluine Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency*

1-31 .51*6 .372 .272 .301 BB 2-4 .362 .362 .347 .367 BB 2-5 .409 .328 .276 .337 BB 2-6 .420 .301 .301 .268 BB

2-9 -553 .301 • 357 .289 B 2-19 .502 .289 .333 .310 N 2-2 6 • 377 .252 .256 .289 B 2-26 .415 .272 .319 .268 N

2-26 .482 .284 .328 .328 B 2-27 .463 .256 .289 .310 B Avg. _ .453 .302 .308 .307 * N - no noticeable bluing after four days at room temperature. B - Incipient bluing of inner petals after four days at room temp BB - Inner petals definitely bluing after four days at room temp.

Appendix R. Relative Concentration of Anthocyanln Pigments and Tannins In Petals of Better Times Roses. Plot III-9. High Potassium, High Nitrate.

Date O.D. of Piiassnt Extract O.D. of Tannin Test Soln. Bluing Cut Fresh Petals Old Petals Fresh Petals Old Petals Tendency

1-31 • 531 .415 .301 .280 BB 2-4 .319 .260 .293 .310 B 2-4 .523 .268 .252 .301 BB 2-9 .415 .268 .276 .268 B

2-19 .475 .357 .310 .219 B 2-20 .456 .387 .324 .293 BB 2-23 .438 .256 .387 .301 B 2-25 .463 .347 .284 .272 HB

2-29 .432 .280 .362 .301 B 3-1 .456 .367 • 333 • 377 B Avg. > 5 1 .322 ■ ■ , .302 * N - no noticeable bluing after four days at room temperature. B - Incipient bluing of inner petals after four days at room temp. BB - inner petals definitely bluing after four days at room temp. - 92-

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